antibody molecules for cd73 and uses thereof

专利摘要:
The present invention relates to antibody molecules that bind to CD73. The anti-CD73 antibody molecules can be used to treat, prevent and / or diagnose cancer.
公开号:BR112019027104A2
申请号:R112019027104-0
申请日:2018-06-21
公开日:2020-08-18
发明作者:Ansgar Brock；Rachel W. O'Connor；Michael Warren；Pamela Holland；Kulandayan Kasi Subramanian；Marie-Louise Fjaellskog；Dirksen Bussiere；Wei Shu；Mikias Woldegiorgis；Michael GLADSTONE；John Delmas Venable Iii；Viviana Cremasco；Christine Miller；Jonathan Hill；Catherine Anne Sabatos-Peyton；Glenn Dranoff；Bianka Prinz；Jerry M. Thomas；Scott Chappel；Andrew Lake；Alison Paterson
申请人:Novartis Ag；Surface Oncology, Inc.；
IPC主号:

专利说明:

[001] [001] This request claims the priority of US Serial No. 62 / 523,481 deposited on June 22, 2017 and US Serial No. 62 / 636,510 filed on February 28, 2018, the contents of which are each incorporated in this document to reference title in its entirety. SEQUENCE LISTING
[002] [002] The present application contains a Sequence Listing that was sent electronically in ASCII format and is incorporated here by way of reference in its entirety. The referred copy of ASCII, created on June 15, 2018, is called N2067-7123WO SL.txt and is 497,959 bytes in size. BACKGROUND
[003] [003] The Differentiation Group 73 (CD73), also known as ecto-5'-nucleotidase (ecto-5'NT), is a cell surface enzyme linked to glycosyl phosphatidylinositol (GPI) found in most from tissues, and particularly expressed in endothelial cells and subsets of hematopoietic cells (Resta et al., Immunol Rev161: 95 to 109 (1998) and Colgan et al., Prinergic Signal 2: 351-60 (2006)). CD73 catalyzes the conversion of adenosine monophosphate (AMP) into adenosine. Adenosine is a signaling molecule that mediates its biological effects through several receptors, including adenosine receptors A1, A2A, AZ2B and A3. The A2A receptor has received particular attention due to its wide expression in immune cells. Adenosine has pleiotropic effects on the tumor microenvironment, including expansion of regulatory T cells (Tregs), inhibition of interferon-mediated effector (Teff) responses (IFN) -y and expansion of myeloid-derived suppressor cells (MDSCs).
[004] [004] CD73 is also expressed in cancer cells, including colon, lung, pancreas, ovary, bladder, leukemia, glioma, glioblastoma, melanoma, thyroid, esophagus, prostate and breast (Jin et al., Cancer Res 70: 2,245 -55 (2010) and Stagg et al., PNAS 107: 1.547-52 (2010); Zhang et al., Cancer Res 70: 6407-11 (2010)). It was reported that the high expression of CD73 is correlated to the unsatisfactory result through several indications of cancer, such as lung cancers, melanoma, triple negative breast, scaly head and neck and colorectal. See, for example, Allard B, et al., Expert Opin Ther Targets 18: 863 to 881 (2014); Leclerc BG, et al., Clin Cancer Res 22: 158 to 166 (2016); Ren ZH, et al., Oncotarget 7: 61,690 to 61,702 (2016); Ren ZH, et al., Oncol Lett 12: 556 to 562 (2016); and Turcotte M, et al., Cancer Res 75: 4,494 to 4,503 (2015).
[005] [005] Considering the continuous need for improved strategies to target diseases such as cancer, new compositions and methods to regulate CD73 activity and related therapeutic agents are highly desired. SUMMARY
[006] [006] The antibody molecules that bind to CD73 (Differentiation Group 73) with high affinity and specificity are disclosed in this document. Also provided are nucleic acid molecules encoding the antibody molecules, expression vectors, host cells and methods for preparing the antibody molecules. Immunoconjugates, multi-specific or bispecific antibody molecules and pharmaceutical compositions that comprise the antibody molecules are also provided. The anti-CD73 antibody molecules disclosed in this document can be used (alone or in combination with other agents or mo-
[007] [007] Consequently, in certain aspects, this disclosure provides an antibody molecule (for example, an isolated or recombinant antibody molecule) that has one or more (for example, 2, 3, 4, 5, 6, 7 , 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25 or more, for example, all) of the following properties: (i) binds to CD73, for example, human CD73, with high affinity, for example, with a dissociation constant (Kp) of less than about 100 nM, for example, less than about 10 nM, 1 NM, 0.1 nM or 0.01 nM, for example, when the antibody molecule is tested as a bivalent antibody molecule using Octet; (ii) binds to soluble human CD73 or membrane-bound human CD73, or both; (iii) binds substantially to a non-human primate CD73, for example, CD73 cinomolgo, with a dissociation constant (Kp) of less than about 100 nM, for example, less than about 10 nM, 1 nM, 0.1 nM or 0.01 nM, for example, when the antibody molecule is tested as a bivalent antibody molecule using Octet; (iv) does not bind to murine CD73, for example, as determined with the use of Octet, for example, as described in Example 1 (v) inhibits or reduces the enzymatic activity of CD73 (for example, soluble human CD73 or Human CD73 linked to the
[008] [008] In one aspect, an antibody molecule that binds to a human CD73 dimer is disclosed herein, wherein said dimer consisting of a first CD73 monomer and a second CD73 monomer, where, when the antibody molecule comprises a first antigen binding domain and a second antigen binding domain, where the first antigen binding domain binds to the first CD73 monomer and the second antigen binding domain binds to the second mon-
[009] [009] In one aspect, a composition is disclosed herein comprising a plurality of antibody molecules that binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second - the CD73 monomer, where each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where , when the antibody molecules in plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at least 30%, 35% or 40% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, each of which said complex consists of an antibody molecule and a CD73 dimer, for example, when measured using size exclusion chromatography and the percentage value is obtained by determining the amount of the antibody molecules in the complex in relation to the total amount of antibody binding molecules to CD73 (excluding unbound antibody molecules), for example, as described in Example 2. In one embodiment, a document is disclosed in this document. composition comprising a plurality of antibody molecules that bind to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second CD73 monomer, each monomer comprising the amino acid sequence residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where, when the plurality antibody molecules each comprise the same first domain antigen-binding domain and the same second antigen-binding domain, at least 30%, 35% or 40% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, each of which mplex consists of an antibody molecule and a CD73 dimer, for example, when measured using a size exclusion chromatography (SEC) assay that comprises the following steps: (i) incubate a sample containing an equivalent amount of the divalent antibody molecule and the CD73 monomer overnight at 4 ° C; (li) pass the sample through an SEC column (for example, a Shodex Protein KW-803 column (8 x 300 mm | ID)) at room temperature in a buffer containing 90% 2x PBS and 10% isopropanol in volume; and (ili) analyzing SEC peaks to obtain the relative percentage values of distinguishable antibody CD73 species by determining the amount of antibody molecules in the complex in relation to the total amount of antibody binding molecules to CD73 (excluding unbound antibody molecules).
[0010] [0010] In one aspect, a composition is disclosed herein comprising a plurality of antibody molecules that binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second - the CD73 monomer, where each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where , when the antibody molecules in plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at least 70%, 75% or 80% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, each of which said complex consists of an antibody molecule and a CD73 dimer, for example, when measured using size exclusion chromatography and the percentage value is obtained by determining the amount age of the antibody molecules in the complex in relation to the total amount of antibody binding molecules to CD73 (excluding unbound antibody molecules), for example, as described in Example 2. In one embodiment, a document is disclosed in this document. composition comprising a plurality of antibody molecules that bind to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second CD73 monomer, each monomer comprising the amino acid sequence residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where, when the plurality antibody molecules each comprise the same first domain antigen-binding domain and the same second antigen-binding domain, at least 70%, 75% or 80% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, wherein each of said c The complex consists of an antibody molecule and a CD73 dimer, for example, when measured using a size exclusion chromatography (SEC) assay that comprises the following steps:
[0011] [0011] In one aspect, a plurality of an antibody molecule that binds to a human CD73 dimer is disclosed herein, wherein said dimer consists of a first CD73 monomer and a second CD73 monomer, each monomer - a number comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where, when the antibody molecules in plurality, each one, comprise the same first antigen binding domain and the same second antigen binding domain, at most 60%, 65% or 70% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, in that each of the complex comprises two or more antibody molecules and two or more dimers of CD73, for example, when measured with the use of size exclusion chromatography and the percentage value is obtained by determining the quantity of molecules those of antibody in the complex in relation to the total amount of antibody binding molecules to CD73 (excluding unbound antibody molecules), for example, as described in Example 2. In one embodiment, it is disclosed herein documents a plurality of antibody molecules that bind to a human CD73 dimer, wherein said dimer consists of a first CD73 monomer and a second CD73 monomer, each monomer comprising the amino acid sequence of the residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where, when the plurality antibody molecules each comprise the same first domain of binding antigen and the same second antigen binding domain, at most 60%, 65% or 70% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, wherein each of said complex comprises two or more ant molecules icorpo and two or more dimers of CD73, for example, when measured using a size exclusion chromatography (SEC) assay comprising the following steps: (i) incubating a sample containing an equivalent amount of bivalent antibody molecule and the CD73 monomer overnight at 4 ºC; (li) pass the sample through an SEC column (for example, a Shodex Protein KW-803 column (8 x 300 mm | D)) at room temperature in a buffer containing 90% 2x PBS and 10% isopropanol in volume; and (iii) analyzing SEC peaks to obtain the relative percentage values of distinguishable antibody CD73 species by determining the amount of antibody molecules in the complex in relation to the total amount of antibody binding molecules to CD73 (excluding unbound antibody molecules).
[0012] [0012] In one aspect, a plurality of antibody molecules that bind to a human CD73 dimer is disclosed herein, wherein said dimer consists of a first CD73 monomer and a second CD73 monomer, each monomer-
[0013] [0013] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, wherein the antibody molecule preferably binds to an open conformation, for example, a catalytically inactive conformation, of CD73 relative to tion to a closed conformation, for example, a catalytically active conformation, of CD73, for example, does not bind or bind to the closed conformation, for example, the catalytically active conformation, of CD73 with lower affinity, for example , 50%, 60%, 70%, 80%, 90%, 95%, or 99% lower affinity than when the antibody molecule binds to the open conformation, for example, the catalytically inactive conformation of CD73.
[0014] [0014] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, wherein the antibody molecule prevents or reduces the conversion of human CD73 from a catalytically inactive open conformation to a catalytically closed conformation active, for example, reduces conversion by at least 1.5 times, 2 times, 5 times, 10 times, 20 times, 30 times, 40 times, 50 times, 60 times, 70 times, 80 times, 90 times or 100 ve-
[0015] [0015] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, wherein the antibody molecule reduces the hydrogen-deuterium exchange in one or more regions of a protein comprising the sequence of amino acids from residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171) when bound to it, where one or more regions are selected from of the group consisting of residues 158 to 172, residues 206 to 215, residues 368 to 387 and residues 87 to 104 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a divalent antibody molecule with the use of hydrogen deuterium exchange mass spectrometry, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example , pH 7.5) and room temperature.
[0016] [0016] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, wherein the antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the average hydrogen-deuterium exchange in the residue (or residues) Xc selected from the core region C ( residues 368 to 387 of SEQ ID NO: 105) to a greater extent than in residue (or residues) Xa selected from core A region (residues 158 to 172 of SEQ ID NO: 105), residue (or residues) Xg selected from core B region (residues 206 to 215 of SEQ ID NO: 105) or residue (or residues) Xp selected from core D region (residues 297 to 309 of SEQ ID NO: 105), for example, when the
[0017] [0017] In one embodiment, the antibody molecule, when attached to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the average hydrogen-deuterium exchange in 20 selected residues from the core C region to a greater extent than in: (i) 14 selected residues from the core A region, (ii) 10 selected residues from the core B region, or (ili) 13 selected residues from the core D region, for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry, for example, as described in Example 2, for example, mass spectrometry of hydrogen deuterium exchange conducted for 1 minute in exchange for pH 7 to 8 (for example, pH 7.5) and room temperature. In one embodiment, the antibody binding reduces the average hydrogen-deuterium exchange in the C-region to a greater extent than in the A-region. In one embodiment, the antibody binding reduces the average hydrogen-deuterium exchange. in the C region to a greater extent than in the B region. In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in the C region to a greater extent than in the D region. In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in the C-region to a greater extent than in the A-region and the B-region. In one embodiment, the antibody-binding reduces the hydrogen-deuterium exchange average in the core C region to a greater extent than in the core A region and the core D region. In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in the core C region to a greater extent than in the region of nucleus B and region of nucleus D. In In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in the region of nucleus C to a greater extent than in the region of nucleus A, region of nucleus B and region of nucleus D.
[0018] [0018] In one embodiment, the antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the average hydrogen-deuterium exchange in 15 selected residues from the core C region to a greater extent than in: (i) 10 selected residues from the core A region, (ii) 8 selected residues from the core B region, or (iii) 10 selected residues from the core D region, for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature. In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in the C-region to a greater extent than in the A-region.
[0019] [0019] In one embodiment, the antibody molecule, when attached to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the average hydrogen-deuterium exchange in 10 selected residues from the core C region to a greater extent than in: (i) 7 selected residues from the core A region, (ii) 5 selected residues from the nucleus B region, or (iii) 7 residues selected from the nucleus D region, for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mas-Sa spectrometry, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature. In one embodiment, the antibody bond reduces the average hydrogen-deuterium exchange in the C-region region to a greater extent than in the A-region. In one embodiment, the antibody bond reduces the hydrogen exchange. nio-deuterium medium in the C region to a greater extent than in the B region. In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in the C region to a greater extent than in the nucleus region D. In one embodiment, the antibody binding reduces the average hydrogen-deuterium exchange in the C region to a greater extent than in the A region and the B region. In one embodiment, the antibody binding reduces the exchange of average hydrogen-deuterium in the region of nucleus C to a greater extent than in the region of nucleus A and region of nucleus D. In one embodiment, antibody binding reduces the exchange of average hydrogen-deuterium in the region of nucleus C a greater extent than in the region of nucleus B and region of nucleus D In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in the region of nucleus C to a greater extent than in the region of nucleus A, region of nucleus B and region of nucleus D.
[0020] [0020] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, wherein the antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the average hydrogen-deuterium exchange in residues 368 to 387 of SEQ ID NO: 105 to a greater extent than in residues 158 to 172, residues 206 to 215 or residues 297 to 309 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule with the use of mass exchange spectrometry of hydrogen deuterium, for example
[0021] [0021] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, wherein the antibody molecule, when bound to a protein that comprises the
[0022] [0022] In one embodiment, the antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the average hydrogen-deuterium exchange in 14 selected residues from the core A region to a greater extent than in: (i) 10 selected residues from the core B region, (ii) 20 selected residues from the nucleus C region, or (iii) 13 residues selected from nucleus D region, for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature. In one embodiment, the antibody bond reduces the average hydrogen-deuterium exchange in the region of nucleus A to a greater extent than in the region of nucleus B. In one embodiment, the antibody bond reduces the exchange of hydrogen. nio-deuterium medium in the region of nucleus A to a greater extent than in the region of nucleus C. In one embodiment, antibody binding reduces the exchange of hydrogen-deuterium medium in the region of nucleus A to a greater extent than in the region of nucleus D. In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in the region of nucleus A to a greater extent than in the region of nucleus B and region of nucleus C. In one embodiment, the binding of antibody reduces the exchange average hydrogen-deuterium in the region of nucleus A to a greater extent than in the region of nucleus B and region of nucleus D. In one embodiment, antibody binding reduces the exchange of average hydrogen-deuterium in the region of nucleus A a greater extent than in the core C region and the core D region In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in the region of nucleus A to a greater extent than in the region of nucleus B, region of nucleus C and region of nucleus D.
[0023] [0023] In one embodiment, the antibody molecule, when attached to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the average hydrogen-deuterium exchange in 10 selected residues from the core A region to a greater extent than in: (i) 8 selected residues from the core B region,
[0024] [0024] In one embodiment, the antibody molecule, when attached to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the average hydrogen-deuterium exchange in 7 selected residues from the core A region to a greater extent than in: (i) 5 selected residues from the core B region, (ii) 10 residues selected from nucleus C region, or (ili) 7 residues selected from nucleus D region, for example, when the antibody molecule is tested as a bivalent antibody molecule using deuterium exchange mass spectrometry hydrogen, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature. In one embodiment, the antibody bond reduces the average hydrogen-deuterium exchange in the region of nucleus A to a greater extent than in the region of nucleus B. In one embodiment, the antibody bond reduces the exchange of hydrogen. nio-deuterium medium in the region of nucleus A to a greater extent than in the region of nucleus C. In one embodiment, antibody binding reduces the exchange of hydrogen-deuterium medium in the region of nucleus A to a greater extent than in the region of nucleus D. In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in the region of nucleus A to a greater extent than in the region of nucleus B and region of nucleus C. In one embodiment, the binding of antibody reduces the exchange average hydrogen-deuterium in the region of nucleus A to a greater extent than in the region of nucleus B and region of nucleus D. In one embodiment, antibody binding reduces the exchange of average hydrogen-deuterium in the region of nucleus A a greater extent than in the region of nucleus C and region of nucleus D In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in the region of nucleus A to a greater extent than in the region of nucleus B, region of nucleus C and region of nucleus D.
[0025] [0025] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, wherein the antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the average hydrogen-deuterium exchange in residues 158 to 172 of SEQ ID NO: 105 to a greater extent than residues 206 to 215 of SEQ ID NO: 105, residues 368 to 387 of SEQ ID NO: 105 or residues 297 to 309 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody using hydrogen deuterium exchange mass spectrometry, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, PH 7.5) and room temperature.
[0026] [0026] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, wherein the antibody molecule reduces the hydrogen-deuterium exchange in one or more regions of a protein comprising the sequence of amino acids from residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171) when bound to it, where one or more regions are selected from of the group consisting of residues 158 to 172, residues 206 to 215, residues 368 to 387 and residues 87 to 104 of SEQ ID NO: 105, in which the region with the greatest reduction in the average hydrogen-deuterium exchange among one or more regions does not consist of residues 206 to 215 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry, for example example, as described in Example 2, for example, mas spectrometry hydrogen deuterium exchange sa conducted for 1 minute in exchange at pH 7 to 8 (eg pH 7.5) and room temperature.
[0027] [0027] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, wherein the antibody molecule, when bound to a protein that comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein that consists of the amino acid sequence
[0028] [0028] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, wherein the antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), leads to a reduction in the average hydrogen-deuterium exchange of less than, for example, 0.05, 0.04 , 0.03 or 0.02 Da per residue in residues 206 to 215 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule using exchange mass spectrometry of hydrogen deuterium, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature.
[0029] [0029] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, in which the antibody molecule contacts, for example, directly or indirectly, at least one, two, three or four residues within residues 158 to 172 of SEQ ID NO: 105. In one aspect, an antibody molecule that binds to human CD73, in which the antibody molecule contacts, for example, directly or indirectly, is disclosed in the present document.
[0030] [0030] In one embodiment, the antibody molecule binds to at least one, two, three or four residues within residues 158 to 172 of SEQ ID NO: 105. In one embodiment, the antibody molecule binds to at least one, two, three, four or five residues within residues 206 to 215 of SEQ ID NO: 105.
[0031] [0031] Alternatively, or in combination with a modality described in this document, the antibody molecule reduces the identification of tandem mass tag (TMT) in residue K136 (numbered according to SEQ ID NO: 105) of a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 when bound to it, for example, when the antibody molecule is tested as a bivalent antibody molecule using isotope identification of TMT, for example, using the methods described in Example 9, for example, using the TMT isotope identification conducted by an identification time of 30 s, optionally in which the antibody molecule further reduces the identification of TMT in one or more of residues K133, K162, K179, K206, K214, K285, K291 and K341 (numbered according to SEQ ID NO: 105) of the protein, for example, in one or more of residues K162, K206, K214, K285, K291 and K341 (numbered according to SEQ ID NO: 105) of the protein.
[0032] [0032] Alternatively, or in combination with a modality described in this document, the antibody molecule binds to one or more CD73 residues, for example, through an electrostatic interaction and / or a hydrogen bond, in which the one or more residues are selected from the group consisting of residues Y110, K136, L132, L157, K162, S155 and T209, numbered according to SEQ ID NO: 105, for example, as measured using analysis crystal structure, for example, using the methods described in Example 8. In one embodiment, the antibody molecule comprises a variable region of heavy chain and a variable region of light chain, in which the molecule of antibody has one or more (for example, 1, 2, 3, 4 or all) of the following properties: (i). the antibody molecule (for example, the heavy chain variable region, for example, R54 of the heavy chain variable region, numbered according to Kabat numbering) binds to Y110 (for example, the main chain carbonyl) Y110) or K136 (for example, K136 backbone carbonyl) from CD73 (nu-
[0033] [0033] In one embodiment, the antibody molecule binds to one or more regions of CD73, for example, through complementary format and / or a Van der Waal interaction, in which one or more regions are selected from the group consisting of residues 155 to 170, 136 to 138 and 209 to 210, numbered according to SEQ ID NO: 105, for example, as measured using crystal structure analysis, for example, with the use of methods described in Example 8. In one embodiment, the antibody molecule comprises a heavy chain variable region and a light chain variable region, where the antibody molecule has one or more (for example, 1, 2 or all) of the following properties: (i). the antibody molecule (for example, the heavy chain variable region, for example, residue 33, 50, 52, 56, 97, 98, 100 or 100º of the heavy chain variable region, numbered according to the numbering of Kabat) binds to residues 155 to 170 of CD73 (numbered according to SEQ ID NO: 105), for example, through format complementarity and / or a Van der Waal interaction, for example, as measured with use analysis of the crystal structure, for example, using the methods described in Example 8, (ii) the antibody molecule (for example, the
[0034] [0034] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, wherein the antibody molecule comprises at least one, two, three, four, five or six complementarity determining regions (CDRs) ) (or collectively all CDRs) of a heavy variable region and / or a light chain variable region comprising an amino acid sequence shown in Table 1 (for example, of the heavy and light chain variable region sequences of a antibody disclosed in Table 1, for example, antibody 918, 350, 356, 358, 930, 373, 374, 376, 377, 379, 363, 366, 407, 893, 939, 430, or 398), or encoded by a nucleotide sequence shown in Table 1. In some embodiments, CDRs conform to the definition of Kabat (for example, as shown in Table 1). In some modalities, CDRs are in accordance with the definition of Chothia (for example, as shown in Table 1). In some modalities, CDRs are according to the combined definition based on the definition
[0035] [0035] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, comprising a heavy chain variable region (VH) that comprises a sequence of amino acids from the complementarity determining region heavy chain 3 (VHCDR3) from GGLYGSGSYLSDFDL (SEQ ID NO: 37).
[0036] [0036] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, comprising: (i)): a heavy chain variable region (VH) comprising one, two or three among : an amino acid sequence from the heavy chain complementarity determining region 1 (VHCDR1) of X: X2AMS (SEQ ID NO: 88), where X: is R, You S, and X2 is Y or N; a VHCDR2 amino acid sequence from XIIX2GX3GXaX5TYYADSVKG (SEQ ID NO: 89), where Xx is A or S, XzéSouT, XséSouT, XéM GouS, eXséN S, LouY; an amino acid sequence of VHCDSGL GL IDS (GGL ID: 37GL) ); and / or (ii), a light chain variable region (VL) that comprises one, two or three among: a sequence of amino acids of the light chain complementarity determining region 1
[0037] [0037] In some embodiments, the anti-CD73 antibody molecule comprises: (i)) a VH comprising one, two or three of: a VHCDR1 amino acid sequence of SEQ ID NO: 38, a VHCDR amino acid sequence 2 of SEQ ID NO: 36 and a VHCDR3 amino acid sequence of SEQ ID NO: 37 (or a sequence that has at least about 85%, 90%, 95%, or 99% identity with the same sequence, and / or that has one, two, three or more substitutions, insertions or deletions, for example, conserved substitutions); and a VL comprising one, two or three of: a VLCDR1 amino acid sequence of SEQ ID NO: 48, a VLCDR2 amino acid sequence of SEQ ID NO: 49 and a VLCDR3 amino acid sequence of SEQ ID NO: 48 : 50 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with the same, and / or that has one, two, three or more substitutions, insertions or deletions, for example, preserved substitutions); (ii) a VH comprising one, two or three of: a VÓHCDR1 amino acid sequence of SEQ ID NO: 72, a VÓHCDR2 amino acid sequence of SEQ ID NO: 71 and a VÓHCDR3 amino acid sequence of SEQ ID NO: 37 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with the same, and / or that has one, two, three or more substitutions , insertions or deletions, for example, preserved substitutions); and a VL comprising one, two or three of: a VLCDR1 amino acid sequence of SEQ ID NO: 48, a VLCDR2 amino acid sequence of SEQ ID NO: 49 and a VLCDR3 amino acid sequence of SEQ ID NO: 48 : 50 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with the same, and / or that has one, two, three or more substitutions, insertions or deletions, for example, preserved substitutions);
[0038] [0038] In some embodiments, the anti-CD73 antibody molecule comprises a variable region of heavy chain that comprises the amino acid sequence of SEQ ID NO: 44, 77, 84, 142, 151 or 159, or a sequence of amino acids that have at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 44, 77, 84, 142, 151 or 159.
[0039] [0039] In some embodiments, the anti-CD73 antibody molecule comprises a light chain variable region that comprises the amino acid sequence of SEQ ID NO: 55, or an amino acid sequence that is at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 55.
[0040] [0040] In some embodiments, the anti-CD73 antibody molecule comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 46, 79, 86, 114, 116 or 117, or an amino acid sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 46, 79, 86, 114, 116 or 117.
[0041] [0041] In some embodiments, the anti-CD73 antibody molecule comprises a light chain comprising the amino acid sequence of SEQ ID NO: 57, or an amino acid sequence that is at least about 85%, 90%, 95% , or 99% sequence identity with SEQ ID NO: 57.
[0042] [0042] In some embodiments, the anti-CD73 antibody molecule comprises:
[0043] [0043] In some embodiments, the anti-CD73 antibody molecule comprises: (i): a heavy chain comprising the amino acid sequence of SEQ ID NO: 46 (or a sequence that is at least about 85%, 90%, 95%, or 99% sequence identity therewith) and a light chain comprising the amino acid sequence of SEQ ID NO: 57 (or a sequence that is at least about 85%, 90 %, 95%, or 99% sequence identity with the same); (li) a heavy chain comprising the amino acid sequence of SEQ ID NO: 114 (or a sequence having at least about 85%, 90%, 95%, or 99% sequence identity thereto) and a chain light that comprises the amino acid sequence of SEQ ID NO: 57 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity therewith); (ili))) a heavy chain comprising the amino acid sequence of SEQ ID NO: 79 (or a sequence that has at least
[0044] [0044] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein which comprises a heavy chain variable region (VH) comprising a VHCDR3 amino acid sequence of ESQESPYNNWFDP (SEQ ID
[0045] [0045] In one aspect, an antibody molecule that binds to human CD73 is disclosed in the present document, comprising: (i). a heavy chain variable region (VH) comprising an amino acid sequence of heavy chain complementarity determining region 1 (VHCDR1) from X: X2YWS (SEQ ID NO: 90), where X: is R, G or S, and X> is Y or R; a VÓHCDR2 amino acid sequence from YIYX: X2GSTX3; YNPSLKS (SEQ ID NO: 91), where X1 is G or S, Cc is R, Sou T, and Xg is N or K; and a VÓHCDR3 amino acid sequence from ESQESPYNNWFDP (SEQ ID NO: 3); and / or (li) a light chain variable region (VL) comprising an amino acid sequence of light chain complementarity determining region 1 (VLCDR1) from RASQGISSWLA (SEQ ID NO: 14); an AASSLOS VLCDR2 amino acid sequence (SEQ ID NO: 15); and a VLCDR3 amino acid sequence from QQOGNSFPRT (SEQ ID NO: 16).
[0046] [0046] In some embodiments, the anti-CD73 antibody molecule comprises: (i)) a VH comprising one, two or three of: a VHCDR1 amino acid sequence of SEQ ID NO: 61, a VHCDR amino acid sequence 2 of SEQ ID NO: 60 and a VÓMCDR3 amino acid sequence of SEQ ID NO: 3 (or a sequence that has at least about 85%, 90%, 95%, or 99% identity with the same sequence, and / or that has one, two, three or more substitutions, insertions or deletions, for example, conserved substitutions); and a VL comprising one, two or three of: a VLCDR1 amino acid sequence of SEQ ID NO: 14, a VLCDR2 amino acid sequence of SEQ ID NO: 15 and a VLCDR3 amino acid sequence of SEQ ID NO: : 16 (or a
[0047] [0047] In some embodiments, the anti-CD73 antibody molecule comprises a heavy chain variable region that comprises the amino acid sequence of SEQ ID NO: 66, 31, 10 or 168, or an amino acid sequence that has at least minus about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 66, 31, 10 or 168.
[0048] [0048] In some embodiments, the anti-CD73 antibody molecule comprises a light chain variable region that comprises the amino acid sequence of SEQ ID NO: 21, or an amino acid sequence that is at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 21.
[0049] [0049] In some embodiments, the anti-CD73 antibody molecule comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 68, 33, 12, 115, 113 or 112, or an amino acid sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 68, 33, 12, 115, 113 or 112.
[0050] [0050] In some embodiments, the anti-CD73 antibody molecule comprises a light chain comprising the amino acid sequence of SEQ ID NO: 23, or an amino acid sequence that is at least about 85%, 90%, 95% , or 99% sequence identity with SEQ ID NO: 23.
[0051] [0051] In some embodiments, the anti-CD73 antibody molecule comprises: (i)): a variable heavy chain region comprising the amino acid sequence of SEQ ID NO: 66 (or a sequence that is at least about 85%, 90%, 95%, or 99% sequence identity therewith) and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 21 (or a sequence having at least about 85 %, 90%, 95%, or 99% sequence identity therewith); (ii) a heavy chain variable region that comprises the amino acid sequence of SEQ ID NO: 31 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 21 (or a sequence which has at least about 85%, 90%, 95%, or 99% sequence identity thereto) ; (iii) a heavy chain variable region that comprises the amino acid sequence of SEQ ID NO: 10 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 21 (or a sequence which has at least about 85%, 90%, 95%, or 99% sequence identity thereto) ; or (iv) a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 168 (or a sequence that has at least about 85%, 90%, 95%, or 99% identity of sequence with the same) and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 21 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with the same).
[0052] [0052] In some embodiments, the anti-CD73 antibody molecule comprises: (i)) a heavy chain comprising the amino acid sequence of SEQ ID NO: 68 (or a sequence that is at least about 85%, 90%, 95%, or 99% sequence identity therewith) and a light chain comprising the amino acid sequence of SEQ ID NO: 23 (or a sequence which is at least about 85%, 90 %, 95%, or 99% sequence identity with the same); (li) a heavy chain comprising the amino acid sequence of SEQ ID NO: 115 (or a sequence having at least about 85%, 90%, 95%, or 99% sequence identity thereto) and a chain light comprising the amino acid sequence of SEQ ID NO: 23 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity therewith); (ili))) a heavy chain comprising the amino acid sequence of SEQ ID NO: 33 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with the same ) and a light chain comprising the amino acid sequence of SEQ ID NO: 23 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with the same bad); (iv) a heavy chain comprising the amino acid sequence of SEQ ID NO: 113 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity thereto) and a chain light comprising the amino acid sequence of SEQ ID NO: 23 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity therewith); (v) a heavy chain comprising the amino acid sequence of SEQ ID NO: 12 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity therewith) and a light chain comprising the amino acid sequence of SEQ ID NO: 23 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity to it) ; or (vi) a heavy chain comprising the amino acid sequence of SEQ ID NO: 112 (or a sequence which has at least about 85%, 90%, 95%, or 99% sequence identity thereto) and a light chain comprising the amino acid sequence of SEQ ID NO: 23 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity therewith).
[0053] [0053] In certain embodiments, the anti-CD73 antibody molecule comprises a variable region of heavy chain that has an amino acid sequence derived from a human VH3-23 germline sequence. In certain embodiments, the anti-CD73 antibody molecule comprises a light chain variable region that has an amino acid sequence derived from a human VK3-15 germline sequence. In certain embodiments, the anti-CD73 antibody molecule comprises a heavy chain variable region that has an amino acid sequence derived from a human VH3-23 germline sequence and a light chain variable region that has an amino acid sequence derived from a human VK3-15 germline sequence.
[0054] [0054] In certain embodiments, the anti-CD73 antibody molecule comprises a variable region of heavy chain that has an amino acid sequence derived from a human VH4-59 germline sequence. In certain embodiments, the anti-CD73 antibody molecule comprises a light chain variable region that has an amino acid sequence derived from a human VK1-12 germline sequence. In certain embodiments, the anti-CD73 antibody molecule comprises a heavy chain variable region that has an amino acid sequence derived from a human VH4-59 germline sequence and a light chain variable region that has an amino acid sequence derived from a human VK1-12 germline sequence.
[0055] [0055] In certain embodiments, the anti-CD73 antibody molecule comprises a heavy chain variable region that has an amino acid sequence derived from a human VH3-23 germline sequence. In certain embodiments, the anti-CD73 antibody molecule comprises a light chain variable region that has an amino acid sequence derived from a human VK1-05 germline sequence. In certain embodiments, the anti-CD73 antibody molecule comprises a heavy chain variable region that has an amino acid sequence derived from a human VH3-23 germline sequence and a light chain variable region that has an amino acid sequence derived from a human VK1-05 germline sequence.
[0056] [0056] In certain embodiments, the anti-CD73 antibody molecule comprises a variable heavy chain region that has an amino acid sequence derived from a human VH1-02 germline sequence. In certain embodiments, the anti-CD73 antibody molecule comprises a light chain variable region that has an amino acid sequence derived from a human VK1-12 germline sequence. In certain embodiments, the anti-CD73 antibody molecule comprises a heavy chain variable region that has an amino acid sequence derived from a human VH1-02 germline sequence and a light chain variable region that has an amino acid sequence derived from a human VK1-12 germline sequence.
[0057] [0057] In certain embodiments, the anti-CD73 antibody molecule comprises a variable region of heavy chain that has an amino acid sequence derived from a human VH3-07 germline sequence. In certain embodiments, the anti-CD73 antibody molecule comprises a light chain variable region that has an amino acid sequence derived from a human VK4-01 germline sequence. In certain embodiments, the anti-CD73 antibody molecule comprises a heavy chain variable region that has an amino acid sequence derived from a human VH3-07 germline sequence and a light chain variable region that has an amino acid sequence derived from a human VK4-01 germline sequence.
[0058] [0058] In certain embodiments, the anti-CD73 antibody molecule comprises a variable region of heavy chain that has an amino acid sequence derived from a human VH1-69 germline sequence. In certain embodiments, the anti-CD73 antibody molecule comprises a light chain variable region that has an amino acid sequence derived from a human VK3-15 germline sequence. In certain embodiments, the anti-CD73 antibody molecule comprises a heavy chain variable region that has an amino acid sequence derived from a human VH1-69 germline sequence and a light chain variable region that has an amino acid sequence derived from a human VK3-15 germline sequence.
[0059] [0059] In certain embodiments, the anti-CD73 antibody molecule comprises a variable region of heavy chain that has an amino acid sequence derived from a human VH4-34 germline sequence. In certain embodiments, the anti-CD73 antibody molecule comprises a light chain variable region that has an amino acid sequence derived from a human VK1-12 germline sequence. In certain embodiments, the anti-CD73 antibody molecule comprises a heavy chain variable region that has an amino acid sequence derived from a human VH4-34 germline sequence and a light chain variable region that has an amino acid sequence derived from a human VK1-12 germline sequence.
[0060] [0060] In certain embodiments, the anti-CD73 antibody molecule is a monoclonal antibody or an antibody with unique specificity. In certain embodiments, the anti-CD73 antibody molecule is a multispecific or bispecific antibody. The heavy and light chains of the anti-CD73 antibody molecule can be full length (for example, an antibody can include at least one or at least two complete heavy chains, and at least one or at least two complete light chains) or include an antigen binding fragment (for example, a Fab, F (ab ') 2, Fv, a single chain Fv fragment, a single domain antibody, a diabody (dAb), a divalent or bispecific antibody or fragment thereof, a single domain variant thereof or a camelid antibody).
[0061] [0061] In certain embodiments, the anti-CD73 antibody molecule has a heavy chain constant region (Fc) chosen, for example, from the heavy chain constant regions of I9G1, IgG2, I9G3, I9G4, IgM, IgA1, IgA2, 1gD and IgE; particularly, chosen, for example, from the IgG1, IgG2, IgG3 and IgG4 heavy chain constant regions, more particularly, the IgG4 heavy chain constant region. In some modalities, the heavy chain constant region is human IgG4. In some embodiments, the anti-CD73 antibody molecule has a light chain constant region chosen, for example, from the kappa or lambda light chain constant regions. In some embodiments, the light chain constant region is kappa (for example, human kappa). In some modalities, the constant region is altered, for example, mutated, to modify the properties of the anti-CD73 antibody molecule (for example, to increase or decrease one or more of: Fc receptor binding, glycosylation of antibody, the number of cysteine residues, effector cell function or complement function). In some embodiments, the heavy chain constant region of an IgG4, for example, a human IgG4, is mutated at position 228 according to Eu numbering (for example, S to P), for example, as shown in Table 3. In certain embodiments, the anti-CD73 antibody molecules comprise a human IgG4 mutated at position 228 according to the numbering of Eu (for example, S to P), for example, as shown in Table 3; and a kappa light chain constant region, for example, as shown in Table 3. In some modalities, the heavy chain constant region of an IgG4, for example, a human I9gG4, is mutated at position 228 ( for example, S to P) and position 235 (for example, L to E) according to the numbering of Eu, for example, as shown in Table 3. In certain modalities,
[0062] [0062] In some embodiments, the anti-CD73 antibody molecule comprises a heavy chain constant region selected from IgG1, IgG2, IgG3 and IgG4 and a light chain constant region chosen from constant regions of kappa or lambda light chain. In some embodiments, the anti-CD73 antibody molecule comprises a heavy chain constant region that comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 92 to 103, 119 and 120 and a constant region of light chain comprising the amino acid sequence of SEQ ID NO: 104.
[0063] [0063] In one aspect, the invention features an antibody molecule that competes with a monoclonal antibody, for example, an antibody molecule described in this document, for binding to human CD73. The invention also features an antibody molecule that binds to the same (or substantially the same) epitope or an overlapping (or substantially overlapping) epitope as a monoclonal antibody, for example, an antibody molecule described herein, the human CD73 .
[0064] [0064] In one embodiment, the monoclonal antibody comprises a heavy chain variable region (VH) comprising an amino acid sequence of heavy chain complementarity determining region 3 (VHCDR3) of GGLYGSGSYLSDFDL (SEQ ID NO: 37).
[0065] [0065] In certain embodiments, the monoclonal antibody comprises: (i): a heavy chain variable region (VH) comprising an amino acid sequence of X heavy chain complementarity determining region (VHCDR1) ;: X2AMS (SEQ ID NO: 88), where Xx is R, Y or S, and Xo is Y or N; a VHCDR2 amino acid sequence of XIX2GX3; GXaXsTYYADSVKG (SEQ ID NO: 89), where X is Aou S, Xoé Sou T, Xgé Sou T, 4 É M, GouS, and Xs is N, S, Lou Y; and a VHCDR3 amino acid sequence from GGLYGSGSYLSDFDL (SEQ ID NO: 37); and / or (i))) a light chain variable region (VL) comprising an amino acid sequence of RASQSVGSNLA light chain 1 complementarity determining region (VLCDR1) (SEQ ID NO: 48); a VLCDR2 amino acid sequence from GASTRAT (SEQ ID NO: 49); and a VLCDR3 amino acid sequence from QQHNAFPYT (SEQ ID NO: 50).
[0066] [0066] In certain embodiments, the monoclonal antibody comprises: (i)) a VH that comprises a VÓHCDR1 amino acid sequence of SEQ ID NO: 38, a VHCDR2 amino acid sequence of SEQ ID NO: 36 and a VHCDR3 amino acid sequence of SEQ ID NO: 37; and a VL that comprises a sequence
[0067] [0067] In one embodiment, the monoclonal antibody comprises a heavy chain variable region (VH) comprising an amino acid sequence of ESQESPYNNWFDP heavy chain complementarity determining region 3 (VHCDR3) (SEQ ID NO: 3).
[0068] [0068] In certain embodiments, the monoclonal antibody comprises: (i). a heavy chain variable region (VH) comprising an amino acid sequence of heavy chain complementarity determining region 1 (VHCDR1) from X: X2YWS (SEQ ID NO: 90), where X: is R, G or S, and Xo is Y or R; a VHCDR2 amino acid sequence from YIYX: X2GSTX; YNPSLKS (SEQ ID NO: 91), where X1 is G or S, Cc is R, Sou T, and Xg is N or K; and a VHCDR3 amino acid sequence from ESQESPYNNWFDP (SEQ | D NO: 3); and / or (ii), a light chain variable region (VL) comprising an amino acid sequence of RASQGISSWLA light chain 1 complementarity determining region (VLCDR1) (SEQ ID NO: 14); an AASSLOS VLCDR2 amino acid sequence (SEQ ID NO: 15); and a VLCDR3 amino acid sequence from QQGNSFPRT (SEQ ID NO: 16).
[0069] [0069] In certain embodiments, the monoclonal antibody comprises
[0070] [0070] In one aspect, the invention features a nucleic acid molecule comprising one or both sequences of nucleotides that encode variable regions of heavy and light chain, CDRs, hypervariable loops, framework regions of anti-antibody molecules CD73, as described in this document. In certain embodiments, the nucleotide sequence encoding the anti-CD73 antibody molecule is codon-optimized. For example, the invention features a first and second variable regions of heavy and light chain encoding nucleic acid, respectively, of an anti-CD73 antibody molecule chosen, for example, from any one of 918, 350, 356, 358, 930, 373, 374, 376, 377, 379, 363, 366, 407, 893, 939, 430 or 398, as summarized in Table 1, or a sequence substantially identical thereto. For example, the nucleic acid may comprise a sequence of nucleotides as shown in Table 1, or a sequence substantially identical to the same (for example, a sequence that is at least about 85%, 90%, 95%, or 99 % sequence identity with the same, or that differs by no more than 3, 6, 15, 30 or 45 nucleotides from the sequences shown in Table 1).
[0071] [0071] In one aspect, nucleic acids comprising nucleotide sequences encoding variable regions of heavy and light chain and CDRs of anti-CD73 antibody molecules, as described herein, are disclosed. For example, the disclosure provides a first and second variable regions of heavy and light chain encoding nucleic acid, respectively, of an anti-CD73 antibody molecule according to Table 1 or a sequence substantially identical thereto. For example, the nucleic acid can comprise a nucleotide sequence that encodes an anti-CD73 antibody molecule according to Table 1, or a sequence substantially identical to such a nucleotide sequence (for example, a sequence that has at least about
[0072] [0072] In certain embodiments, the nucleic acid may comprise a nucleotide sequence that encodes at least one, two or three CDRs, or hypervariable loops, of a variable region of heavy chain that has an amino acid sequence as shown. - seated in Table 1, or a sequence substantially homologous to the same (for example, a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity to the same and / or that has one, two, three or more substitutions, insertions or deletions, for example, preserved substitutions).
[0073] [0073] In certain embodiments, the nucleic acid can comprise a sequence of nucleotides that encodes at least one, two or three CDRs, or hypervariable loops, of a variable region of light chain that has an amino acid sequence as it presents - as shown in Table 1, or a sequence substantially homologous to the same (for example, a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity to the same and / or that has one, two, three or more substitutions, insertions or deletions, for example, preserved substitutions).
[0074] [0074] In certain embodiments, the nucleic acid may comprise a sequence of nucleotides that encodes at least one, two, three, four, five or six CDRs or hypervariable loops, from variable regions of heavy and light chain that have an amino acid sequence as shown in Table 1, or a sequence substantially homologous to it (for example, a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with the same and / or one, two, three or more substitutions, insertions or deletions, for example, conserved substitutions).
[0075] [0075] In certain embodiments, the nucleic acid encodes a variable region of heavy chain, where the nucleic acid comprises the nucleotide sequence of SEQ ID NO: 45, 78, 85, 143, 152, 160, 67, 32 , 11 or 169, or a nucleotide sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 45, 78, 85, 143, 152, 160, 67, 32, 11 or 169.
[0076] [0076] In certain embodiments, the nucleic acid encodes a heavy chain, wherein the nucleic acid comprises the nucleotide sequence of SEQ ID NO: 47, 80, 87, 69, 34 or 13, or a sequence of nucleotides that have at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 47, 80, 87, 69, 34 or
[0077] [0077] In certain embodiments, the nucleic acid encodes a variable region of light chain, where the nucleic acid comprises the nucleotide sequence of SEQ ID NO: 56, 144, 22 or 170, or a sequence of nucleotides which has at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 56, 144, 22 or 170.
[0078] [0078] In certain embodiments, the nucleic acid encodes a light chain, wherein the nucleic acid comprises the nucleotide sequence of SEQ ID NO: 58 or 24, or a nucleotide sequence that is at least about 85 %, 90%, 95%, or 99% sequence identity with SEQ ID NO: 58 or 24.
[0079] [0079] In one aspect, this disclosure features host cells and vectors containing the nucleic acids described in this document. Nucleic acids can be present in a single vector or separate vectors present in the same host cell or separate host cells. Also provided in the present document is a method for producing an anti-C73 antibody molecule, the method comprising cultivating a disseminated host cell
[0080] [0080] In one aspect, the present disclosure provides a method for providing an antibody molecule described in the present document. The method may include: providing a CD73 antigen (for example, an antigen comprising at least a portion of a CD73 epitope, for example, the N-terminal domain of a CD73 antigen); that obtains an antibody molecule that binds to the CD73 antigen; and assessing whether the antibody molecule binds to the CD73 antigen, or assessing the effectiveness of the antibody molecule in modulation, for example, stimulating or inhibiting CD73 activity. The method may additionally include administering the antibody molecule to an individual, for example, a human or non-human animal.
[0081] [0081] In one aspect, the disclosure provides compositions, for example, pharmaceutical compositions, which include a pharmaceutically acceptable stabilizer, excipient or carrier, and at least one of the anti-CD73 antibody molecules described in the present document. In one embodiment, the composition, for example, the pharmaceutical composition, includes a combination of the antibody molecule and one or more agents, for example, a therapeutic agent or another antibody molecule, as described herein. In some embodiments, the antibody molecule is conjugated to an identification or a therapeutic agent. In some embodiments, the compositions, for example, pharmaceutical compositions, comprise a combination of the antibody molecule and a second agent, for example, a therapeutic agent or two or more of the aforementioned antibody molecules, as further described herein.
[0082] [0082] The anti-CD73 antibody molecules disclosed in this document may inhibit, reduce or neutralize one or more activities of CD73, for example, inhibit or reduce the enzymatic activity of
[0083] [0083] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (i) the antibody molecule binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second CD73 monomer (for example, each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, for example, each monomer consists of the amino acid sequence of SEQ ID NO : 171), where, when the antibody molecule comprises a first antigen binding domain and a second antigen binding domain, where the first antigen binding domain binds to the first CD73 monomer and the second antigen binding binds to the second CD73 monomer, for example, when tested using size exclusion chromatography; and (ii) the antibody molecule, when linked to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the average hydrogen-deuterium exchange in residues 368 to 387 of SEQ ID NO: 105 to a greater extent than in residues 158 to 172, residues 206 to 215 or residues 297 to 309 of SEQ ID NO: 105 , for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry, for example, as described in Example 2, for example, exchange mass spectrometry of hydrogen deuterium conducted for 1 minute in exchange at pH 7 to 8 (eg pH 7.5) and room temperature.
[0084] [0084] In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in residues 368 to 387 of SEQ ID NO: 105 to a greater extent than in residues 158 to 172 of SEQ ID NO: 105. In one embodiment , antibody binding reduces the average hydrogen-deuterium exchange in residues 368 to 387 of SEQ ID NO: 105 to a greater extent than in residues 206 to 215 of SEQ ID NO:
[0085] [0085] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (i) the antibody molecule binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second CD73 monomer (for example, each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, for example, each monomer consists of the amino acid sequence of SEQ ID NO : 171), where, when the antibody molecule comprises a first antigen binding domain and a second antigen binding domain, where the first antigen binding domain binds to the first CD73 monomer and the second antigen binding binds to the second CD73 monomer, for example, when tested using size exclusion chromatography; and (ii) the antibody molecule, when linked to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the average hydrogen-deuterium exchange in residues 158 to 172 of SEQ ID NO: 105 to a greater extent than in residues 206 to 215 of SEQ ID NO: 105, residues 368 to 387 of SEQ ID NO: 105 or residues 297 to 309 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a divalent antibody molecule using hydrogen deuterium exchange mass spectrometry, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature.
[0086] [0086] In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in residues 158 to 172 of SEQ ID NO: 105 to a greater extent than in residues 206 to 215 of SEQ ID NO:
[0087] [0087] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (i) the antibody molecule binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second CD73 monomer (for example, each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, for example, each monomer consists of the amino acid sequence of SEQ ID NO : 171), where, when the antibody molecule comprises a first antigen binding domain and a second antigen binding domain, where the first antigen binding domain binds to the first CD73 monomer and the second antigen binding binds to the second CD73 monomer, for example, when tested using size exclusion chromatography; and (ii) the antibody molecule reduces hydrogen-deuterium exchange in one or more regions of a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the sequence amino acids of SEQ ID NO: 171) when linked to it, in which one or more regions are selected from the group consisting of residues 158 to 172, residues 206 to 215, residues 368 to 387 and residues 87 to 104 of SEQ ID NO: 105, in which the region that has the greatest reduction in the average hydrogen-deuterium exchange among one or more regions does not consist of residues 206 to 215 of SEQ ID NO: 105, for example , when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry, for example, as described in Example 2, for example, conducted hydrogen deuterium exchange mass spectrometry for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and time environment rature.
[0088] [0088] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, in which: (i) the antibody molecule binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second CD73 monomer (for example, each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, for example, each monomer consists of the amino acid sequence of SEQ ID NO : 171), where, when the antibody molecule comprises a first antigen binding domain and a second antigen binding domain, where the first antigen binding domain binds to the first CD73 monomer and the second antigen binding binds to the second CD73 monomer, for example, when tested using size exclusion chromatography; and (ii) the antibody molecule, when linked to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), leads to a reduction in the average hydrogen-deuterium exchange of more than, for example, 0.02, 0.03, 0.04, 0.05 or 0.06 Da per residue in residues 368 to 387 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry, for example, as described in Example 2, for example , hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature.
[0089] [0089] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (i) the antibody molecule binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second CD73 monomer (for example, each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, for example, each monomer consists of the amino acid sequence of SEQ ID NO : 171), where, when the antibody molecule comprises a first antigen binding domain and a second antigen binding domain, where the first antigen binding domain binds to the first CD73 monomer and the second antigen binding binds to the second CD73 monomer, for example, when tested using size exclusion chromatography; and (ii) the antibody molecule, when linked to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), leads to a reduction in the average hydrogen-deuterium exchange of less than, for example, 0.05, 0.04, 0.03 or 0.02 Da per residue in residues 206 to 215 of SEQ ID NO: 105, example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry, for example, as described in Example 2, for example, mass mass spectrometry hydrogen deuterium exchange conducted for 1 minute in exchange at pH 7 to 8 (eg pH 7.5) and room temperature.
[0090] [0090] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (i) the antibody molecule binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second CD73 monomer (for example, each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, for example, each monomer consists of the amino acid sequence of SEQ ID NO : 171), where, when the antibody molecule comprises a first antigen binding domain and a second antigen binding domain, where the first antigen binding domain binds to the first CD73 monomer and the second antigen binding binds to the second CD73 monomer, for example, when tested using size exclusion chromatography; and (ii) the antibody molecule binds to at least one, two, three or four residues within residues 158 to 172 of SEQ ID NO: 105, and / or at least one, two, three, four or five residues within from residues 206 to 215 of SEQ ID NO: 105.
[0091] [0091] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (i) the antibody molecule binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second CD73 monomer (for example, each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, for example, each monomer consists of the amino acid sequence of SEQ ID NO : 171), where, when the antibody molecule comprises a first antigen binding domain and a second antigen binding domain, where the first antigen binding domain binds to the first CD73 monomer and the second antigen binding binds to the second CD73 monomer, for example, when tested using size exclusion chromatography; and (ii) the antibody molecule inhibits at least about 60%, 70%, 80%, or 90% of the enzyme activity of human membrane-bound CD73, for example, when the antibody molecule is tested as an antibody molecule bivalent with the use of a modified Cell Titration Glo (CTG) assay, for example, as described in Example 1.
[0092] [0092] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (i) the antibody molecule binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second CD73 monomer (for example, each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, for example, each monomer consists of the amino acid sequence of SEQ ID NO : 171), where, when the antibody molecule comprises a first antigen binding domain and a second antigen binding domain, where the first antigen binding domain binds to the first CD73 monomer and the second antigen binding binds to the second CD73 monomer, for example, when tested using size exclusion chromatography; and (ii) the antibody molecule comprises a heavy chain variable region (VH) comprising an amino acid sequence of heavy chain complementarity determining region 3 (VHCDR3) from GGLYGSGSYLSDFDL (SEQ ID NO: 37) .
[0093] [0093] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, in which: (i) the antibody molecule binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second CD73 monomer (for example, each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, for example, each monomer consists of the amino acid sequence of SEQ ID NO : 171), where, when the antibody molecule comprises a first antigen binding domain and a second antigen binding domain, where the first antigen binding domain binds to the first CD73 monomer and the second antigen binding binds to the second CD73 monomer, for example, when tested using size exclusion chromatography; and (ii) the antibody molecule comprises: (a) a heavy chain variable region (VH) comprising an amino acid sequence of X heavy chain complementarity determining region (VHCDR1); X2AMS (SEQ ID NO: 88), where Xx is R, Y or S, and X2 is Y or N; a VHCDR2 amino acid sequence of XIX2GX3GXaXsTYYADSVKG (SEQ ID NO: 89), where X1 is Aou S, Xoé Sou T, Xgé Sou T, 4 É M, GouS,
[0094] [0094] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (1) the antibody molecule binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second CD73 monomer (for example, each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, for example, each monomer consists of the amino acid sequence of SEQ ID NO: 171 ), where, when the antibody molecule comprises a first antigen binding domain and a second antigen binding domain, where the first antigen binding domain binds to the first CD73 monomer and the second CD73 binding domain. antigen binds to the second CD73 monomer, for example, when tested using size exclusion chromatography; and (ii) the antibody molecule comprises: (a) a VH comprising a VÓHCDR1 amino acid sequence of SEQ ID NO: 38, a VÓHCDR2 amino acid sequence of SEQ ID NO: 36 and a sequence of VHCDR3 amino acids of SEQ ID NO: 37; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 48, a VLCDR2 amino acid sequence of SEQ ID NO: 49 and a VLCDR3 amino acid sequence of SEQ ID NO: 50;
[0095] [0095] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (i) the antibody molecule binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second CD73 monomer (for example, each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, for example, each monomer consists of the amino acid sequence of SEQ ID NO : 171), where, when the antibody molecule comprises a first antigen binding domain and a second antigen binding domain, where the first antigen binding domain binds to the first CD73 monomer and the second antigen binding binds to the second CD73 monomer, for example, when tested using size exclusion chromatography; and (ii) the antibody molecule comprises a heavy chain variable region (VH) comprising an amino acid sequence of heavy chain complementarity determining region 3 (VHCDR3) from ESQESPYNNWFDP (SEQ | D NO: 3 ).
[0096] [0096] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (i) the antibody molecule binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second CD73 monomer (for example, each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, for example, each monomer consists of the amino acid sequence of SEQ ID NO : 171), where, when the antibody molecule comprises a first antigen binding domain and a second antigen binding domain, where the first antigen binding domain binds to the first CD73 monomer and the second antigen binding binds to the second CD73 monomer, for example, when tested using size exclusion chromatography; and (ii) the antibody molecule comprises: (a) a heavy chain variable region (VH) comprising an amino acid sequence of heavy chain complementarity determining region 1 (VHCDR1) from X: X2YWS (SEQ ID NO: 90), where X: is R, G or S, and X> is Y or R; a VÓHOCDR2 amino acid sequence from YIYX: X2GSTX; YNPSLKS (SEQ ID NO: 91), where X1 is G or S, Cc is R, Sou T, and Xg is N or K; and a VÓHCDR3 amino acid sequence from ESQESPYNNWFDP (SEQ ID NO: 3) and (b) a light chain variable region (VL) comprising an amino acid sequence from the light chain complementarity determining region 1 ( VLCDR1) from RASQGISSWLA (SEQ ID NO: 14); an AASSLOS VLCDR2 amino acid sequence (SEQ ID NO: 15); and a VLCDR3 amino acid sequence from QOGNSFPRT (SEQ ID NO: 16).
[0097] [0097] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (i) the antibody molecule binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second CD73 monomer (for example, each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, for example, each monomer consists of the amino acid sequence of SEQ ID NO : 171), where, when the antibody molecule comprises a first antigen binding domain and a second antigen binding domain, where the first antigen binding domain binds to the first CD73 monomer and the second antigen binding binds to the second CD73 monomer, for example, when tested using size exclusion chromatography; and
[0098] [0098] In one aspect, a composition is disclosed herein comprising a plurality of antibody molecules that binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second - the CD73 monomer, where each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where : (i) when the antibody molecules in plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at least 30%, 35%, 40%, 45%, 50% , 55%, 60%, 65%, 70%, 75%, or 80% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, each of which complex consists of an antibody molecule and a CD73 dimer, for example, when measured using size exclusion chromatography and the value Percentage is obtained by determining the amount of antibody molecules in the complex in relation to the total amount of antibody binding molecules to CD73 (excluding unbound antibody molecules), for example, as described in the Example 2; and (ii) the antibody molecule, when linked to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the exchange of
[0099] [0099] In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in residues 368 to 387 of SEQ ID NO: 105 to a greater extent than in residues 158 to 172 of SEQ ID NO:
[00100] [00100] In one aspect, a composition is disclosed herein comprising a plurality of antibody molecules that binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second - the CD73 monomer, where each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where : (i) when the antibody molecules in plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at least 30%, 35%, 40%, 45%, 50% , 55%, 60%, 65%, 70%, 75%, or 80% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, each of which complex consists of an antibody molecule and a CD73 dimer, for example, when measured using size exclusion chromatography and the value Percentage is obtained by determining the amount of antibody molecules in the complex in relation to the total amount of antibody binding molecules to CD73 (excluding unbound antibody molecules), for example, as described in the Example 2; and (ii) the antibody molecule, when linked to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the average hydrogen-deuterium exchange in residues 158 to 172 of SEQ ID NO: 105 to a greater extent than in residues 206 to 215 of SEQ ID NO: 105,
[00101] [00101] In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in residues 158 to 172 of SEQ ID NO: 105 to a greater extent than in residues 206 to 215 of SEQ ID NO:
[00102] [00102] In one aspect, a composition is disclosed herein comprising a plurality of antibody molecules that binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second - the CD73 monomer, where each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where : (i) when the antibody molecules in plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at least 30%, 35%, 40%, 45%, 50% , 55%, 60%, 65%, 70%, 75%, or 80% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, each of which complex consists of an antibody molecule and a CD73 dimer, for example, when measured using size exclusion chromatography and the value Percentage is obtained by determining the amount of antibody molecules in the complex in relation to the total amount of antibody binding molecules to CD73 (excluding unbound antibody molecules), for example, as described in the Example 2; and (ii) the antibody molecule reduces hydrogen-deuterium exchange in one or more regions of a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the sequence amino acids of SEQ ID NO: 171) when linked to it, in which one or more regions are selected from the group consisting of residues 158 to 172, residues 206 to 215, residues 368 to 387 and residues 87 to 104 of SEQ ID NO: 105, in which the region that has the greatest reduction in the average hydrogen-deuterium exchange among one or more regions does not consist of residues 206 to 215 of SEQ ID NO: 105, for example , when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry, for example, as described in Example 2, for example, conducted hydrogen deuterium exchange mass spectrometry for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and time environment rature.
[00103] [00103] In one aspect, a composition is disclosed herein comprising a plurality of antibody molecules that binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second - the CD73 monomer, where each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where : (i) when the antibody molecules in plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at least 30%, 35%, 40%, 45%, 50% , 55%, 60%, 65%, 70%, 75%, or 80% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, each of which complex consists of an antibody molecule and a CD73 dimer, for example, when measured using size exclusion chromatography and the value Percentage is obtained by determining the amount of antibody molecules in the complex in relation to the total amount of antibody binding molecules to CD73 (excluding unbound antibody molecules), for example, as described in the Example 2; and (ii) the antibody molecule, when linked to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), leads to a reduction in the average hydrogen-deuterium exchange of more than, for example, 0.02, 0.03, 0.04, 0.05 or 0.06 Da per residue in residues 368 to 387 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry, for example, as described in Example 2, for example , hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature.
[00104] [00104] In one aspect, a composition is disclosed herein comprising a plurality of antibody molecules that binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second - the CD73 monomer, where each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where : (i) when the antibody molecules in plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at least 30%, 35%, 40%, 45%, 50% , 55%, 60%, 65%, 70%, 75%, or 80% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, each of which complex consists of an antibody molecule and a CD73 dimer, for example, when measured using size exclusion chromatography and the value percentage is obtained by determining the amount of antibody molecules in the complex in relation to the total amount of antibody binding molecules to CD73 (excluding
[00105] [00105] In one aspect, a composition is disclosed herein comprising a plurality of antibody molecules that binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second - the CD73 monomer, where each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where : (i) when the antibody molecules in plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at least 30%, 35%, 40%, 45%, 50% , 55%, 60%, 65%, 70%, 75%, or 80% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, each of which complex consists of an antibody molecule and a CD73 dimer, for example, when measured using size exclusion chromatography and the value Percentage is obtained by determining the amount of antibody molecules in the complex in relation to the total amount of antibody binding molecules to CD73 (excluding unbound antibody molecules), for example, as described in the Example 2; and (ii) the antibody molecule binds to at least one, two, three or four residues within residues 158 to 172 of SEQ ID NO: 105, and / or at least one, two, three, four or five residues within from residues 206 to 215 of SEQ ID NO: 105.
[00106] [00106] In one aspect, a composition is disclosed herein comprising a plurality of antibody molecules that binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second - the CD73 monomer, where each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where : (i) when the antibody molecules in plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at least 30%, 35%, 40%, 45%, 50% , 55%, 60%, 65%, 70%, 75%, or 80% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, each of which complex consists of an antibody molecule and a CD73 dimer, for example, when measured using size exclusion chromatography and the value Percentage is obtained by determining the amount of antibody molecules in the complex in relation to the total amount of antibody binding molecules to CD73 (excluding unbound antibody molecules), for example, as described in the Example 2; and
[00107] [00107] In one aspect, a composition is disclosed herein comprising a plurality of antibody molecules that binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second - the CD73 monomer, where each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where : (i) when the antibody molecules in plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at least 30%, 35%, 40%, 45%, 50% , 55%, 60%, 65%, 70%, 75%, or 80% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, each of which complex consists of an antibody molecule and a CD73 dimer, for example, when measured using size exclusion chromatography and the value Percentage is obtained by determining the amount of antibody molecules in the complex in relation to the total amount of antibody binding molecules to CD73 (excluding unbound antibody molecules), for example, as described in the Example 2; and (ii) the antibody molecule comprises a heavy chain variable region (VH) comprising an amino acid sequence of heavy chain complementarity determining region 3 (VHCDR3) from GGLYGSGSYLSDFDL (SEQ ID NO: 37) .
[00108] [00108] In one aspect, a composition is disclosed herein comprising a plurality of antibody molecules that binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second - the CD73 monomer, where each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where : (i) the antibody molecules in the plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at least 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, or 80% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, each of which complex consists of a antibody molecule and a CD73 dimer, for example, when measured using size exclusion chromatography and the value of per percentage is obtained by determining the amount of antibody molecules in the complex in relation to the total amount of antibody binding molecules to CD73 (excluding unbound antibody molecules), for example, as described in Example 2 ; and (ii) the antibody molecule comprises: (a) a heavy chain variable region (VH) comprising an amino acid sequence of the heavy chain complementarity determining region 1 (VHCDR1) of X; X2AMS (SEQ ID NO: 88), where Xx is R, Y or S, and Xo is Y or N; a VHCDR2 amino acid sequence of XIX2GX; 3GXaXsTYYADSVKG (SEQ ID NO: 89), where X1 is Aou S, Xoé S or T, Xgé Sou T, x É M, GouS, and Xs is N, S, Lou Y; and a VHCDR3 amino acid sequence of
[00109] [00109] In one aspect, a composition is disclosed herein comprising a plurality of antibody molecules that binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second - the CD73 monomer, where each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where : (i) when the antibody molecules in plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at least 30%, 35%, 40%, 45%, 50% , 55%, 60%, 65%, 70%, 75%, or 80% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, each of which complex consists of an antibody molecule and a CD73 dimer, for example, when measured using size exclusion chromatography and the value Percentage is obtained by determining the amount of antibody molecules in the complex in relation to the total amount of antibody binding molecules to CD73 (excluding unbound antibody molecules), for example, as described in the Example 2; and (ii) the antibody molecule comprises: (a) a VH comprising an amino acid sequence
[00110] [00110] In one aspect, a composition is disclosed herein comprising a plurality of antibody molecules that binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second - the CD73 monomer, where each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where : (i) when the antibody molecules in plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at least 30%, 35%, 40%, 45%, 50% , 55%, 60%, 65%, 70%, 75%, or 80% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, each of which complex consists of an antibody molecule and a CD73 dimer, for example, when measured using size exclusion chromatography and the value percentage is obtained by determining the amount of antibody molecules in the complex in relation to the total amount of antibody binding molecules to CD73 (excluding
[00111] [00111] In one aspect, a composition is disclosed herein comprising a plurality of antibody molecules that binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second - the CD73 monomer, where each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where : (i) when the antibody molecules in plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at least 30%, 35%, 40%, 45%, 50% , 55%, 60%, 65%, 70%, 75%, or 80% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, each of which complex consists of an antibody molecule and a CD73 dimer, for example, when measured using size exclusion chromatography and the value Percentage is obtained by determining the amount of antibody molecules in the complex in relation to the total amount of antibody binding molecules to CD73 (excluding unbound antibody molecules), for example, as described in the Example 2; and (ii) the antibody molecule comprises: (a) a heavy chain variable region (VH) comprising an amino acid sequence of heavy chain complementarity determining region 1 (VHCDR1) from X: X2YWS (SEQ ID NO: 90), where X: is R, G or S, and X> is Y or R; a VÓHCDR2 amino acid sequence from YIYX: X2GSTX:; YNPSLKS (SEQ ID NO: 91), where X: is G or S, Cc is R, S or T, and Xg is Nou K; and a VÓHCDR3 amino acid sequence of ESQESPYNNWFDP (SEQ | D NO: 3) and (b) a light chain variable region (VL) that comprises a sequence of amino acids from the light chain complementarity determining region 1 (VLCDR1) from RASQGISSWLA (SEQ ID NO: 14); an AASSLOS VLCDR2 amino acid sequence (SEQ ID NO: 15); and a VLCDR3 amino acid sequence from QQOGNSFPRT (SEQ ID NO: 16).
[00112] [00112] In one aspect, a composition is disclosed herein comprising a plurality of antibody molecules that binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second - the CD73 monomer, where each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where : (i) when the antibody molecules in plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at least 30%, 35%, 40%, 45%, 50% , 55%, 60%, 65%, 70%, 75%, or 80% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, each of which complex consists of an antibody molecule and a CD73 dimer, for example, when measured using size exclusion chromatography and the value percentage is obtained by determining the amount of antibody molecules in the complex in relation to the amount
[00113] [00113] In one aspect, a composition is disclosed herein comprising a plurality of antibody molecules that binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second - the CD73 monomer, where each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where : (i) when the antibody molecules in plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at most 20%, 25%, 30%, 35%, 40% , 45%, 50%, 55%, 60%, 65%, or 70% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, each of which complex comprises two or more antibody molecules and two or more dimers of CD73, for example, when measured using exclusion chromatography in size and percentage value is obtained by determining the amount of antibody molecules in the complex in relation to the total amount of antibody binding molecules to CD73 (excluding unbound antibody molecules), for example , as described in Example 2; and (ii) the antibody molecule, when linked to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the average hydrogen-deuterium exchange in residues 368 to 387 of SEQ ID NO: 105 to a greater extent than in residues 158 to 172, residues 206 to 215 or residues 297 to 309 of SEQ ID NO: 105 , for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry, for example, as described in Example 2, for example, exchange mass spectrometry of hydrogen deuterium conducted for 1 minute in exchange at pH 7 to 8 (eg pH 7.5) and room temperature.
[00114] [00114] In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in residues 368 to 387 of SEQ ID NO: 105 to a greater extent than in residues 158 to 172 of SEQ ID NO:
[00115] [00115] In one aspect, a composition is disclosed herein comprising a plurality of antibody molecules that binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second - the CD73 monomer, where each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where :
[00116] [00116] In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in residues 158 to 172 of SEQ ID NO: 105 to a greater extent than in residues 206 to 215 of SEQ ID NO:
[00117] [00117] In one aspect, a composition comprising a plurality of anti-inflammatory molecules is disclosed herein.
[00118] [00118] In one aspect, a composition is disclosed herein comprising a plurality of antibody molecules that binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second - the CD73 monomer, where each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where : (i) when the antibody molecules in plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at most 20%, 25%, 30%, 35%, 40% , 45%, 50%, 55%, 60%, 65%, or 70% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, each of which complex comprises two or more antibody molecules and two or more dimers of CD73, for example, when measured using exclusion chromatography in size and percentage value is obtained by determining the amount of antibody molecules in the complex in relation to the total amount of antibody binding molecules to CD73 (excluding unbound antibody molecules), for example , as described in Example 2; and (ii) the antibody molecule, when linked to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), leads to a reduction in the average hydrogen-deuterium exchange of more than, for example, 0.02, 0.03, 0.04, 0.05 or 0.06 Da per residue in residues 368 to 387 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry, for example, as described in Example 2, for example , hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature.
[00119] [00119] In one aspect, a composition is disclosed herein comprising a plurality of antibody molecules that binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second - the CD73 monomer, where each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where : (i) when the antibody molecules in plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at most 20%, 25%, 30%, 35%, 40% , 45%, 50%, 55%, 60%, 65%, or 70% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, each of which complex comprises two or more antibody molecules and two or more dimers of CD73, for example, when measured using exclusion chromatography in size and percentage value is obtained by determining the amount of antibody molecules in the complex in relation to the total amount of antibody binding molecules to CD73 (excluding unbound antibody molecules), for example , as described in Example 2; and (ii) the antibody molecule, when linked to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), leads to a reduction in the average hydrogen-deuterium exchange of less than, for example, 0.05, 0.04, 0.03 or 0.02 Da per residue in residues 206 to 215 of SEQ ID NO: 105, example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry, for example, as described in Example 2, for example, mass mass spectrometry hydrogen deuterium exchange conducted for 1 minute in exchange at pH 7 to 8 (eg pH 7.5) and room temperature.
[00120] [00120] In one aspect, a composition is disclosed herein comprising a plurality of antibody molecules that binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second - the CD73 monomer, where each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where : (i) when the antibody molecules in plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at most 20%, 25%, 30%, 35%, 40% , 45%, 50%, 55%, 60%, 65%, or 70% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, each of which complex comprises two or more antibody molecules and two or more dimers of CD73, for example, when measured using exclusion chromatography in size and the percentage value is obtained by determining the amount of antibody molecules in the complex in relation to the total amount of antibody-binding molecules
[00121] [00121] In one aspect, a composition is disclosed herein comprising a plurality of antibody molecules that binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second - the CD73 monomer, where each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where : (i) when the antibody molecules in plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at most 20%, 25%, 30%, 35%, 40% , 45%, 50%, 55%, 60%, 65%, or 70% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, each of which complex comprises two or more antibody molecules and two or more dimers of CD73, for example, when measured using exclusion chromatography in size and percentage value is obtained by determining the amount of antibody molecules in the complex in relation to the total amount of antibody binding molecules to CD73 (excluding unbound antibody molecules), for example , as described in Example 2; and (ii) the antibody molecule inhibits at least about 60%, 70%, 80%, or 90% of the enzyme activity of human membrane-bound CD73, for example, when the antibody molecule is tested as an antibody molecule bivalent with the use of a modified Cell Titration Glo (CTG) assay, for example, as described in Example 1.
[00122] [00122] In one aspect, a composition is disclosed herein comprising a plurality of antibody molecules that binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second - the CD73 monomer, where each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where : (i) when the antibody molecules in plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at most 20%, 25%, 30%, 35%, 40% , 45%, 50%, 55%, 60%, 65%, or 70% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, each of which complex comprises two or more antibody molecules and two or more dimers of CD73, for example, when measured using exclusion chromatography in size and percentage value is obtained by determining the amount of antibody molecules in the complex in relation to the total amount of antibody binding molecules to CD73 (excluding unbound antibody molecules), for example , as described in Example 2; and (ii) the antibody molecule comprises a heavy chain variable region (VH) comprising an amino acid sequence of heavy chain complementarity determining region 3 (VHCDR3) from GGLYGSGSYLSDFDL (SEQ ID NO: 37) .
[00123] [00123] In one aspect, a composition is disclosed herein comprising a plurality of anti-inflammatory molecules
[00124] [00124] In one aspect, a composition is disclosed herein comprising a plurality of antibody molecules that binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second - the CD73 monomer, where each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where : (i) when the antibody molecules in plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at most 20%, 25%, 30%, 35%, 40% , 45%, 50%, 55%, 60%, 65%, or 70% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, each of which complex comprises two or more antibody molecules and two or more dimers of CD73, for example, when measured using exclusion chromatography in size and percentage value is obtained by determining the amount of antibody molecules in the complex in relation to the total amount of antibody binding molecules to CD73 (excluding unbound antibody molecules), for example , as described in Example 2; and (ii) the antibody molecule comprises: (a) a VH comprising a VÓHCDR1 amino acid sequence of SEQ ID NO: 38, a VHCDR2 amino acid sequence of SEQ ID NO: 36 and an amino acid sequence of VHCDR3 of SEQ ID NO: 37; and a VL that comprises a sequence
[00125] [00125] In one aspect, a composition is disclosed herein comprising a plurality of antibody molecules that binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second - the CD73 monomer, where each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where : (i) when the antibody molecules in plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at most 20%, 25%, 30%, 35%, 40% , 45%, 50%, 55%, 60%, 65%, or 70% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, each of which complex comprises two or more antibody molecules and two or more dimers of CD73, for example, when measured using exclusion chromatography in size and percentage value is obtained by determining the amount of antibody molecules in the complex in relation to the total amount of antibody binding molecules to CD73 (excluding unbound antibody molecules), for example , as described in Example 2; and (ii) the antibody molecule comprises a variable region
[00126] [00126] In one aspect, a composition is disclosed herein comprising a plurality of antibody molecules that binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second - the CD73 monomer, where each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where : (i) when the antibody molecules in plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at most 20%, 25%, 30%, 35%, 40% , 45%, 50%, 55%, 60%, 65%, or 70% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, each of which complex comprises two or more antibody molecules and two or more dimers of CD73, for example, when measured using exclusion chromatography in size and percentage value is obtained by determining the amount of antibody molecules in the complex in relation to the total amount of antibody binding molecules to CD73 (excluding unbound antibody molecules), for example , as described in Example 2; and (ii) the antibody molecule comprises: (a) a heavy chain variable region (VH) comprising an amino acid sequence of heavy chain complementarity determining region 1 (VHCDR1) from X: X2YWS (SEQ ID NO: 90), where X: is R, G or S, and X> is Y or R; a YÓYX VÓHCDR2 amino acid sequence: X2GSTX; YNPSLKS (SEQ ID NO:
[00127] [00127] In one aspect, a composition is disclosed herein comprising a plurality of antibody molecules that binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second - the CD73 monomer, where each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), where : (i) when the antibody molecules in plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at most 20%, 25%, 30%, 35%, 40% , 45%, 50%, 55%, 60%, 65%, or 70% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, each of which complex comprises two or more antibody molecules and two or more dimers of CD73, for example, when measured using exclusion chromatography in size and percentage value is obtained by determining the amount of antibody molecules in the complex in relation to the total amount of antibody binding molecules to CD73 (excluding unbound antibody molecules), for example , as described in Example 2; and
[00128] [00128] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (i) the antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the average hydrogen-deuterium exchange in residues 368 to 387 of SEQ ID NO: 105 a greater than in residues 158 to 172, residues 206 to 215 or residues 297 to 309 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule with the use of spectrometry of hydrogen deuterium exchange mass, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature; and (ii) the antibody molecule inhibits at least about 60%, 70%, 80%, or 90% of the enzyme activity of human membrane-bound CD73, for example, when the antibody molecule is tested as an antibody molecule bivalent with the use of a modified Cell Titration Glo (CTG) assay, for example, as described in Example 1.
[00129] [00129] In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in residues 368 to 387 of SEQ ID NO: 105 to a greater extent than in residues 158 to 172 of SEQ ID NO:
[00130] [00130] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, in which: (i) the antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the average hydrogen-deuterium exchange in residues 158 to 172 of SEQ ID NO: 105 to a greater extent than in residues 206 to 215 of SEQ ID NO: 105, residues 368 to 387 of SEQ ID NO: 105 or residues 297 to 309 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule with the use of hydrogen deuterium exchange mass spectrometry, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange for pH 7 to 8 (e.g., pH 7.5) and room temperature; and (ii) the antibody molecule inhibits at least about
[00131] [00131] In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in residues 158 to 172 of SEQ ID NO: 105 to a greater extent than in residues 206 to 215 of SEQ ID NO:
[00132] [00132] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, in which: (i) the antibody molecule reduces the hydrogen-deuterium exchange in one or more regions of a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171) when linked to it, where one or more regions are selected from the group consisting of residues 158 to 172, residues 206 to 215, residues 368 to 387 and residues 87 to 104 of SEQ ID NO: 105, in which the region with the greatest reduction in hydrogen-deuterium exchange mean between one or more regions does not consist of residues 206 to 215 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule using deuterium exchange mass spectrometry hydrogen, for example, as described in Example 2, for example, spectrometry d and hydrogen deuterium exchange mass conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature; and (ii) the antibody molecule inhibits at least about 60%, 70%, 80%, or 90% of the enzyme activity of human membrane-bound CD73, for example, when the antibody molecule is tested as an antibody molecule bivalent with the use of a modified Cell Titration Glo (CTG) assay, for example, as described in Example 1.
[00133] [00133] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, in which: (i) the antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), leads to a reduction in the average hydrogen-deuterium exchange of more than, for example, 0.02,
[00134] [00134] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, wherein: (i) the antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), leads to a reduction in the average hydrogen-deuterium exchange of less than, for example, 0.05, 0 , 04, 0.03 or 0.02 Da per residue in residues 206 to 215 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule using mass spectrometry hydrogen deuterium exchange, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature; and (ii) the antibody molecule inhibits at least about 60%, 70%, 80%, or 90% of the enzyme activity of human membrane-bound CD73, for example, when the antibody molecule is tested as an antibody molecule bivalent with the use of a modified Cell Titration Glo (CTG) assay, for example, as described in Example 1.
[00135] [00135] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (i) the antibody molecule binds to at least one, two, three or four residues within residues 158 to 172 of SEQ ID NO: 105, and / or at least one, two, three, four or five residues within residues 206 to 215 of SEQ ID NO: 105; and (ii) the antibody molecule inhibits at least about 60%, 70%, 80%, or 90% of the enzyme activity of human membrane-bound CD73, for example, when the antibody molecule is tested as an antibody molecule bivalent with the use of a modified Cell Titration Glo (CTG) assay, for example, as described in Example 1.
[00136] [00136] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, in which: (i) the antibody molecule, when bound to a protein that comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the average hydrogen-deuterium exchange in residues 368 to 387 of SEQ ID NO: 105 a greater than in residues 158 to 172, residues 206 to 215 or residues 297 to 309 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule with the use of spectrometry of hydrogen deuterium exchange mass, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature; and
[00137] [00137] In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in residues 368 to 387 of SEQ ID NO: 105 to a greater extent than in residues 158 to 172 of SEQ ID NO:
[00138] [00138] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, in which:
[00139] [00139] In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in residues 158 to 172 of SEQ ID NO: 105 to a greater extent than in residues 206 to 215 of SEQ ID NO:
[00140] [00140] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, in which: (i) the antibody molecule reduces the hydrogen-deuterium exchange in one or more regions of a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171) when linked to it, where one or more regions are selected from the group consisting of residues 158 to 172, residues 206 to 215, residues 368 to 387 and residues 87 to 104 of SEQ ID NO: 105, in which the region with the greatest reduction in hydrogen-deuterium exchange mean between one or more regions does not consist of residues 206 to 215 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule using deuterium exchange mass spectrometry hydrogen, for example, as described in Example 2, for example, spectrometry d and hydrogen deuterium exchange mass conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature; and (ii) the antibody molecule comprises a variable region
[00141] [00141] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, in which: (i) the antibody molecule, when bound to a protein that comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), leads to a reduction in the average hydrogen-deuterium exchange of more than, for example, 0.02, 0.03, 0.04, 0.05 or 0.06 Da per residue in residues 368 to 387 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule with the use of hydrogen deuterium exchange mass spectrometry, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature; and (ii) the antibody molecule comprises a heavy chain variable region (VH) comprising an amino acid sequence of heavy chain complementarity determining region 3 (VHCDR3) from GGLYGSGSYLSDFDL (SEQ ID NO: 37) .
[00142] [00142] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, wherein: (i) the antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), leads to a reduction in the average hydrogen-deuterium exchange of less than, for example, 0.05, 0 , 04, 0.03 or 0.02 Da per residue in residues 206 to 215 of SEQ ID NO: 105, for example, when the antibody molecule is tested co-
[00143] [00143] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (i) the antibody molecule binds to at least one, two, three or four residues within residues 158 to 172 of SEQ ID NO: 105, and / or at least one, two, three, four or five residues within residues 206 to 215 of SEQ ID NO: 105; and (ii) the antibody molecule comprises a heavy chain variable region (VH) comprising an amino acid sequence of heavy chain complementarity determining region 3 (VHCDR3) from GGLYGSGSYLSDFDL (SEQ ID NO: 37) .
[00144] [00144] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, in which: (i) the antibody molecule, when bound to a protein that comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the average hydrogen-deuterium exchange in residues 368 to 387 of SEQ ID NO: 105 a greater than in residues 158 to 172, residues 206 to 215 or residues 297 to 309 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule with the use of spectrometry of hydrogen deuterium exchange mass, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature; and (ii) the antibody molecule comprises: (a) a heavy chain variable region (VH) comprising an amino acid sequence of heavy chain complementarity determining region 1 (VHCDR1) from X: X2AMS (SEQ ID NO: 88), where Xx is R, Y or S, and Xo is Y or N; a VHCDR2 amino acid sequence from X1IX2GX3GXaXsTYYADSVKG (SEQ ID NO: 89), where X is A or S, Xoé S or T, Xgé Sou T, x É M, GouS, and Xs is N, S, Lou Y; and a VHCDR3 amino acid sequence from GGLYGSGSYLSDFDL (SEQ ID NO: 37) and (b) a light chain variable region (VL) comprising a light chain 1 complementarity determining region amino acid sequence (VLCDR1) from RASQSVGSNLA (SEQ ID NO: 48); a VLCDR2 amino acid sequence from GASTRAT (SEQ ID NO: 49); and a VLCDR3 amino acid sequence from QQHNAFPYT (SEQ ID NO: 50).
[00145] [00145] In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in residues 368 to 387 of SEQ ID NO: 105 to a greater extent than in residues 158 to 172 of SEQ ID NO:
[00146] [00146] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, in which: (i) the antibody molecule, when bound to a protein that comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the average hydrogen-deuterium exchange in residues 158 to 172 of SEQ ID NO: 105 a a greater extent than in residues 206 to 215 of SEQ ID NO: 105, residues 368 to 387 of SEQ ID NO: 105 or residues 297 to 309 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule with the use of hydrogen deuterium exchange mass spectrometry, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange for pH 7 to 8 (e.g., pH 7.5) and room temperature; and (ii) the antibody molecule comprises: (a) a heavy chain variable region (VH) comprising an amino acid sequence of X1X2AMS heavy chain complementarity determining region (VHCDR1) (SEQ ID NO: 88), where X1 is R You S and Xcé You N; a VHCDR2 amino acid sequence from X1IX2GX3GX4aX5TYYADSVKG (SEQ ID NO: 89), where Xx is A or S, X2éSouT, X éSouT, X: is M, GouS, eXséN, S, LouY; (SEQ ID NO: 37); and (b) a light chain variable region (VL) comprising an amino acid sequence of light chain complementarity determining region 1 (VLCDR1) from RASQSVGSNLA (SEQ ID NO: 48); a VLCDR2 amino acid sequence from GASTRAT (SEQ ID NO: 49); and a VLCDR3 amino acid sequence from QQHNAFPYT (SEQ ID NO: 50).
[00147] [00147] In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in residues 158 to 172 of SEQ ID NO: 105 to a greater extent than in residues 206 to 215 of SEQ ID NO:
[00148] [00148] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, in which: (i) the antibody molecule reduces the hydrogen-deuterium exchange in one or more regions of a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171) when linked to it, where one or more regions are selected from the group consisting of residues 158 to 172, residues 206 to 215, residues 368 to 387 and residues 87 to 104 of SEQ ID NO: 105, in which the region with the greatest reduction in hydrogen-deuterium exchange mean between one or more regions does not consist of residues 206 to 215 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule using deuterium exchange mass spectrometry hydrogen, for example, as described in Example 2, for example, spectrometry d and hydrogen deuterium exchange mass conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature; and (ii) the antibody molecule comprises: (a) a heavy chain variable region (VH) comprising an amino acid sequence of heavy chain complementarity determining region 1 (VHCDR1) from X; X2AMS (SEQ
[00149] [00149] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, wherein: (i) the antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), leads to a reduction in the average hydrogen-deuterium exchange of more than, for example, 0.02, 0.03, 0.04, 0.05 or 0.06 Da per residue in residues 368 to 387 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule with the use of hydrogen deuterium exchange mass spectrometry, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature; and (ii) the antibody molecule comprises: (a) a heavy chain variable region (VH) comprising an amino acid sequence of heavy chain complementarity determining region 1 (VHCDR1) from X: X2AMS (SEQ ID NO: 88), where X: is R, You S, and Xo is Y or N; a VÓHCDR2 amino acid sequence of XIX2GX3GXaXsTYYADSVKG (SEQ ID NO: 89), where X is Aou S, Xoé Sou T, Xgé Sou T, 4 É M, GouS, and Xs is N, S, Lou Y; and a VHCDR3 amino acid sequence from GGLYGSGSYLSDFDL (SEQ ID NO: 37) and (b) a light chain variable region (VL) comprising a light chain 1 complementarity determining region amino acid sequence (VLCDR1) from RASQSVGSNLA (SEQ ID NO: 48); a VLCDR2 amino acid sequence from GASTRAT (SEQ ID NO: 49); and a VLCDR3 amino acid sequence from QQHNAFPYT (SEQ ID NO: 50).
[00150] [00150] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, in which: (i) the antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), leads to a reduction in the average hydrogen-deuterium exchange of less than, for example, 0.05, 0 , 04, 0.03 or 0.02 Da per residue in residues 206 to 215 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule using mass spectrometry hydrogen deuterium exchange, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature; and (ii) the antibody molecule comprises: (a) a heavy chain variable region (VH) comprising an amino acid sequence of the heavy chain complementarity determining region 1 (VHCDR1) of X; X2AMS (SEQ ID NO: 88), where Xx is R, You S, and Xo is Y or N; a VHCDR2 amino acid sequence of XIX2GX3; GXaXsTYYADSVKG (SEQ ID
[00151] [00151] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (i) the antibody molecule binds to at least one, two, three or four residues within residues 158 to 172 of SEQ ID NO: 105, and / or at least one, two, three, four or five residues within residues 206 to 215 of SEQ ID NO: 105; and (ii) the antibody molecule comprises: (a) a heavy chain variable region (VH) comprising an amino acid sequence of X heavy chain complementarity determining region (VHCDR1); X2AMS (SEQ ID NO: 88), where X: is R, You S, and X2o is Y or N; a XIX2GX VHCDR2 amino acid sequence; GXaXsTYYADSVKG (SEQ ID NO: 89), where X1 is Aou S, Xoé S or T, Xgé Sou T, x is M, GouS, and Xs is N, S, Lou Y; and a VHCDR3 amino acid sequence from GGLYGSGSYLSDFDL (SEQ ID NO: 37) and (b) a light chain variable region (VL) comprising a light chain 1 complementarity determining region amino acid sequence (VLCDR1) from RASQSVGSNLA (SEQ ID NO: 48); a VLCDR2 amino acid sequence from GASTRAT (SEQ ID NO: 49); and a VLCDR3 amino acid sequence from QQHNAFPYT (SEQ ID NO: 50).
[00152] [00152] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, in which:
[00153] [00153] In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in residues 368 to 387 of SEQ ID NO:
[00154] [00154] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (i) the antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the average hydrogen-deuterium exchange in residues 158 to 172 of SEQ ID NO: 105 to greater than in residues 206 to 215 of SEQ ID NO: 105,
[00155] [00155] In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in residues 158 to 172 of SEQ ID NO: 105 to a greater extent than in residues 206 to 215 of SEQ ID NO:
[00156] [00156] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, in which: (i) the antibody molecule reduces the hydrogen-deuterium exchange in one or more regions of a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171) when linked to it, where one or more regions are selected from the group consisting of residues 158 to 172, residues 206 to 215, residues 368 to 387 and residues 87 to 104 of SEQ ID NO: 105, in which the region with the greatest reduction in hydrogen-deuterium exchange mean between one or more regions does not consist of residues 206 to 215 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule using deuterium exchange mass spectrometry hydrogen, for example, as described in Example 2, for example, spectrometry d and hydrogen deuterium exchange mass conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature; and
[00157] [00157] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (i) the antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), leads to a reduction in the average hydrogen-deuterium exchange of more than, for example, 0.02, 0.03, 0.04, 0.05 or 0.06 Da per residue in residues 368 to 387 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule with the use of hydrogen deuterium exchange mass spectrometry, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature; and (ii) the antibody molecule comprises:
[00158] [00158] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, in which: (i) the antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), leads to a reduction in the average hydrogen-deuterium exchange of less than, for example, 0.05, 0 , 04, 0.03 or 0.02 Da per residue in residues 206 to 215 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule using mass spectrometry hydrogen deuterium exchange, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature; and (ii) the antibody molecule comprises a heavy chain variable region (VH) comprising an amino acid sequence of heavy chain complementarity determining region 3 (VHCDR3) from GGLYGSGSYLSDFDL (SEQ ID NO: 37) .
[00159] [00159] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (i) the antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the average hydrogen-deuterium exchange in residues 368 to 387 of SEQ ID NO: 105 a greater than in residues 158 to 172, residues 206 to 215 or residues 297 to 309 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule with the use of spectrometry of hydrogen deuterium exchange mass, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature; and (ii) the antibody molecule comprises a heavy chain variable region (VH) comprising an amino acid sequence of heavy chain complementarity determining region 3 (VHCDR3) from ESQESPYNNWFDP (SEQ | D NO: 3 ).
[00160] [00160] In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in residues 368 to 387 of SEQ ID NO: 105 to a greater extent than in residues 158 to 172 of SEQ ID NO:
[00161] [00161] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, in which: (i) the antibody molecule, when bound to a protein that comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the average hydrogen-deuterium exchange in residues 158 to 172 of SEQ ID NO: 105 to a greater extent than in residues 206 to 215 of SEQ ID NO: 105, residues 368 to 387 of SEQ ID NO: 105 or residues 297 to 309 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule with the use of hydrogen deuterium exchange mass spectrometry, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange for pH 7 to 8 (e.g., pH 7.5) and room temperature; and (ii) the antibody molecule comprises a heavy chain variable region (VH) comprising an amino acid sequence of heavy chain complementarity determining region 3 (VHCDR3) from ESQESPYNNWFDP (SEQ | D NO: 3 ).
[00162] [00162] In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in residues 158 to 172 of SEQ ID NO: 105 to a greater extent than in residues 206 to 215 of SEQ ID NO:
[00163] [00163] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, in which: (i) the antibody molecule reduces the hydrogen-deuterium exchange in one or more regions of a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171) when linked to it, where one or more regions are selected from the group consisting of residues 158 to 172, residues 206 to 215, residues 368 to 387 and residues 87 to 104 of SEQ ID NO: 105, in which the region with the greatest reduction in hydrogen-deuterium exchange mean between one or more regions does not consist of residues 206 to 215 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule using deuterium exchange mass spectrometry hydrogen, for example, as described in Example 2, for example, spectrometry d and hydrogen deuterium exchange mass conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature; and (ii) the antibody molecule comprises a heavy chain variable region (VH) comprising an amino acid sequence of heavy chain complementarity determining region 3 (VHCDR3) from ESQESPYNNWFDP (SEQ | D NO: 3 ).
[00164] [00164] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, wherein: (i) the antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), leads to a reduction in the average hydrogen-deuterium exchange of more than, for example, 0.02, 0.03, 0.04, 0.05 or 0.06 Da per residue in residues 368 to 387 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule with the use of hydrogen deuterium exchange mass spectrometry, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature; and (ii) the antibody molecule comprises a heavy chain variable region (VH) comprising an amino acid sequence of ESQESPYNNWFDP heavy chain complementarity determining region 3 (VHCDR3) (SEQ ID NO: 3) .
[00165] [00165] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (i) the antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 SEQ ID NO: 105 (for example, a protein consisting of the
[00166] [00166] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (i) the antibody molecule binds to at least one, two, three or four residues within residues 158 to 172 of SEQ ID NO: 105, and / or at least one, two, three, four or five residues within residues 206 to 215 of SEQ ID NO: 105; and (ii) the antibody molecule comprises a heavy chain variable region (VH) comprising an amino acid sequence of heavy chain complementarity determining region 3 (VHCDR3) from ESQESPYNNWFDP (SEQ | D NO: 3 ).
[00167] [00167] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, wherein: (i) the antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the average hydrogen-deuterium exchange in residues 368 to 387 of SEQ ID NO: 105 a greater than in residues 158 to 172, residues 206 to 215 or residues 297 to 309 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule with the use of spectrometry of hydrogen deuterium exchange mass, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature; and (ii) the antibody molecule comprises: (a) a heavy chain variable region (VH) comprising an amino acid sequence of heavy chain complementarity determining region 1 (VHCDR1) from X: X2YWS (SEQ ID NO: 90), where X: is R, G or S, and Xo is Y or R; a VÓHCDR2 amino acid sequence from YIYX: X2GSTX3; YNPSLKS (SEQ ID NO: 91), where X1 is Gou S, co is R, Sou T, and Xg is Nou K; and a VHCDR3 amino acid sequence of ESQESPYNNWFDP (SEQ ID NO: 3) and (b) a light chain variable region (VL) comprising a light chain complementarity determining region amino acid sequence 1 ( VLCDR1) from RASQGISSWLA (SEQ ID NO: 14); an AASSLOS VLCDR2 amino acid sequence (SEQ ID NO: 15); and a VLCDR3 amino acid sequence from QQOGNSFPRT (SEQ ID NO: 16).
[00168] [00168] In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in residues 368 to 387 of SEQ ID NO: 105 to a greater extent than in residues 158 to 172 of SEQ ID NO:
[00169] [00169] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (i) the antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the average hydrogen-deuterium exchange in residues 158 to 172 of SEQ ID NO: 105 to a greater extent than in residues 206 to 215 of SEQ ID NO: 105, residues 368 to 387 of SEQ ID NO: 105 or residues 297 to 309 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule with the use of hydrogen deuterium exchange mass spectrometry, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange for pH 7 to 8 (e.g., pH 7.5) and room temperature; and (ii) the antibody molecule comprises: (a) a heavy chain variable region (VH) comprising an amino acid sequence of heavy chain complementarity determining region 1 (VHCDR1) from X: X2YWS (SEQ ID NO: 90), where X: is R, G or S, and X> is Y or R; a VÓHCDR2 amino acid sequence from YIYX: X2GSTX3; YNPSLKS (SEQ ID NO: 91), where X1 is G or S, Cc is R, Sou T, and Xg is N or K; and a VÓHCDR3 amino acid sequence from ESQESPYNNWFDP (SEQ ID NO: 3) and (b) a light chain variable region (VL) comprising an amino acid sequence from the light chain complementarity determining region 1 ( VLCDR1) from RASQGISSWLA (SEQ ID NO: 14); an AASSLOS VLCDR2 amino acid sequence (SEQ ID NO: 15); and a VLCDR3 amino acid sequence from QQOGNSFPRT (SEQ ID NO: 16).
[00170] [00170] In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in residues 158 to 172 of SEQ ID NO: 105 to a greater extent than in residues 206 to 215 of SEQ ID NO:
[00171] [00171] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, in which: (i) the antibody molecule reduces the hydrogen-deuterium exchange in one or more regions of a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171) when linked to it, where one or more regions are selected from the group consisting of residues 158 to 172, residues 206 to 215, residues 368 to 387 and residues 87 to 104 of SEQ ID NO: 105, in which the region with the greatest reduction in hydrogen-deuterium exchange mean between one or more regions does not consist of residues 206 to 215 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule using deuterium exchange mass spectrometry hydrogen, for example, as described in Example 2, for example, spectrometry d and hydrogen deuterium exchange mass conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature; and (ii) the antibody molecule comprises:
[00172] [00172] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, in which: (i) the antibody molecule, when bound to a protein that comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), leads to a reduction in the average hydrogen-deuterium exchange of more than, for example, 0.02, 0.03, 0.04, 0.05 or 0.06 Da per residue in residues 368 to 387 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule with the use of hydrogen deuterium exchange mass spectrometry, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature; and (ii) the antibody molecule comprises: (a) a heavy chain variable region (VH) that comprises
[00173] [00173] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, in which: (i) the antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), leads to a reduction in the average hydrogen-deuterium exchange of less than, for example, 0.05, 0 , 04, 0.03 or 0.02 Da per residue in residues 206 to 215 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule using mass spectrometry hydrogen deuterium exchange, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature; and (ii) the antibody molecule comprises: (a) a heavy chain variable region (VH) comprising an amino acid sequence of heavy chain complementarity determining region 1 (VHCDR1) from X: X2YWS (SEQ ID NO: 90), where X: is R, G or S, and X> is Y or R; a VÓHCDR2 amino acid sequence from YIYX: X2GSTX:; YNPSLKS (SEQ ID NO: 91), where X: is G or S, Cc is R, S or T, and Xg is Nou K; and a VÓHCDR3 amino acid sequence of ESQESPYNNWFDP (SEQ | D NO: 3) and (b) a light chain variable region (VL) that comprises a sequence of amino acids from the light chain complementarity determining region 1 (VLCDR1) from RASQGISSWLA (SEQ ID NO: 14); an AASSLOS VLCDR2 amino acid sequence (SEQ ID NO: 15); and a VLCDR3 amino acid sequence from QQOGNSFPRT (SEQ ID NO: 16).
[00174] [00174] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (i) the antibody molecule binds to at least one, two, three or four residues within residues 158 to 172 of SEQ ID NO: 105, and / or at least one, two, three, four or five residues within residues 206 to 215 of SEQ ID NO: 105; and (ii) the antibody molecule comprises: (a) a heavy chain variable region (VH) comprising an amino acid sequence of heavy chain complementarity determining region 1 (VHCDR1) from X: X2YWS (SEQ ID NO: 90), where X: is R, G or S, and Xo is Y or R; a VÓHCDR2 amino acid sequence from YIYX: X2GSTX; YNPSLKS (SEQ ID NO: 91), where X1 is G or S, co is R, Sou T, and Xg is Nou K; and a VHCDR3 amino acid sequence of ESQESPYNNWFDP (SEQ ID NO: 3) and (b) a light chain variable region (VL) comprising a light chain complementarity determining region amino acid sequence 1 ( VLCDR1) from RASQGISSWLA
[00175] [00175] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (i) the antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the average hydrogen-deuterium exchange in residues 368 to 387 of SEQ ID NO: 105 a greater than in residues 158 to 172, residues 206 to 215 or residues 297 to 309 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule with the use of spectrometry of hydrogen deuterium exchange mass, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature; and (ii) the antibody molecule comprises: (a) a VH comprising a VHCDR1 amino acid sequence of SEQ ID NO: 61, a VHCDR2 amino acid sequence of SEQ ID NO: 60 and an amino acid sequence of VHCDR3 of SEQ ID NO: 3; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 14, a VLCDR2 amino acid sequence of SEQ ID NO: 15 and a VLCDR3 amino acid sequence of SEQ ID NO: 16; (b) a VH comprising a VHCDR1 amino acid sequence of SEQ ID NO: 4, a VHCDR 2 amino acid sequence of SEQ ID NO: 26 and a VHCDR3 amino acid sequence of SEQ ID NO: 3; and a VL that comprises a sequence
[00176] [00176] In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in residues 368 to 387 of SEQ ID NO: 105 to a greater extent than in residues 158 to 172 of SEQ ID NO:
[00177] [00177] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (i) the antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the average hydrogen-deuterium exchange in residues 158 to 172 of SEQ ID NO: 105 to a greater extent than in residues 206 to 215 of SEQ ID NO: 105, residues 368 to 387 of SEQ ID NO: 105 or residues 297 to 309 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule with the use of hydrogen deuterium exchange mass spectrometry, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange for pH 7 to 8 (e.g., pH 7.5) and room temperature; and (ii) the antibody molecule comprises: (a) a VH comprising a VHCDR1 amino acid sequence of SEQ ID NO: 61, a VHCDR2 amino acid sequence of SEQ ID NO: 60 and an amino acid sequence of
[00178] [00178] In one embodiment, antibody binding reduces the average hydrogen-deuterium exchange in residues 158 to 172 of SEQ ID NO: 105 to a greater extent than in residues 206 to 215 of SEQ ID NO:
[00179] [00179] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, in which: (i) the antibody molecule reduces the hydrogen-deuterium exchange in one or more regions of a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171) when linked to it, where one or more regions are selected from the group consisting of residues 158 to 172, residues 206 to 215, residues 368 to 387 and residues 87 to 104 of SEQ ID NO: 105, in which the region with the greatest reduction in hydrogen-deuterium exchange mean between one or more regions does not consist of residues 206 to 215 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule using deuterium exchange mass spectrometry hydrogen, for example, as described in Example 2, for example, spectrometry d and hydrogen deuterium exchange mass conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature; and
[00180] [00180] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, in which: (i) the antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), leads to a reduction in the average hydrogen-deuterium exchange of more than, for example, 0.02, 0.03, 0.04, 0.05 or 0.06 Da per residue in residues 368 to 387 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule with the use of hydrogen deuterium exchange mass spectrometry, for example, as described in Example 2, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature; and (ii) the antibody molecule comprises: (a) a VH comprising an amino acid sequence of VHCDR1 of SEQ ID NO: 61, an amino acid sequence of VHCDR2 of SEQ ID NO: 60 and an amino acid sequence of VHCDR3 of SEQ ID NO: 3; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 14, a VLCDR2 amino acid sequence of SEQ ID NO: 15 and a VLCDR3 amino acid sequence of SEQ ID NO: 16; (b) a VH comprising an amino acid sequence of VHCDR1 of SEQ ID NO: 4, an amino acid sequence of
[00181] [00181] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, in which: (i) the antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), leads to a reduction in the average hydrogen-deuterium exchange of less than, for example, 0.05, 0 , 04, 0.03 or 0.02 Da per residue in residues 206 to 215 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule using mass spectrometry of hydrogen deuterium exchange, for example, as
[00182] [00182] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (i) the antibody molecule binds to at least one, two, three or four residues within residues 158 to 172 of SEQ ID NO: 105, and / or at least one, two, three, four or five residues within residues 206 to 215 of SEQ ID NO: 105; and (ii) the antibody molecule comprises: (a) a VH comprising a VÓHCDR1 amino acid sequence of SEQ ID NO: 61, a VÓHCDR2 amino acid sequence of SEQ ID NO: 60 and a sequence of VHCDR3 amino acids of SEQ ID NO: 3; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 14, a VLCDR2 amino acid sequence of SEQ ID NO: 15 and a VLCDR3 amino acid sequence of SEQ ID NO: 16; (b) a VH comprising a VÓHCDR1 amino acid sequence of SEQ ID NO: 4, a VHCDR 2 amino acid sequence of SEQ ID NO: 26 and a VHCDR3 amino acid sequence of SEQ ID NO: 3; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 14, a VLCDR2 amino acid sequence of SEQ ID NO: 15 and a VLCDR3 amino acid sequence of SEQ ID NO: 16; (c) a VH comprising a VÓHCDR1 amino acid sequence of SEQ ID NO: 4, a VHCDR2 amino acid sequence of SEQ ID NO: 2 and a VHCDR3 amino acid sequence of SEQ ID NO: 3; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 14, a VLCDR2 amino acid sequence of SEQ ID NO: 15 and a VLCDR3 amino acid sequence of SEQ ID NO: 16; or
[00183] [00183] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (i) the antibody molecule inhibits at least about 60%, 70%, 80%, or 90% of the enzymatic activity of human membrane-bound CD73, for example, when the antibody molecule is tested as a bivalent antibody molecule using a modified Cell Titration (CTG) assay, for example, as described in Example 1 ; and (ii) the antibody molecule comprises: (a) a heavy chain variable region (VH) comprising an amino acid sequence of heavy chain complementarity determining region 1 (VHCDR1) from X: X2AMS (SEQ ID NO: 88), where X: is R, You S, and X2o is Y or N; a VHCDR2 amino acid sequence of XIX2GX3; 3GXaXsTYYADSVKG (SEQ ID NO: 89), where X1 is Aou S, Xoé S or T, Xgé Sou T, x is M, GouS, and Xs is N, S, Lou Y; and a VHCDR3 amino acid sequence from GGLYGSGSYLSDFDL (SEQ ID NO: 37) and (b) a light chain variable region (VL) comprising a light chain 1 complementarity determining region amino acid sequence (VLCDR1) from RASQSVGSNLA (SEQ ID NO: 48); a VLCDR2 amino acid sequence from GASTRAT (SEQ ID NO: 49); and a VLCDR3 amino acid sequence from QQHNAFPYT (SEQ ID NO: 50).
[00184] [00184] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (i) the antibody molecule inhibits at least about 60%, 70%, 80%, or 90% of the enzymatic activity of human membrane-bound CD73, for example, when the antibody molecule is tested as a bivalent antibody molecule using a modified Cell Titration (CTG) assay, for example, as described in Example 1 ; and (ii) the antibody molecule comprises: (a) a VH comprising a VHCDR1 amino acid sequence of SEQ ID NO: 38, a VHCDR2 amino acid sequence of SEQ ID NO: 36 and an amino acid sequence of VHCDR3 of SEQ ID NO: 37; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 48, a VLCDR2 amino acid sequence of SEQ ID NO: 49 and a VLCDR3 amino acid sequence of SEQ ID NO: 50; (b) a VH comprising a VHCDR1 amino acid sequence of SEQ ID NO: 72, a VHCDR2 amino acid sequence of SEQ ID NO: 71 and a VHCDR3 amino acid sequence of SEQ ID NO: 37; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 48, a VLCDR2 amino acid sequence of SEQ ID NO: 49 and a VLCDR3 amino acid sequence of SEQ ID NO: 50;
[00185] [00185] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (i) the antibody molecule inhibits at least about 60%, 70%, 80%, or 90% of the enzymatic activity of human membrane-bound CD73, for example, when the antibody molecule is tested as a bivalent antibody molecule using a modified Cell Titration (CTG) assay, for example, as described in Example 1 ; and; (ii) the antibody molecule comprises a heavy chain variable (VH) region comprising an amino acid sequence of ESQESPYNNWFDP heavy chain complementarity determining region 3 (VHCDR3) (SEQ ID NO: 3).
[00186] [00186] In one aspect, an antibody molecule that binds to human CD73 is disclosed in this document, in which: (i) the antibody molecule inhibits at least about 60%, 70%, 80%, or 90% of the enzyme activity of membrane-bound human CD73, for example, when the antibody molecule is tested as a bivalent antibody molecule using a modified Cell Titration Glucose (CTG) assay, for example, as described in Example 1; and (ii) the antibody molecule comprises: (a) a heavy chain variable region (VH) comprising an amino acid sequence of heavy chain complementarity determining region 1 (VHCDR1) from X: X2YWS (SEQ ID NO: 90), where X: is R, G or S, and X> is Y or R; a VÓHCDR2 amino acid sequence from YIYX: X2GSTX; YNPSLKS (SEQ ID NO: 91), where X1 is G or S, c is R, Sou T, and Xg is N or K; and a
[00187] [00187] In one aspect, an antibody molecule that binds to human CD73 is disclosed herein, in which: (i) the antibody molecule inhibits at least about 60%, 70%, 80%, or 90% of the enzymatic activity of human membrane-bound CD73, for example, when the antibody molecule is tested as a bivalent antibody molecule using a modified Cell Titration (CTG) assay, for example, as described in Example 1 ; and; (ii) the antibody molecule comprises: (a) a VH comprising a VHCDR1 amino acid sequence of SEQ ID NO: 61, a VHCDR2 amino acid sequence of SEQ ID NO: 60 and a VHCDR3 amino acid sequence SEQ ID NO: 3; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 14, a VLCDR2 amino acid sequence of SEQ ID NO: 15 and a VLCDR3 amino acid sequence of SEQ ID NO: 16; (b) a VH comprising a VHCDR1 amino acid sequence of SEQ ID NO: 4, a VHCDR 2 amino acid sequence of SEQ ID NO: 26 and a VHCDR3 amino acid sequence of SEQ ID NO: 3; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 14, a VLCDR2 amino acid sequence of SEQ ID NO: 15 and a amino acid sequence
[00188] [00188] The antibody molecules disclosed in the present document can modulate (for example, intensify, stimulate, increase, inhibit, reduce or neutralize) one or more CD73 activities. In some embodiments, the antibody molecule results in one or more of: inhibiting or reducing the enzymatic activity of CD73; inhibit or reduce the conversion of adenosine monophosphate (AMP) to adenosine; and increasing the proliferation of stimulated anti-CD3 / anti-CD28 T cells, for example, CD4 + T cells, in the presence of adenosine monophosphate (AMP).
[00189] [00189] In some embodiments, the antibody molecule inhibits or reduces the enzymatic activity of CD73 (for example, soluble human CD73 or membrane-bound human CD73), for example, conversion by human CD73 of adenosine monophosphate (AMP ) in adenosine, for example, as measured by a method described in this document, for example, a malachite green phosphate (MG) assay or a modified Cell Titration Glucose (CTG) assay, for example, as described in Example 1.
[00190] [00190] In some embodiments, the antibody molecule inhibits at least about 60%, 70%, 80%, or 90% of the enzyme activity of human membrane-bound CD73, for example, when the antibody molecule is tested as a bivalent antibody molecule using a modified Cell Titration Glo (CTG) assay, for example, as described in Example 1. In one embodiment, the antibody molecule inhibits at least about 60%, 70 %, 80%, or 90% of the enzyme activity of membrane-bound human CD73, for example, when the antibody molecule is tested as a divalent antibody molecule using a modified Cell Titration Glo (CTG) assay that comprises the following steps: (i) incubating a dose titration of the antibody molecule (eg 1000 ng / ml) with 20,000 cells / ml of a human cancer cell line that expresses human CD73 (eg - plo, the human breast cancer cell line MDA-MB-231 or the cancer cell line from the human ovary SKOV3) for 240 minutes at 37ºC in the presence of 100 µM AMP; (ii) measure the disappearance of AMP using a modified Cell Titration (CTG) assay as described in Example 1; and (iii) calculate the percentage of inhibition mediated by the antibody molecule with the use of zero time control as 100% inhibition and no antibody control as 0% inhibition.
[00191] [00191] In some embodiments, the antibody molecule increases the proliferation of stimulated anti-CD3 / anti-CD28 T cells, for example, CD4 + T cells, in the presence of adenosine monophosphate
[00192] [00192] In certain respects, a method of modulating (for example, stimulating or inhibiting) an immune response in an individual is provided. The method comprises administering to the individual an anti-CD73 antibody molecule disclosed herein, (for example, a therapeutically effective amount of an anti-CD73 antibody molecule), alone or in combination with one or more agents or procedures (for example, in combination with antitumor therapies, for example, chemotherapies, radiation therapies and / or other immunomodulatory agents), so that the immune response in the individual is modulated. In some embodiments, the antibody molecule inhibits, reduces or neutralizes an immune response in an individual.
[00193] [00193] The individual may be a mammal, for example, a monkey, a primate, preferably a higher primate, for example, a human being (for example, a patient who has, or is at risk for, a disorder described in this document). In some modalities, the individual needs to intensify an immune response, and in some modalities, the individual needs to inhibit an immune response. In one embodiment, the individual has, or is at risk of, having a disorder described in this document, for example, a cancer as described in this document. In certain modalities, the subject is, or is at risk of being, immunocompromised. For example, the subject is being subjected to or has undergone chemotherapeutic treatment and / or radiation therapy.
[00194] [00194] In one aspect, a method of stimulating an immune response in an individual is provided. The method comprises administering to the subject an anti-CD73 antibody molecule described in the present document, for example, a therapeutically effective amount of an anti-CD73 antibody molecule, alone or in combination with one or more agents or procedures.
[00195] [00195] In one aspect, a method of treating (for example, one or more among reducing, inhibiting, or delaying progression) a cancer or tumor in an individual is provided. The method comprises administering to the subject an anti-CD73 antibody molecule described in the present document, for example, a therapeutically effective amount of an anti-CD73 antibody molecule, alone or in combination with one or more agents or procedures.
[00196] [00196] In some embodiments, the antibody molecule is administered in combination with a second procedure or therapeutic agent. In some modalities, the second procedure or therapeutic agent is chosen from one or more among chemotherapy, a targeted anticancer therapy, an oncolytic drug, a cytotoxic agent, an immunologically based therapy, a cytokine, a surgical procedure, a procedure radiation, an activator of a co-stimulating molecule, an inhibitor of an inhibitory molecule (for example, an inhibitor of a checkpoint inhibitor), a vaccine or cell therapy. In some embodiments, the second therapeutic agent is chosen from one or more of: 1) a protein C kinase inhibitor (PKC); 2) a heat shock protein inhibitor 90 (HSP90); 3) a phospho-inositide 3-kinase inhibitor (PISK) and / or rapamycin target (mMTOR); 4) a cytochrome P450 inhibitor (for example, a CYP17 inhibitor or a 17alpha-hydroxylase / C17-20 lyase inhibitor); 5) an iron chelating agent; 6) an aromatase inhibitor; 7) a p53 inhibitor, for example, an inhibitor of a p53 / Mdm 2 interaction; 8) an apoptosis inducer; 9) an angiogenesis inhibitor; 10) an aldosterone synthase inhibitor; 11) a smoothed receptor inhibitor (SMO); 12) a prolactin receptor inhibitor (PRLR); 13) a Wnt signaling inhibitor; 14) an inhibitor
[00197] [00197] In some embodiments, the anti-CD73 antibody molecule is administered in combination with a PD-1 inhibitor. In some embodiments, the PD-1 inhibitor is selected from the group consisting of PDROO01, Nivolumab, Pembrolizumab, Pidilizumab, MEDIO680, REGN2810, TSR-042, PF-O06801591 and AMP-224. In some embodiments, the PD-1 inhibitor is an anti-PD-1 antibody molecule. In some embodiments, the anti-PD-1 antibody molecule is administered, for example, intravenously, at a dose of about 250 mg to 350 mg, about 350 mg to 450 mg or about 450 mg to 550 mg , for example, at a dose of about 300 mg or about 400 mg, for example, once every three weeks (Q3W) or once every four weeks (Q4W), for example, at a dose of about 300 mg Q3W or at a dose of about 400 mg Q4W.
[00198] [00198] In some embodiments, the anti-HIV antibody molecule
[00199] [00199] In some embodiments, the anti-CD73 antibody molecule is administered in combination with a CTLA4 inhibitor. In some embodiments, the CTLA-4 inhibitor is Ipilimumab or Tremalimumab.
[00200] [00200] In some embodiments, the anti-CD73 antibody molecule is administered in combination with a TIM-3 inhibitor. In some embodiments, the TIM-3 inhibitor is chosen from MGB453, TSR-022 or LY3321367.
[00201] [00201] In some embodiments, the anti-CD73 antibody molecule is administered in combination with a LAG-3 inhibitor. In some embodiments, the LAG-3 inhibitor is selected from the group consisting of LAG525, BMS-986016, TSR-033, MK-4280 and REGN3767.
[00202] [00202] In some embodiments, the anti-CD73 antibody molecule is administered in combination with a GITR agonist. In some embodiments, the GITR agonist is selected from the group consisting of GWN323 (Novartis), BMS-986156 (BMS), MK-4166 or MK-1248 (Merck), TRX518 (Leap Therapeutics), IN-CAGN1876 ( Incyte / Agenus), AMG 228 (Amgen) and INBRX-110 (Inhibrx).
[00203] [00203] In some embodiments, the anti-CD73 antibody molecule is administered in combination with a multispecific anti-CD3 antibody molecule. In some embodiments, the anti-CD3 multispecific antibody molecule is a bispecific anti-CD3 x anti-CD123 antibody molecule (for example, XENP14045), or a bispecific anti-CD3 x anti-CD123 antibody molecule CD20 (for example, XENP13676).
[00204] [00204] In some embodiments, the anti-CD73 antibody molecule is administered in combination with a cytokine molecule. In some embodiments, the cytokine molecule is IL-15 complexed with a soluble form of the IL-15 alpha receptor (IL-15Ra).
[00205] [00205] In some embodiments, the anti-CD73 antibody molecule is administered in combination with a STING agonist.
[00206] [00206] In some embodiments, the anti-CD73 antibody molecule is administered in combination with a macrophage colony stimulating factor inhibitor (M-CSF), optionally where the M-CSF inhibitor is MCS110.
[00207] [00207] In some embodiments, the anti-CD73 antibody molecule is administered in combination with a CSF-1R inhibitor, optionally wherein the CSF-1R inhibitor is BLZ945.
[00208] [00208] In some embodiments, the anti-CD73 antibody molecule is administered in combination with an indolamine 2,3-dioxigenase (IDO) and / or 2,3-dioxigenase tryptophan (TDO) inhibitor.
[00209] [00209] In some embodiments, the anti-CD73 antibody molecule is administered in combination with a TGF-B inhibitor.
[00210] [00210] In some embodiments, the anti-CD73 antibody molecule is administered in combination with an adenosine APAR antagonist. In some embodiments, the adenosine A2AR antagonist is selected from the group consisting of PBF509, CPI444, AZD4635, Vipadenante, GBV-2034 and AB928. In some embodiments, the adenosine A2AR antagonist is selected from the group consisting of 5-bromo-2,6-di- (1H-pyrazol-1-yl) pyrimidine-4-amine; (S) -7- (5-methylfuran-2-i1) -3 - ((6 - ((((tetrahydrofuran-3-yl) Oxy) methyl) pyridin-2-yl) methyl) - 3H- [1 , 2,3] triazolo [4,5-d] pyrimidin-5-amine; (R) -7- (5-methylfuran-2-yl) -3 - ((6 - ((((tetrahydrofuran-3-yl) oxy) methyl) pyridin-2-yl) methyl) -3H- [1,2,3] triazolo [4,5-d] pyrimidin-5-amine or racemate thereof; 7- (5-methylfuran-2-yl) -3 - ((6 - ((((tetrahydrofuran-3-yl) oxy) Methyl) pyridin-2-
[00211] [00211] In some embodiments, the anti-CD73 antibody molecule is administered in combination with a PD-1 inhibitor and an adenosine A2AR antagonist. In some embodiments, the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 5 mg to 100 mg, about 100 mg to 500 mg, about 500 mg to 1,000 mg , about 1,000 mg to 1,500 mg, about 1,500 mg to 2,000 mg, about 2,000 mg to 2,500 mg, about 2,500 mg to 3,000 mg, about 3,000 mg to 3,500 mg, or about
[00212] [00212] In some embodiments, the anti-CD73 antibody molecule is administered in combination with a PD-L1 inhibitor and an adenosine A2AR antagonist.
[00213] [00213] In some embodiments, the anti-CD73 antibody molecule is administered in combination with a chimeric antigen receptor (CAR) T cell therapy. In some embodiments, T CAR cell therapy is CTLO019.
[00214] [00214] In some embodiments, the anti-CD73 antibody molecule is administered in combination with one or more agents disclosed in Table 18, for example, one or more of: 1) a protein C kinase inhibitor (PKC) ; 2) a heat shock protein inhibitor 90 (HSP90); 3) an inhibitor of a phosphoinositide 3-kinase (PI3K) and / or rapamycin target (mMTOR); 4) a cytochrome P450 inhibitor (for example, a CYP17 inhibitor or a 17alpha-hydroxylase / C 17-20 lyase inhibitor); 5) an iron chelating agent; 6) an aromatase inhibitor; 7) a p53 inhibitor, for example, an inhibitor of a p53 / Mdm 2 interaction; 8) an apoptosis inducer; 9) an angiogenesis inhibitor; 10) an aldosterone synthase inhibitor; 11) a smoothed receptor inhibitor (SMO); 12) a prolactin receptor inhibitor (PRLR); 13) a Wnt signaling inhibitor; 14) a CDK4 / 6 inhibitor; 15) a fibroblast growth factor 2 receptor (FGFR2) / fibroblast growth factor 4 (FGFR4) receptor; 16) a macrophage colony stimulating factor inhibitor (M-CSF); 17) an inhibitor of one or more of c-KIT, histamine release, FIt3 (for example, FLK2 / STK1) or PKC; 18) an inhibitor of one or more of VEGFR-2 (for example, FLK-1 / KDR), PDGFRbeta, c-KIT or Raf kinase C; 19) a somatostatin agonist inhibitor and / or a growth hormone release; 20) an anaplastic lymphoma kinase inhibitor (ALK); 21) an insulin-like growth factor receptor inhibitor (IGF-1R); 22) a P-glycoprotein 1 inhibitor; 23) an inhibitor of vascular endothelial growth factor receptor (VEGFR); 24) a BCR-ABL kinase inhibitor; 25) an FGFR inhibitor; 26) a CYP11B2 inhibitor; 27) an inhibitor of HDM 2, for example, an inhibitor of HDM2-p53 interaction; 28) a tyrosine kinase inhibitor; 29) a c-MET inhibitor; 30) a JAK inhibitor; 31) a DAC inhibitor; 32) an 11B-hydroxylase inhibitor; 33) an IAP inhibitor; 34) a PIM kinase inhibitor; 35)
[00215] [00215] In certain embodiments, cancer treated with the anti-CD73 antibody molecule, alone or in combination with a second procedure or therapeutic agent, includes, but is not limited to, a solid tumor, a hematological cancer (e.g., leukemia, lymphoma, myeloma, for example, multiple myeloma) and a metastatic lesion. In some modalities, cancer is chosen from lung cancer (eg, non-small cell lung cancer), pancreatic cancer (eg, pancreatic ductal adenocarcinoma), breast cancer (eg, triple negative breast cancer), melanoma, head and neck cancer (eg, scaly head and neck cancer), colorectal cancer (eg, microsatellite stable colorectal cancer (MSS)), ovarian cancer or kidney cancer (for example, renal cell carcinoma). In certain embodiments, the anti-CD73 antibody molecule, used alone or in combination with a second procedure or therapeutic agent, controls tumor growth, reduces metastasis and / or improves survival.
[00216] [00216] In certain embodiments, the antibody molecule is administered at a dose of about 100 mg to 1,600 mg, about 100 mg at
[00217] [00217] In certain embodiments, the antibody molecule is administered
[00218] [00218] Still further, this disclosure provides methods for enhancing an immune response to an antigen in an individual, which comprises administering to the individual: (i) the antigen; and (ii) an anti-CD73 antibody molecule disclosed herein, so that an immune response to the antigen in the individual is enhanced. The antigen can be, for example, a tumor antigen, a viral antigen, a bacterial antigen, or a pathogen antigen.
[00219] [00219] The anti-CD73 antibody molecule can be administered to the individual systemically (for example, orally, parenterally, subcutaneously, intravenously, rectally, intramuscularly, intraperitoneally, intranasally, transdermally or by inhalation or intracavitary installation) or locally. In one embodiment, the anti-CD73 antibody molecule is administered intravenously.
[00220] [00220] The anti-CD73 antibody molecule can be used alone in unconjugated form or can be attached to a substance, for example, a cytotoxic agent or chemical portion (for example, a therapeutic drug; a compound that emits radiation; molecules of plant, fungal or bacterial origin, or a biological protein (for example, a protein toxin) or particle (for example, a recombinant viral particle, for example, by means of a viral coating protein). anti-CD73 antibody can be coupled to a radioactive isotope such as an emitter of a, B or y, or an emitter of B and y.
[00221] [00221] The dosages and therapeutic regimens of the anti-CD73 antibody molecule can be determined by a skilled person.
[00222] [00222] In another aspect, a method for detecting CD73 in a biological sample or in an individual is provided. In a modality, the method comprises (i) placing the sample or the individual (and, optionally, a sample or reference individual) in contact with the antibody molecule according to any one of claims 1 to 57 under conditions that allow the interaction of the antibody molecule and CD73 to occur, and (ii) detect the formation of a
[00223] [00223] The antibody molecules described in this document are preferred for use in the methods described in this document, although other anti-CD73 antibodies can be used instead, or in combination with an anti-CD73 antibody molecule from invention. BRIEF DESCRIPTION OF THE DRAWINGS
[00224] [00224] Figure 16 is a graph showing the binding of anti-CD73 antibodies or an isotype control antibody to CD8 + T cells, measured by flow cytometry. The MFI values are plotted against antibody concentrations. The tested antibodies are anti-CD73 antibodies 350, 356, 358, 373, 374, 377 and 379, as well as an isotype control antibody, all expressed in .B format.
[00225] [00225] Figures 2A and 2B are graphs showing results of a malachite green inorganic phosphate assay that tests the ability of the anti-CD73 antibody to inhibit the human CD73-mediated conversion of adenosine monophosphate (AMP) into adenosine . The release rate of inorganic phosphate (Pi) is plotted against tested AMP concentrations. In Figure 2A, the recombinant human CD73 was incubated with the AMP substrate with buffer alone ("hCD73 Km") or in the presence of an I9G1 isotype control antibody ("ISO.C"). In Figure 2B, the recombinant CD73 was incubated with AMP with buffer alone ("hCD73 Km") or in the presence of anti-CD73 350.C antibody at the indicated concentrations ("1 µg / ml 350.C", "0.3 µg / ml
[00226] [00226] Figures 3A, 3B and 3C are graphs that show results from a malachite green phosphate (MG) assay that tests the ability of the anti-CD73 antibody to inhibit the enzymatic activity of CD73 human or recombinant soluble cinomolg. The formation of% INH phosphate is plotted against concentrations of anti-CD73 antibody for studies using recombinant human CD73 (Figures 3A and 3B) or CD73 cinomolgo (Figure 3C). The tested antibodies are anti-CD73 antibodies 350, 356, 373 and 374, expressed in .A or .B format.
[00227] [00227] Figure 4 is a graph showing results of a modified Cell Titration Glo (CTG) assay that examines the enzyme inhibiting activity of anti-CD73 antibodies against CD73 spilled from a cell line of MDA-MB- breast cancer
[00228] [00228] Figure 5 is a graph showing results of a modified Cell Titration Glo (CTG) assay that examines the ability of anti-CD73 antibodies to inhibit the enzymatic activity of CD73 in the serum of a patient of pancreatic cancer. AMP conversion of% INH is plotted against concentrations of anti-CD73 antibody. The anti-CD73 antibodies tested are 350, 356, 358, 373, 374, 377 and 379, all expressed in .B format.
[00229] [00229] Figure 6 is a graph showing the inhibition of CD73 expressed on the surface of a breast cancer cell line MDA-MB-231, measured using a malachite green phosphate assay. AMP conversion of% INH is plotted over a range of anti-CD73 antibody concentrations. The anti-CD73 antibodies tested are 350, 356, 358, 373, 374, 377 and 379, all in the .B format.
[00230] [00230] Figures 7A, 7B, 7C and 7D are bar graphs showing results of a Cell Titration Glo (CTG) assay
[00231] [00231] Figures 8A and 8B are graphs showing the enzyme inhibiting activity of anti-CD73 antibodies against surface CD73 expressed in a human breast cancer cell line MDA-MB-231 or a line SKOV3 human ovarian cancer cell, measured by a modified Cell Titration Glucose (CTG) assay. % INH is plotted against concentrations of anti-CD73 antibody. The anti-CD73 antibodies tested are 350 and 373, expressed in .A or .B format.
[00232] [00232] Figures 9A and 9B are graphs similar to Figures 8A and 8B. The tested antibodies are anti-CD73 antibodies 350, 356, 373 and 374, in .A or .B format.
[00233] [00233] Figure 10 is a graph showing the inhibition of overexpressed human CD73 in HEK 293 cells by anti-CD73 antibodies, as measured by a modified Cell Titration (CTG) assay. AMP conversion of% INH is plotted against a range of anti-CD73 antibody concentrations. The tested antibodies are anti-CD73 antibodies 350, 356, 373 and 374, in .A or .B format.
[00234] [00234] Figures 11A and 11B are graphs showing the inhibition of CD73 expressed in primary human PBMCs isolated from two donors separated by anti-CD73 antibodies, measured by a modified Glo Cell Titration (CTG) assay. % INH is plotted against a range of anti-CD73 antibody concentrations. The anti-CD73 antibodies tested are 350, 356 and 358, all in .B format.
[00235] [00235] Figures 12A and 12B are graphs showing the proliferation of stimulated anti-CD3 / 28 CD4 + T cells in the presence of AMP and an anti-CD73 antibody. The proliferation index, a measure of T cell division, is plotted against a range of antibody concentrations. Figure 12A shows results of a study that tests anti-CD73 antibodies 350, 356, 358, 374, 377 and 379, all in the .B format. Figure 12B shows results of a study using antibodies 350 and 372, expressed in .A or .B format.
[00236] [00236] Figure 13 is a bar graph showing the results of a xenograft study that tests the ability of the anti-CD73 antibody to inhibit the enzymatic activity of CD73 in vivo. The y-axis shows levels of adenosine and inosine in the serum of immunocompromised mice implanted with a breast cancer cell line with high CD73 expression (MDA-MB-231) as measured by mass spectrometry. The anti-CD73 antibodies 350, 356, 373 and 374, expressed in .A or .B format, were administered intraperitoneally to 20 or 200 pug / mice. The control polyclonal human IgG antibody was administered to 200 upug / mouse.
[00237] [00237] Figure 14 is a graph showing the comparison of protection profiles of 373.A and 373.B, as measured by fragmentation hydrogen deuterium exchange mass spectrometry (HDx-MS). Figure 14 shows results for 1 min in exchange at pH 7.5 and room temperature.
[00238] [00238] Figure 15 is a graph showing the comparison of protection profiles of 350.A2 and 350.B, as measured by fragmentation hydrogen deuterium exchange mass spectrometry (HDx-MS). Figure 15 shows results for 1 min in exchange at pH 7.5 and room temperature.
[00239] [00239] Figure 16A is a graph showing the CD73 conformational exchange between the closed / active (Protein Database (PDB) 4H2I) and open / inactive (PDB 4H2F) conformation. Figure 16B is a graph showing the open / open conformation of CD73 dimer constructed from two PDB 4H2F units after alignment of the c-terminal domains with PDB 4H1S.
[00240] [00240] Figures 17A and 17B are graphs showing SEC profiles of the CD73-373.A and CD73-373.B complexes, respectively.
[00241] [00241] Figures 18A and 18B are graphs showing SEC profiles of the CD73-350.A2 and CD73-350.B complexes, respectively.
[00242] [00242] Figure 19 is a graph showing the relative percentages of CD73 / mAb species calculated using the integration method.
[00243] [00243] Figure 20 is a schematic graph showing an oligomerization model to unify HDx and SEC interpretations.
[00244] [00244] Figures 21A and 21B are graphs showing the expression of CD73 in stromal cell subsets analyzed by flow cytometry in 4T1 tumors from mice treated with an anti-TGFB pan antibody or hlgG2 isotype control . In Figure 21A, the expression of CD73 is displayed as fluorescence intensity, compared to samples stained with a compatible isotype control. A representative sample is shown. In Figure 21B, the quantification of CD73 expression, measured as mean fluorescence intensity, is represented. Each symbol represents a sample. P values are indicated where it is significant. CAF = cancer-associated fibroblasts.
[00245] [00245] Figures 22A, 22B and 22C are graphs showing results
[00246] [00246] Figure 23 is a panel of graphs showing individual tumor volumes for the indicated groups. The isotype control mice were euthanized on Day 25. In the anti-PD-1 treated group, the mice received 300 µg / mouse for all doses. In the treated group of 350.B, the mice received 600 ug / mouse for the first dose, followed by 400 upg / mouse for the remaining four doses. The same dosing schedules were administered for the combination group. All mice were treated on Days 2, 5, 9, 12 and 17.
[00247] [00247] Figure 24A is a pair of graphs showing the detection of 373.A biotinylated in pre-treated blood samples from
[00248] [00248] Figures 25A and 25B are a graphical panel showing that the proliferation of CD4 + and CD8 + T cells was suppressed by AMP during TCR-mediated activation, and that suppression can be restored by antibody 373.A.
[00249] [00249] Figures 26A and 26B show CD73 residues (bold, italic and underlined with a line) that interact with the 350.A2 heavy Fab chain through direct enthalpic interactions (Y110, L132, K136, S155 , L157 and K162, numbered according to SEQ ID NO: 105) (Figure 26A) or hydrophobic interactions and Van der Waal (residues 136 to 138 and 155 to 170 numbered according to SEQ ID NO: 105) (Figure 26B ), as measured in Example 8. Figures 26C and 26D show the Fab heavy chain of 350.A2 residues (bold, italic and underlined with a line) that interact with CD73 through direct enthalpic interactions (R31, R54 , E95, E98 and S99, numbered according to Kabat numbering; or R31, R54, E98, E101 and S102, numbered according to their linear positions in SEQ ID NO: 331) (Figure 26C) or hydrophobic and Van interactions der Waal (residues 30, 31, 33, 50, 52, 56, 97, 98, 100, and 100a, numbered according to Kabat numbering; or residues 30, 31, 33, 50, 52, 56, 100, 101, 103 , and 104, numbered according to their linear positions in SEQ ID NO: 331) (Figure 26D), as measured in Example 8. Figures 26E and 26F show CD73 residues (bold, italic and underlined with a line) that interact with a 350.A2 Fab light chain through direct enthalpic interactions (T209, numbered according to SEQ ID NO: 105) (Figures 26E) or hydrophobic and Van der Waal interactions (residues 209 and 210, numbered according with SEQ ID NO: 105) (Figure 26F), as measured in Example 8. Figures 26G and 26H show 350.A2 Fab light chain residues (bold, italic and underlined with a line) that interact with CD73 through direct enthalpic interactions (W32, numbered according to
[00250] [00250] Figures 27A and 27B are graphs that show the difference between the identification ratios observed for CD73 / mAb and CD73 complexes alone after an identification pulse of 30 s.
[00251] [00251] Figures 28A and 28B are graphs that show the difference between the identification ratios observed for CD73 / mAb and CD73 complexes alone after identification for 300s. BRIEF DESCRIPTION OF THE TABLES
[00252] [00252] Table 1 provides amino acid and nucleotide sequences for exemplary anti-CD73 antibodies.
[00253] [00253] Table 2 provides consensus CDR sequences for exemplary anti-CD73 antibodies.
[00254] [00254] Table 3 provides amino acid sequences of human IgG heavy chains and human kappa light chain.
[00255] [00255] Table 4 provides exemplary CD73 sequences.
[00256] [00256] Tables 5 and 6 provide amino acid and / or nucleotide sequences of exemplary anti-PD-1 antibody molecules.
[00257] [00257] Tables 7 and 8 provide exemplary amino acid and / or nucleotide sequences of anti-PD-L1 antibody molecules.
[00258] [00258] Tables 9 and 10 provide exemplary amino acid and / or nucleotide sequences of anti-LAG-3 antibody molecules.
[00259] [00259] Tables 11 and 12 provide exemplary amino acid and / or nucleotide sequences of anti-TIM-3 antibody molecules.
[00260] [00260] Tables 13 and 14 provide amino acid and / or nucleotide sequences of exemplary anti-GITR antibody molecules.
[00261] [00261] Table 15 provides amino acid sequences of exemplary bispecific anti-CD3 antibody molecules.
[00262] [00262] Tables 16 and 17 provide exemplary IL15 / IL-15Ra amino acid sequences.
[00263] [00263] Table 18 is a summary of selected therapeutic agents that can be administered in combination with the anti-CD73 antibody molecules described in this document. Table 18 provides, from left to right, the following: the Compound Designation of the second therapeutic agent, the Compound and Patent Publication (or Publications) structure that discloses the Compound.
[00264] [00264] Table 19 provides nomenclatures for two strains of anti-CD73 antibodies.
[00265] [00265] Table 20 provides affinities for anti-CD73 antibodies.
[00266] [00266] Table 21 provides affinities for anti-CD73 Fabs.
[00267] [00267] Table 22 provides provisional dose levels for 373.A.
[00268] [00268] Table 23 provides provisional dose levels for 373.A in combination with PBF509.
[00269] [00269] Table 24 provides provisional dose levels for 373.A in combination with BAPO49-Clone-E.
[00270] [00270] Table 25 provides provisional dose levels for PBF509 in combination with 373.A and BAPO49-Clone-E.
[00271] [00271] Table 26 shows corresponding germline anti-CD73 antibodies. DETAILED DESCRIPTION
[00272] [00272] The term "CD73" as used in this document refers to "Grouping of Differentiation 73", also known as
[00273] [00273] Without wishing to stick to the theory, the human CD73 has two domains. A conserved N-terminal domain (roughly corresponding to residues 29 to 310 of SEQ ID NO: 105) and a conserved C-terminal domain (roughly corresponding to residues 343 to 513 of SEQ ID NO: 105), which they are linked by a single a-helix (approximately corresponding to residues 318 to 336 of SEQ ID NO: 105). The active site is detected primarily in closed conformation and is formed between C-terminal and N-terminal domains. For enzyme catalysis, a motion of -100º domain of the N-terminal domain in relation to the C-terminal domain can enable substrate binding and release, which occurs in the open (catalytically inactive) conformation. Human CD73 forms a dimer through protein-protein interactions between C-terminal domains. The buried surface area, as well as the molecular interactions at the dimer interface, are significantly different between active and inactive conformations of the enzyme. See, for example, Knapp K, et al., Structure 20: 2161-73 (2012), incorporated in this document for reference in its entirety.
[00274] Consequently, the present invention provides, at least in part, antibody molecules that bind to CD73 with high affinity and specificity. In one embodiment, human antibodies that bind to CD73 are disclosed in this document. In one embodiment, antibody molecules that are capable of inhibiting or reducing the enzymatic activity of CD73, for example, human CD73, for example, soluble human CD73 or membrane-bound human CD73, are disclosed herein. In one embodiment, antibody molecules that are capable of inhibiting or reducing the CD73-mediated conversion of adenosine monophosphate (AMP) to adenosine are disclosed herein. Additional aspects of the invention include nucleic acid molecules that encode antibody molecules, expression vectors, host cells and methods for producing the antibody molecules. Immunoconjugates, multispecific or bispecific molecules and pharmaceutical compositions comprising the antibody molecules are also provided. The anti-CD73 antibody molecules disclosed herein may be used to treat, prevent and / or diagnose cancerous or malignant disorders, for example, solid and liquid tumors, for example, lung cancer (for example, cancer non-small cell lung cancer), pancreatic cancer (eg pancreatic ductal adenocarcinoma), breast cancer (eg triple negative breast cancer), melanoma, head and neck cancer (eg head cancer and scaly neck
[00275] [00275] Additional terms are defined below and throughout the order.
[00276] [00276] As used in this document, the articles "one" and "one" refer to one or more than one (for example, at least one) of the grammatical object of the article. As used in this document, "plurality" means two or more.
[00277] [00277] The term "or" is used in this document to mean, and is used interchangeably with, the term "and / or", unless the context clearly indicates otherwise.
[00278] [00278] "About" and "approximately" will generally mean an acceptable degree of error for the measured quantity given the nature or accuracy of the measurements. Exemplary degrees of error are within 20 percent (%), typically within 10% and, more typically, within 5% of a given value or range of values.
[00279] [00279] The compositions and methods disclosed in this document encompass polypeptides and nucleic acids that have the specified sequences or sequences substantially identical or similar to them, for example, sequences that are at least about 85%, 90 % or 95% sequence identity to the specified sequence. In the context of an amino acid sequence, the term "substantially identical" is used in this document to refer to a first amino acid that contains a sufficient or minimal number of amino acid residues that are i) identical to, or ii) substitutions conservative amino acid residues aligned in a second amino acid sequence so that the first and second amino acid sequences can have a common structural domain and / or common functional activity. For example, amino acid sequences that contain a common structural domain that is at least about 85%, 90%. 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity with a reference sequence, for example, a sequence provided in this document.
[00280] [00280] In the context of nucleotide sequences, the term "substantially identical" is used throughout this document to refer to a first nucleic acid sequence that contains a sufficient or minimal number of nucleotides that are identical to nucleotides aligned in a second nucleic acid sequence so that the first and second nucleotide sequences encode a polypeptide that has common functional activity or encode a common structural polypeptide domain or a common functional polypeptide activity. For example, nucleotide sequences that have at least about 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity with a reference sequence, for example, a sequence provided in this document.
[00281] [00281] The term "functional variant" refers to polypeptides that have an amino acid sequence substantially identical to the naturally occurring sequence, or are encoded by a substantially identical sequence of nucleotides, and are capable of having one or more activities of the naturally occurring sequence.
[00282] [00282] The calculations of homology or sequence identity between sequences (the terms are used interchangeably in this document) are performed as follows.
[00283] [00283] To determine the percent identity of two amino acid sequences, or two nucleic acid sequences, the sequences are aligned for optimal comparison purposes (for example, gaps can be introduced in one or both of the first and a second sequence of amino acids or nucleic acids for optimal alignment and non-homologous sequences can be discarded for comparison purposes). In a preferred embodiment, the length of an aligned reference sequence for comparison purposes is at least 30%, for example, at least 40%, 50%, 60%, for example, at least 70%, 80 %, 90%, 100% of the length of the reference sequence. Amino acid or nucleotide residues at corresponding amino acid positions or nucleotide positions are then compared. When a position in the first sequence is occupied by the same amino acid or nucleotide residue as the corresponding position in the second sequence, then the molecules are identical in that position.
[00284] [00284] The percentage identity between the two sequences is a function of the number of identical positions shared by the sequences, considering the number of gaps, and the length of each gap, which needs to be introduced for optimal alignment of the two sequences.
[00285] [00285] The comparison of sequences and the determination of the percent identity between two sequences can be performed with the use of a mathematical algorithm. In some modalities, the percentage of identity between two amino acid sequences is determined using the Needleman and Wunsch algorithm ((1970) J. Mol. Biol. 48: 444 to 453) that was incorporated into the GAP program in the GCG software package (available at http://Www.gcg.com), using a Blossum 62 matrix or a PAM250 matrix, and a span weight of 16, 14, 12, 10 , 8, 6 or 4 and a length weight of 1, 2, 3, 4, 5 or 6. In certain embodiments, the percentage of identity between two nucleotide sequences is determined using the GAP program in the GCG software package (available at http: /Awww.gceg.com), using an NWSgapdna.CMP matrix, and a span weight of 40, 50, 60, 70 or 80 and a length weight of 1, 2, 3, 4, 5 or 6. An appropriate set of parameters (and the one that should be used, unless otherwise specified) is a Blossum 62 score matrix with a span penalty of 12, a penalty span span of 4 and a frame shift span penalty of 5.
[00286] [00286] The percentage identity between two sequences of amino acids or nucleotides can be determined using the algorithm of E. Meyers and W. Miller ((1989) CABIOS, 4:11 to 17) that was incorporated into the program ALIGN (version 2.0), using a PAM120 weight residual table, a gap length penalty of 12 and a gap penalty of 4.
[00287] [00287] The nucleic acid and protein sequences described in this document can be used as a "query sequence" to conduct a search against public databases to, for example, identify other family members or related sequences . Such searches can be carried out using the NBLAST and XBLAST (version 2.0) programs by Altschul, et a /. (1990) J. Mol. Biol. 215: 403-10. BLAST nucleotide searches can be performed with the NBLAST program, score = 100, word length = 12 to obtain nucleotide sequences homologous to a nucleic acid as described in this document. BLAST protein searches can be performed with the XBLAST program, punctuation = 50, word length = 3 to obtain amino acid sequences homologous to the protein molecules described in this document. To obtain gap alignments for comparison purposes, Gapped BLAST can be used as described in Altschul et a /., (1997) Nucleic Acids Res. 25: 3,389 a
[00288] [00288] “As used in this document, the term" hybridizes under conditions of low stringency, medium stringency, high stringency or very high stringency "describes the conditions for hybridization and washing. Guidance for carrying out hybridization reactions can be found in Current Protocols in Molecular Biology, John Wiley & Sons, N.Y. (1989), 6.3.1 to 6.3.6, which is incorporated by reference. Aqueous and non-aqueous methods are described in that reference and any one of them can be used. The specific hybridization conditions mentioned in this document are as follows: 1) the low stringency hybridization conditions in 6X sodium chloride / sodium citrate (SSC) at about 45 ºC, followed by two washes in 0 , 2X SSC, 0.1% SDS at least at 50 ºC (the washing temperature can be increased to 55 ºC for conditions of low rigor); 2) hybridization conditions of medium stringency in 6X SSC at about 45 ºC, followed by one or more washes in 0.2X SSC, 0.1% SDS at 60 “ºC; 3) hybridization conditions high stringency in 6X SSC at about 45 ºC, followed by one or more washes in 0.2X SSC, 0.1% SDS at 65 ºC; and preferably 4) very high stringency hybridization conditions are 0.5M sodium phosphate, 7% SDS at 65 ºC, followed by one or more washes in 0.2X SSC, 1% SDS at 65 ”ºC . Very strict conditions (4) are suitable conditions and those that should be used unless otherwise specified.
[00289] [00289] It is understood that the molecules of the invention may have
[00290] [00290] The term "amino acid" is intended to encompass all molecules, regardless of whether they are natural or synthetic, which includes both an amino functionality and an acidic functionality and capable of being included in a naturally occurring polymer of amino acids. Exemplary amino acids include naturally occurring amino acids; analogues, derivatives and the like; amino acid analogs that have variant side chains; and all stereoisomers of any of the above. As used herein, the term "amino acid" includes both D- or L-optical isomers and peptidomimetics.
[00291] [00291] A "conservative amino acid substitution" is one in which the amino acid residue is replaced by an amino acid residue that has a similar side chain. Families of amino acid residues that have similar side chains have been defined in the specialty. These families include amino acids with basic side chains (eg, lysine, arginine, histidine), acidic side chains (eg, aspartic acid, glutamic acid), uncharged polar side chains (eg, glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine), non-polar side chains (eg, alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, tryptophan), beta-branched side chains (eg, treinein, valine, isoleucine ) and aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, histidine).
[00292] [00292] The terms "polypeptide", "peptide" and "protein" (if there is a single chain) are used interchangeably in the present document to refer to amino acid polymers of any length. The polymer can be linear or branched, it can comprise modified amino acids and can be disrupted by non-amino acids. The terms also encompass a modified amino acid polymer; for example, disulfide bond formation, glycosylation, lipidation, acetylation, phosphorylation or any other manipulation, such as conjugation with an identification component. The polypeptide can be isolated from natural sources, it can be one produced by recombinant techniques from a eukaryotic or prokaryotic host, or it can be a product of synthetic procedures.
[00293] [00293] The terms "nucleic acid", "nucleic acid sequence", "nucleotide sequence" or "polynucleotide sequence" and "polynucleotide" are used interchangeably. They refer to a polymeric form of nucleotides of any length, deoxyribonucleotides or ribonucleotides, or analogues thereof. The polynucleotide can have a single strand or a double strand, and if the single strand can be the coding strand or non-coding (antisense) strand. A polynucleotide can comprise modified nucleotides, such as methylated nucleotides and nucleotide analogues. The nucleotide sequence can be disrupted by non-nucleotide components. A polynucleotide can be further modified after polymerization, as in conjunction with an identification component. The nucleic acid can be a recombinant polynucleotide, or a polynucleotide of genomic origin, cDNA, semisynthetic or synthetic, which does not occur in nature or is bound to another polynucleotide in an unnatural arrangement.
[00294] [00294] The term "isolated" as used here refers to material that is removed from its original or native environment (for example, the natural environment if it occurs naturally). For example, a naturally occurring polynucleotide or polypeptide in a living animal is not isolated, but the same polynucleotide or polypeptide, separately
[00295] [00295] The term "Chimeric Antigen Receptor" or alternatively a "CAR" relates to a set of polypeptides, typically two in the simplest modalities, which, when in an immune effector cell, endow the cell with specificity for a target cell, typically a cancer cell, with intracellular signal generation. In some embodiments, a CAR comprises at least an extracellular antigen binding domain, a transmembrane domain and a cytoplasmic signaling domain (also referred to herein as "an intracellular signaling domain") that comprises a functional signaling derived from a stimulating molecule and / or co-stimulating molecule as defined below. In some respects, the set of polypeptides are contiguous to each other. In some embodiments, the set of polypeptides includes a dimerization switch that, in the presence of a dimerization molecule, can couple the polypeptides together, for example, it can couple an antigen-binding domain to an intracellular signaling domain. In one aspect, the stimulating molecule is the zeta chain associated with the T cell receptor complex. In one aspect, the cytoplasmic signaling domain additionally comprises one or more functional signaling domains derived from at least one molecule co-stimulator as defined below. In one aspect, the co-stimulatory molecule is chosen from the co-stimulatory molecules described here, for example, 4-1BB (i.e., CD137), CD27 and / or CD28. In one aspect, the CAR comprises a chimeric fusion protein comprising an extracellular antigen binding domain, a transmembrane domain and an intracellular signaling domain comprising a functional signaling domain derived from a stimulator molecule. In one aspect, the CAR comprises a chimeric fusion protein comprising an extracellular antigen-binding domain, a trans-membrane domain and an intracellular signaling domain comprising a functional signaling domain derived from a co-stimulating molecule and a signaling domain functional derivative of a stimulating molecule. In one aspect, the CAR comprises a chimeric fusion protein comprising an extracellular antigen binding domain, a transmembrane domain and an intracellular signaling domain comprising two functional signaling domains derived from one or more molecules (or molecules) co-stimulating and a functional signaling domain derived from a stimulating molecule. In one aspect, the CAR comprises a chimeric fusion protein comprising an extracellular antigen binding domain, a transmembrane domain and an intracellular signaling domain comprising at least two functional signaling domains derived from one or more more co-stimulating molecule (s) and a functional signaling domain derived from a stimulating molecule. In one aspect, the CAR comprises an optional leader sequence at the amino (N-ter) terminus of the CAR fusion protein. In one aspect, the CAR further comprises a leader sequence at the N-terminal of the extracellular antigen binding domain, wherein the leader sequence is optionally cleaved from the antigen binding domain (e.g., an scFv) during cell processing and localization of CAR on the cell membrane.
[00296] [00296] The term "signaling domain" relates to the functional portion of a protein that acts by transmitting information
[00297] [00297] An "intracellular signaling domain", as the term is used here, refers to an intracellular portion of a molecule. The intracellular signaling domain generates a signal that promotes an immune effector function of the cell containing the CAR, for example, a CART cell. Examples of immune effector function, for example, in a CART cell, include cytolytic activity and helper activity, including cytokine secretion.
[00298] [00298] In one embodiment, the intracellular signaling domain may comprise a primary intracellular signaling domain. Exemplary primary intracellular signaling domains include those derived from molecules responsible for the primary stimulus or antigen-dependent stimulus. In one embodiment, the intracellular signaling domain may comprise a co-stimulating intracellular domain. Exemplary co-stimulating intracellular signaling domains include those derived from molecules responsible for co-stimulating signals, or antigen independent stimuli. For example, in the case of a CART, a primary intracellular signaling domain may comprise a cytoplasmic sequence of a T cell receptor, and a co-stimulating intracellular signaling domain may comprise a co-stimulating or co-stimulating molecule cytoplasmic sequence.
[00299] [00299] “A primary intracellular signaling domain may comprise a signaling motif that is known as the tyrosine-based activation motif of the immunoreceptor or ITAM. Examples of primary cytoplasmic signaling sequences containing ITAMs include, but are not limited to, those derived from CD3 zeta, FcR common range (FCER1G), Fc gamma Rlla, FcR beta (Fc Epsilon
[00300] [00300] The term "zeta" or alternatively "zeta chain", "CD3-zeta" or "TCR-zeta" is defined as the protein provided as ac number. GenBank BAG36664.1, or equivalent residues of a non-human species, for example, mouse, rodent, monkey, simian and the like, and a "zeta-stimulating domain" or alternatively a "CD3- stimulating domain" zeta "or a" TCR-zeta stimulating domain "is defined as the amino acid residues of the cytoplasmic domain of the zeta chain, or functional derivatives of the zeta chain, which are sufficient to functionally transmit a necessary initial signal for cell activation T. In one aspect, the cytoplasmic domain of zeta comprises residues 52 to 164 of ac number. GenBank BAG36664.1, or the equivalent residues of a non-human species, for example, mouse, rodent, monkey, simian and similar, which are functional orthologs of the same.
[00301] [00301] The term a "co-stimulating molecule" refers to a cognate binding partner in a T cell that binds to a co-stimulating ligand, thereby mediating a co-stimulating response by the T cell, as, but without limitations , proliferation. Co-stimulatory molecules are cell surface molecules different from antigen receptors or their ligands that contribute to an efficient immune response. Co-stimulatory molecules include, but are not limited to, a class MHC molecule, BTLA and a Toll ligand receptor, as well as OX40, CD27, CD28, CDS, ICAM-1, LFA-1 (CD11a / CD18), ICOS ( CD278) and 4-1BB (CD137). Additional examples of such co-stimulating molecules include CDS, ICAM-1, GITR, BAFFR, HVEM (LIGHTR), SLAMF7, NKp80 (KLRF1), NKp44, NKp30, NKp46, CD160, CD19, CD4, CD8alfa, CD8beta, IL2R beta, ILPR gamma, IL7R alpha, ITGAA4, VLA1, CD49a, ITGAA4, IA4, CD49D, ITGA6, VLA-6, CD49f, ITGAD,
[00302] [00302] A co-stimulating intracellular signaling domain can be the intracellular portion of a co-stimulating molecule. A co-stimulating molecule can be represented in the following protein families: TNF receptor proteins, Immunoglobulin type proteins, cytokine receptors, integrins, lymphocyte activation signaling molecules (SLAM proteins) and NK cell activating receptors . Examples of such molecules include CD27, CD28, 4-1BB (CD137), OX40, GITR, CD30, CD40, ICOS, BAFFR, HVEM, ICAM-1, antigen associated with lymphocyte-1 (LFA-1), CD2 function , CDS, CD7, CD287, LIGHT, NKG2C, NKG2D, SLAMF7, NKp80, NKp30, NKp44, NKp46, CD160, B7-H3 and a linker that binds to CD83 and the like.
[00303] [00303] The intracellular signaling domain may comprise the entire intracellular portion, or the entire native intracellular signaling domain, of the molecule from which it is derived, or a fragment or functional derivative thereof.
[00304] [00304] The term "4-1BB" refers to a member of the TNFR superfamily with an amino acid sequence provided as Gen-Bank Accession No. AAA62478.2, or the equivalent residues of a non-human species, for example, mouse, rodent, monkey, simian and the like; and a "4-1BB costimulatory domain" is defined as the amino acid residues 214-255 of GenBank Accession No. AAAG62478.2, or the equivalent residues of a non-human species.
[00305] [00305] —The "immune cell", as this term is used in this document, refers to a cell that is involved in an immune response, for example, in promoting an immune response. Examples of immune effector cells include T cells, for example, alpha / beta T cells and gamma / delta T cells, B cells, natural killer cells (NK), natural killer T cells (NKT), mast cells, and phagocytes derived from the myeloid lineage.
[00306] [00306] The "immuno-defective function" or "immuno-defective response", as such term is used in this document, refers to the function or response, for example, of an immuno-defective cell, which enhances or promotes an immune attack by a cell -target. For example, an immune function or effector response refers to a property of a T or NK cell that promotes the death or inhibition of the growth or proliferation of a target cell. In the case of a T cell, primary stimulation and costimulation are examples of an effective immune function or response.
[00307] [00307] The terms "cancer-associated antigen" or "tumor antigen" are indistinctly related to a molecule (typically a protein, carbohydrate or lipid) that is expressed on the surface of a cancer cell, either entirely or as a fragment (for example, MHC / peptide), and which is useful for the preferential targeting of a pharmacological agent to the cancer cell. In some embodiments, a tumor antigen is a marker expressed by both normal cells and cancer cells, for example, a lineage marker, for example, CD19 on B cells. In some embodiments, a tumor antigen is a surface molecule cell that is overexpressed in a cancer cell compared to a normal cell, for example, overexpression 1 time, overexpression 2 times, overexpression of
[00308] [00308] Various aspects of the compositions and methods in this document are described in greater detail below. Additional definitions are presented throughout the specification. Antibody Molecules
[00309] [00309] In one embodiment, the antibody molecule binds to a mammalian, for example, human CD73. For example, the antibody molecule binds to an epitope, for example, linear or conformational epitope, for example, an epitope as described in the present document, on CD73.
[00310] [00310] As used herein, the term "antibody molecule" refers to a protein, for example, an immunoglobulin chain or fragment thereof, comprising at least one immunoglobulin variable domain sequence. The term "antibody molecule" includes, for example, a monoclonal antibody (including a full-length antibody that has an immunoglobulin Fc region). In one embodiment, an antibody molecule comprises a full-length antibody or a full-length immunoglobulin chain. In one embodiment, an antibody molecule comprises an antigen-binding or functional fragment of a full-length antibody or a full-length immunoglobulin chain.
[00311] [00311] As used in this document, an antibody molecule "binds" to an antigen, since such a link is understood by a person skilled in the art. In one embodiment, an antibody binds to an antigen with a dissociation constant (Kp) of about 1 x 103 M or less, 1 x 10 ° M or less or 1 x 10 ° M or less.
[00312] [00312] In one embodiment, an antibody molecule is a monospecific antibody molecule and binds to a single epitope, for example, a monospecific antibody molecule that has a plurality of immunoglobulin variable domain sequences, each of which which binds to the same epitope.
[00313] [00313] In one embodiment, an antibody molecule is a multispecific antibody molecule, for example, comprises a plurality of immunoglobulin variable domain sequences, wherein a first immunoglobulin variable domain sequence among the plurality has binding specificity for a first epitope and a second immunoglobulin variable domain sequence within the plurality have binding specificity for a second epitope. In one embodiment, the first and second epitopes are on the same antigen, for example, the same protein (or subunit of a multimeric protein). In one embodiment, the first and second epitopes overlap or substantially overlap. In one embodiment, the first and second epitopes do not overlap or substantially do not overlap. In one embodiment, the first and second epitopes are on different antigens, for example, different proteins (or different subunits of a multimeric protein). In one embodiment, a multispecific antibody molecule comprises a third, fourth or fifth variable domain of immunoglobulin. In one embodiment, a multispecific antibody molecule is a bispecific antibody molecule, a triespecific antibody molecule, or a tetra-specific antibody molecule.
[00314] [00314] In one embodiment, a multispecific antibody molecule is a bispecific antibody molecule. A bispecific antibody has specificity for no more than two antigens. A bispecific antibody molecule is characterized by a first immunoglobulin variable domain sequence that has binding specificity for a first epitope and a second immunoglobulin variable domain sequence that has binding specificity for a second epitope. In one embodiment, the first and second epitopes are on the same antigen, for example, the same pro-
[00315] [00315] In one embodiment, an antibody molecule comprises a diabody and a single chain molecule, as well as an antigen-binding fragment of an antibody (for example, Fab, F (ab '): and Fv ). For example, an antibody molecule can include a heavy chain variable domain (H) sequence (abbreviated here as VH) and a light chain variable domain (L) sequence
[00316] [00316] Examples of antigen-binding fragments of an antibody molecule include: (i) a Fab fragment, a monovalent fragment consisting of the VL, VH, CL and CH1 domains; (ii) a fragment F (ab '), a divalent fragment comprising two Fab fragments connected by a disulfide bridge in the hinge region; (iii)
[00317] [00317] The term "antibody" includes intact molecules as well as functional fragments thereof. Antibody constant regions can be altered, for example., Mutated, to modify the properties of the antibody (for example, to increase or decrease one or more of: Fc receptor binding, antibody glycosylation, the number of cysteine, effector cell function or complement function).
[00318] [00318] The antibodies disclosed in this document can also be single domain antibodies. Single domain antibodies can include antibodies whose regions of complementary determination are part of a single domain polypeptide. Examples include, but are not limited to, heavy chain antibodies, antibodies naturally devoid of light chains, single domain antibodies derived from conventional 4 chain antibodies, genetically modified antibodies and single domain frameworks other than those derived from antibodies. Single domain antibodies can be any of the art, or any future single domain antibodies. Single domain antibodies can be derived from any species including, but not limited to, mice
[00319] [00319] The VH and VL regions can be subdivided into hypervariability regions called "complementarity determining regions" (CDR), interspersed with regions that are more conserved, called "framework regions" (FR or FW).
[00320] [00320] The extension of the framework and CDR region was precisely defined by a number of methods (see, Kabat, EA, et al. (1991) Sequences of Proteins of Immunological Interest, Fifth Edition, US Department of Health and Human Services, NIH Publication No. 91-3242; Chothia, C. et al. (1987) J. Mol. Biol. 196: 901-917; and the definition of ADM used by Ox-ford Molecular's ADM antibody modeling software. generally speaking, for example, Protein Sequence and Structure Analysis of Antibody Variable Domains In: Antibody Engineering Lab Manual (Ed .: Duebel, S. and Kontermann, R,., Springer-Verlag, Heidelberg).
[00321] [00321] The terms "complementarity determining region" and
[00322] [00322] The precise amino acid sequence limits of a given CDR can be determined using any of the well-known schemes, including those described by Kabat et al. (1991), "Sequences of Proteins of Immunological Interest", 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD ("Kabat" numbering scheme), Al-Lazikani et a., ( 1997) JMB 273,927 to 948 ("Chothia" numbering scheme). As used in this document, CDRs defined according to the "Chothia" numbering scheme are sometimes also called "hypervariable loops".
[00323] [00323] For example, under Kabat, the amino acid residues of CDR in the heavy chain variable domain (VH) are numbered 31- (HCDR1), 50-65 (HCDR2) and 95-102 (HCDR3); the amino acid residues of CDR in the light chain variable domain (VL) are numbered 24-34 (LCDR1), 50-56 (LCDR2) and 89-97 (LCDR3). Under Chothia, the CDR amino acids in the VH are numbered 26-32 (HCDR1), 52-56 (HCDR2) and 95-102 (HCDR3), and the amino acid residues in VL are numbered 26-32 (LCDR1), 50- 52 (LCDR2) and 91-96 (LCDR3). By combining Kabat and Chothia's CDR definitions, CDRs consist of amino acid residues 26-35 (HCDR1), 50-65 (HCDR2) and 95-102 (HCDR3) in human VH and amino acid residues 24-34 (LCDR1), 50-56 (LCDR2) and 89-97 (LCDR3) in human VL.
[00324] [00324] Under all definitions, each VH and VL typically includes three CDRs and four FRs, arranged from the amino terminal to the carboxy terminal in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FRA.
[00325] [00325] “In general, unless specifically indicated, anti-CD73 antibody molecules can include any combination of one or more Kabat CDRs, Chothia CDRs, combination of Kabat and Chothia CDRs, IMGT CDRs and / or an alternative definition, for example, described in Table 1.
[00326] [00326] As used herein, an "immunoglobulin variable domain sequence" refers to an amino acid sequence that can form the structure of an immunoglobulin variable domain. For example, the sequence may include all or part of the amino acid sequence of a naturally occurring variable domain. For example, the sequence may or may not include one, two or more N- or C-terminal amino acids, or it may include other changes that are compatible with formation of the protein structure.
[00327] [00327] The term "antigen binding site" refers to that part of an antibody molecule that comprises determinants that form an interface that binds to a CD73 polypeptide, or an epitope thereof. In relation to proteins (or protein mimetics), the antigen binding site typically includes one or more loops (of at least, for example, four amino acids or amino acid simulator) that form an interface that binds to a polypeptide of CD73. Typically, the antigen binding site of an antibody molecule includes at least one or two CDRs and / or hypervariable loops or, more typically, at least three, four, five or six CDRs and / or hypervariable loops.
[00328] [00328] “As used herein, the term" Eu numbering "refers to the Eu numbering convention for antibody constant regions, as described in Edelman, G.M. et al., Proc. Natl. Acad. USA, 63, 78 to 85 (1969) and Kabat et al., In "Sequences of Proteins of Immunological Interest", U.S. Dept. Health and Human Ser-
[00329] [00329] The terms "compete" or "compete crosswise" are used interchangeably throughout this document to refer to the ability of an antibody molecule to interfere with the binding of an anti-CD73 antibody molecule, for example for example, an anti-CD73 antibody molecule provided herein, to a target, for example, human CD73. Interference with binding can be direct or indirect (for example, through an allosteric modulation of the antibody molecule or the target). The extent to which an antibody molecule is able to interfere with the binding of another antibody molecule to the target, and therefore the possibility of saying that it competes, can be determined using a competition link, for example, a flow cytometry assay, an ELISA or BIACORE assay. In some modalities, a competition bond test is a quantitative competition test. In some embodiments, a first anti-CD73 antibody molecule is said to compete for binding to the target with a second anti-CD73 antibody molecule when the binding of the first antibody molecule to the target is reduced by 10% or more , for example, 20% or more, 30% or more, 40% or more, 50% or more, 55% or more, 60% or more, 65% or more, 70% or more, 75% or more, 80 % or more, 85% or more, 90% or more, 95% or more, 98% or more, 99% or more in a competition bond test (for example, a competition test described in this document).
[00330] [00330] As used herein, the term "epitope" refers to the chemical portions of an antigen (for example, human CD73) that specifically interact with an antibody molecule. Such chemical moieties, also referred to herein as epitopic determinants, typically comprise, or are part of, elements such as amino acid side chains or sugar side chains. An epitopic determinant can be defined by methods known in the art or disclosed in this document, for example, by crystallography or by hydrogen-deuterium exchange. At least one or some of the chemical moieties in the antibody molecule that specifically interact with an epitopic determinant are typically located on a CDR (or CDRs). Typically, an epitope has specific three-dimensional structural characteristics. Typically, an epitope has specific loading characteristics. Some epitopes are linear epitopes while others are conformational epitopes.
[00331] [00331] In one embodiment, an epitopic determinant is a chemical moiety in the antigen, for example, as an amino acid side chain or sugar side chain, or part of it, which, when the antigen and the antibody molecule are cocrystallized , is at a predetermined distance, for example, at 5 Angstroms, from a chemical moiety in the antibody molecule, referred to herein as a "crystallographic epitopic determinant". The crystallographic epitope determinants of an epitope are collectively called the "crystallographic epitope".
[00332] [00332] A first antibody molecule binds to the same epitope as a second antibody molecule (for example, a reference antibody molecule, for example, an antibody molecule disclosed herein) if the first antibody interacts with the same epitopic determinants on the antigen as well as the second antibody or reference antibody, for example, when the interaction is measured in the same way for both the antibody and the second antibody or reference antibody. Overlapping epitopes share at least one epitope determinant. A first antibody molecule binds to an epitope overlaid with a second antibody molecule (for example, a reference antibody molecule, for example, an antibody disclosed in this document) when both antibody molecules interact with a common epitopic determinant. A first and a second antibody molecule (for example, a reference antibody molecule, for example, an antibody molecule disclosed in this document) bind to substantially overlapping epitopes if at least half of the epitopic determinants of the second antibody or reference antibody are found to be epitopic determinants in the epitope of the first antibody. A first and a second antibody molecule (for example, a reference antibody molecule, for example, an antibody molecule disclosed herein) bind substantially to the same epitope if the first antibody molecule binds to at least half of the epitopic core determinants of the epitope of the second antibody or reference antibody, in which the epitopic core determinants are defined, for example, by crystallography or hydrogen-deuterium exchange.
[00333] [00333] “As used in this document, an antibody molecule" reduces the hydrogen-deuterium exchange "in an antigen fragment when the hydrogen-deuterium exchange in the antigen fragment, in the presence of the molecule antibody, is less than the hydrogen-deuterium exchange in the antigen fragment in the absence of the antibody molecule, as measured in a hydrogen-deuterium exchange assay.
[00334] [00334] As used herein, a reduction in "average hydrogen-deuterium exchange" is determined by the level of normalized hydrogen-deuterium exchange (Da per residue) in an antigen fragment in the absence of an antibody minus the normalized hydrogen-deuterium exchange level (Da per residue) in the antigen fragment in the presence of the antibody.
[00335] [00335] The terms "monoclonal antibody" or "composition of anti-
[00336] [00336] A "effectively human" protein is a protein that does not evoke a neutralizing antibody response, for example, the human anti-murine antibody (HAMA) response. HAMA can be problematic in a number of circumstances, for example, if the antibody molecule is administered repeatedly, for example, in the treatment of a chronic or recurrent disease condition. The HAMA response can make repeated antibody administration potentially ineffective due to an increase in antibody removal from serum (see, for example, Saleh et al., Cancer Immunol. Immunother., 32: 180 to 190 (1990) ) and also due to potential allergic reactions (see, for example, LoBuglio et al., Hybridoma, 5: 5,117 to 5,123 (1986)).
[00337] [00337] The antibody molecule can be a polyclonal antibody or a monocylonal antibody. In other embodiments, the antibody may be produced recombinantly, for example, produced by yeast display, phage display or by combinatorial methods. Alternatively, such antibodies can be selected from antibody presentation systems based on synthetic yeast, such as those described, for example, in Y. Xu et al, Addressing polyspercificity of antibodies selected from an in vitro yeast presentation system: a FACS-based, high-throughput selection and analytical tool. PEDS 26.10, 663-70 (2013) documents WO2Z2009036379; WOZ2010105256; and WO2012009568, in this document incorporated
[00338] [00338] In one embodiment, the antibody is a fully human antibody (for example, an antibody produced by yeast display, an antibody produced by phage display, or an antibody produced in a mouse that has been genetically modified to produce an antibody of a human immunoglobulin sequence) or a non-human antibody, for example, an antibody of rodent (mouse or rat), goat, primate (for example, monkey) or chamomile. Methods for producing rodent antibodies are known in the art.
[00339] [00339] Human monoclonal antibodies can be generated using transgenic mice that carry the human immunoglobulin genes instead of the mouse system. The splenocytes of these transgenic mice immunized with the antigen of interest are used to produce hybridomas that secrete human mAbs with specific affinities for epitopes of a human protein (see, for example, Wood et al. International Order WO 91/00906, Kucherlapati et al. PCT publication WO 91/10741; Lonberg et al. International application WO 92/03918; Kay et al. International application 92/03917; Lonberg, N. et al. 1994 Nature 368: 856-859 ; Green, LL et a /. 1994 Nature Genet. 7:13 to 21; Morrison, SL et al. 1994 Proc. Natl. Acad. Sci. USA 81: 6,851 to 6,855; Bruggeman et al. 1993 Year Immunol 7:33 to 40; Tuaillon et a /. 1993 PNAS 90: 3720-3724; Bruggeman et al. 1991 Eur J Immunol 21: 1,323 to 1,326).
[00340] [00340] An antibody can be one in which the variable region, or a portion of it, for example, CDRs, is generated in a non-human organism, for example, a rat or mouse. Chimeric, CDR-grafted, and humanized antibodies are covered by the invention. Antibodies generated in a non-human organism, for example, a rat or mouse, and then modified
[00341] [00341] Antibodies can be produced by any suitable recombinant DNA techniques known in the art (see Robinson et al, International Patent Publication PCT / US86 / 02269; Akira, et al., European Patent Application 184,187; Taniguchi, M., European Patent Application 171,496; Morrison et al., European Patent Application 173,494; Neuberger et a / l., International Application WO 86/01533; Cabilly et al. US Patent No. 4,816,567; Cabilly et al., European Patent Application 125,023; Better et a /. (1988 Science 240: 1,041 to 1,043); Liu et al. (1987) PNAS 84: 3439 to 3443; Liu et al, 1987, J. Immunol. 139: 3521 to 3526 ; Sun et al. (1987) PNAS 84: 214 to 218; Nishimura et al /., 1987, Canc. Res. 47: 999 to 1005; Wood et al. (1985) Nature 314: 446 to 449; and Shaw et a /., 1988, J. Natl Cancer Inst. 80: 1553 to 1559).
[00342] [00342] A humanized antibody or with CDR graft will have at least one or two, but generally all three recipient CDRs (from heavy and light immunoglobulin chains) replaced by a donor CDR. The antibody can be replaced by at least a portion of a non-human CDR or only some of the CDRs can be replaced by non-human CDRs. It is only necessary to replace the number of CDRs required for binding the humanized antibody to CD73. In some embodiments, the donor is a rodent antibody, for example, a mouse or mouse antibody and the recipient is a human framework or a human consensus framework. Typically, the immunoglobulin that provides the CDRs is called "donor" and the immunoglobulin that provides the framework is called "acceptor". In one embodiment, the donor immunoglobulin is a non-human (for example, rodent). The acceptor framework is a naturally occurring framework (for example, a human framework) or a consensus framework, or a sequence about 85% or more, for example, 90%, 95%, 99% or more identical at the same time.
[00343] [00343] “As used in this document, the term" consensus sequence "refers to the sequence formed from the most frequently occurring amino acids (or nucleotides) in a family of related sequences (See, for example, Winnaker, From Genes to Clones (Verlagsgesellschaft, Weinheim, Germany, 1987). In a family of proteins, each position in the consensus sequence is occupied by the amino acid that occurs most often in the position in the family. If two amino acids occur with equal frequency, either of them can be included in the consensus sequence, a "consensus framework" refers to the framework region in the consensus immunoglobulin sequence.
[00344] [00344] “An antibody can be humanized by the methods known in the art (see, for example, Morrison, SL, 1985, Science 229: 1202 to 1207, by Oi et al., 1986, Bio Techniques 4: 214, and by Queen et al. No. 5,585,089, No. 5,693,761 and No. 5,693,762, the contents of which are all incorporated herein by reference).
[00345] [00345] Humanized antibodies or with CDR grafting can be produced by CDR grafting or CDR substitution, in which one, two or all CDRs of an immunoglobulin chain can be substituted. See, for example, U.S. Patent No. 5,225,539; Jones et al. 1986 Nature 321: 552-525; Verhoeyan et al. 1988 Science 239: 1534; Beidler et al. 1988 J. Immunol. 141: 4053 to 4060; Winter US 5,225,539, the contents of which all are expressly incorporated herein by reference.
[00346] [00346] Also covered by the scope of the invention are anti-
[00347] [00347] The antibody molecule can be a single chain antibody. A single chain antibody (scFV) can be genetically modified (see, for example, Colcher, D. et a /. (1999) Ann NY Acad Sci 880: 263-80; and Reiter, Y. (1996) Clin Cancer Res 2: 245-52). The single chain antibody can be dimerized or multimerized to generate multivalent antibodies that have specificities for different epitopes on the same target protein.
[00348] [00348] In yet other embodiments, the antibody molecule has a heavy chain constant region chosen from, for example, the IgG1, IgG2, 1gG3, IgG4, IGM, IgA1, IgA2, IgD and heavy chain constant regions IgE; particularly, chosen from, for example, the heavy chain constant regions (e.g., human) of IgG1, IgG2, IgG3 and IgG4. In another embodiment, the antibody molecule has a light chain constant region chosen from, for example, Kappa or lambda light chain (e.g., human) constant regions. The constant region can be altered, for example, mutated, to modify the properties of the antibody (for example, to increase or decrease one or more of: Fc receptor binding, antibody glycosylation, the number of cysteine residues , effector cell function and / or complement function). In some modalities, the antibody has an effector function and can fix the complement. In other embodiments, the antibody does not recruit effector cells or fix complement. In certain embodiments, the antibody has reduced capacity or is unable to bind to an Fc receptor. For example, it may be an isotype or subtype, fragment or other mutant, which does not support binding to an Fc receptor, for example, it has a mutated or deleted Fc receptor binding region.
[00349] [00349] Methods for altering an antibody constant region are known in the art. Antibodies with altered function, for example, altered affinity for an effector ligand, such as FcR in a cell, or the complement component C1 can be produced by replacing at least one amino acid residue in the constant portion of the antibody with a different residue ( see, for example, documents EP 388,151 A1, US Pat. No. 5,624,821 and US Pat. No. 5,648,260, the contents of which are all incorporated by reference). Amino acid mutations that stabilize the antibody structure, such as S228P (Eu numbering) in human IgG4, are also contemplated. The similar type of changes could be described, which, if applied to murine, or immunoglobulin of another species, would reduce or eliminate these functions.
[00350] [00350] An antibody molecule can be derived from or linked to another functional molecule (for example, another peptide or protein). As used herein, a "deprivatized" antibody molecule is one that has been modified. Derivatization methods include, but are not limited to, the addition of a fluorescent chemical moiety, a radionucleotide, a toxin, an enzyme or an affinity linker such as biotin. Accordingly, the antibody molecules of the invention are intended to include derivatized or otherwise modified forms of the antibodies described herein, including immunoassay molecules. For example, an antibody molecule can be functionally linked (by chemical coupling, genetic fusion, non-covalent association or otherwise) to one or more other molecular entities, such as another antibody (for example, a bispecific antibody or a body ), a detectable agent, a cytotoxic agent, a pharmaceutical agent and / or a protein or peptide that can mediate the association of the antibody or portion of antibody with another molecule (such as a streptavidin core region or a polyhistidine tag).
[00351] [00351] A type of derivatized antibody molecule is produced by cross-linking two or more antibodies (of the same or different types, for example, to create bispecific antibodies). Suitable crosslinkers include those that are heterobifunctional, that have two reactive groups distinctly separated by a suitable spacer (for example, m-maleimidobenzoyl-N-hydroxysuccinimide ester) or homobifunctional (for example, disuccinimidyl suberate). Such binders are available from Pierce Chemical Company, Rockford, Ill.
[00352] [00352] Useful detectable agents with which an antibody molecule of the invention can be derivatized (or identified) to include fluorescent compounds, various enzymes, prosthetic groups, luminescent materials, bioluminescent materials, fluorescent emitted metal atoms, for example, europium (Eu) and other antanids and radioactive materials (described below). Exemplary detectable fluorescent agents include fluorescein, fluorescein isothiocyanate, rhodamine, 5-dimethylamine-1-naphthalenesulfonyl chloride, phycoerythrin and the like. An antibody can also be derivatized with detectable enzymes, such as alkaline phosphatase, strong root peroxidase, B-galactosidase, acetylcholinesterase, glucose oxidase and the like. When an antibody is derivatized with a detectable enzyme, it is detected by adding additional reagents that the enzyme uses to produce a detectable reaction product. For example
[00353] [00353] The identified antibody molecule can be used, for example, diagnostically and / or experimentally in numerous contexts, including (i) to isolate an antigen predetermined by standard techniques, such as affinity chromatography or immunoprecipitation; (ii) to detect a predetermined antigen (for example, in a cell layer or cell supernatant) to assess the abundance and pattern of protein expression; (iii) to monitor protein levels in tissue as part of a clinical testing procedure, for example, to determine the effectiveness of a given treatment regimen.
[00354] [00354] An antibody molecule can be conjugated to another molecular entity, typically an identification or a therapeutic chemical agent or portion (for example, immunomodulatory, immunostimulating, cytotoxic or cytostatic). Radioactive isotopes can be used in diagnostic or therapeutic applications. Radioactive isotopes that can be coupled to anti-CD73 antibodies include, but are not limited to, a, À or y emitters, or B and y emitters. Such radioactive isotopes include, but are not limited to, iodine ("91 or 1251), yttrium (ººY), lutetium (Lu), actinium (2PPAc), praseodymium, astatin
[00355] [00355] The invention provides radioidentified antibody molecules and methods for identifying them. In one embodiment, a method for identifying an antibody molecule is disclosed. The method includes putting an antibody molecule in contact with a chelating agent to thereby produce a conjugated antibody. The conjugated antibody is radioidentified with a radioisotope, for example, “Indium, yttrium and Lutetium to thereby produce an identified antibody molecule.
[00356] [00356] As discussed above, antibody molecules can be conjugated to a therapeutic agent. Therapeutically active radioisotopes have already been mentioned. Examples of other therapeutic agents include taxol, cytochalasin B, gramicidin D, ethidium bromide, emetin, mitomycin, etoposide, tenoposide, vincristine, vinblastine, colchicine, doxorubicin, daunorubicin, dihydroxy anthracine dione, mitoxantrine, mitramycin, actinomine D, 1-dehydrotestosterone, glucocorticoids, procaine, tetracaine, lidocaine, propranolol, puromycin, maytansinoids, for example maytansinol (see U.S. Pat. No. 5,208,020), CC-1065 (see U.S. Pat. No. 5,475,092, U.S. 5,585,499, No. 5,846, 545) and analogues or counterparts thereto. Therapeutic agents include, but are not limited to, antimetabolites
[00357] [00357] In one aspect, the invention features a method for providing a target binding molecule that binds to a CD73 receptor. For example, the target binding molecule is an antibody molecule. The method includes: providing a target protein that comprises at least a portion of non-human protein, the portion of which is homologous to (at least 70, 75, 80, 85, 87, 90, 92, 94, 95, 96, 97, 98, or 99% identical to) a corresponding portion of a human target protein, but different by at least one amino acid (for example, at least one, two, three, four, five, six, seven, eight or nine amino acids); obtaining an antibody molecule that binds to the antigen; and to evaluate the effectiveness of the binding agent in modulating the activity of the target protein. The method may additionally include administering the binding agent (e.g., antibody molecule) or a derivative (e.g., a humanized antibody molecule) to a human subject.
[00358] [00358] In certain embodiments, the antibody molecule is a multispecific antibody molecule (for example, bispecific or triespecific). Protocols for generating bispecific or heterodimeric antibody molecules are known in the art; including, but not limited to, for example, the "knob in a hole" approach
[00359] [00359] In other embodiments, the anti-CD73 antibody molecule (for example, a monospecific, bispecific or multispecific antibody molecule) is covalently linked, for example, fused, to another partner, for example, a protein, for example, one, two or more cytokines, for example, as a fusion molecule, for example, a fusion protein.
[00360] [00360] A "fusion protein" and a "fusion polypeptide" refer to a polypeptide that has at least two covalently linked portions, each of which is a polypeptide that has a different property. The property can be a biological property, such as in vitro or in vivo activity. The property can also be a simple physical or chemical property, such as binding to a target molecule, catalysis of a reaction, etc. The two portions can be linked directly by a single peptide bond or via a peptide linker, but they are in reading frame with each other.
[00361] [00361] This invention provides an isolated nucleic acid molecule that encodes the above antibody molecule, vectors and host cells thereof. The nucleic acid molecule includes, but is not limited to, RNA, genomic DNA and cDNA. Exemplary anti-CD73 Antibody Molecules
[00362] [00362] In some embodiments, the anti-CD73 antibody molecule comprises at least one antigen binding region, for example, a variable region or an antigen binding fragment thereof, of an antibody described herein, for example , an antibody chosen from 918, 350, 356, 358, 930, 373, 374, 376, 377, 379, 363, 366, 407, 893, 939, 430, or 398; or co-
[00363] [00363] In certain embodiments, the anti-CD73 antibody molecule comprises at least one, two, three or four variable regions of an antibody described herein, for example, an antibody chosen from 918, 350, 356 , 358, 930, 373, 374, 376, 377, 379, 363, 366, 407, 893, 939, 430, or 398; or as described in Table 1; or encoded by a nucleotide sequence in Table 1; or a substantially identical sequence (for example, which has at least about 80%, 85%, 90%, 92%, 95%, 97%, 98%, or 99% sequence identity) with any of the above sequences - tadas.
[00364] [00364] In some embodiments, the anti-CD73 antibody molecule comprises at least one or two variable heavy chain regions of an antibody described in this document, for example, an antibody chosen from 918, 350, 356, 358, 930, 373, 374, 376, 377, 379, 363, 366, 407, 893, 939, 430, or 398; or as described in Table 1; or encoded by a nucleotide sequence in Table 1; or a substantially identical sequence (for example, having at least about 80%, 85%, 90%, 92%, 95%, 97%, 98%, or 99% sequence identity) with any of the aforementioned sequences.
[00365] [00365] In certain embodiments, the anti-CD73 antibody molecule comprises at least one or two variable light chain regions of an antibody described herein, for example, an antibody chosen from 918, 350, 356, 358, 930, 373, 374, 376, 377, 379, 363, 366, 407, 893, 939, 430, or 398; or as described in
[00366] [00366] In one embodiment, the anti-CD73 antibody molecule includes an IgG4 heavy chain constant region, for example, a human IgG4. In another embodiment, human IgG4 includes a substitution (for example, a substitution from Ser to Pro) at position 228 according to Eu numbering. In yet another embodiment, the anti-CD73 antibody molecule includes a chain constant region weight of an IgG1, for example, a human IgG1. In one embodiment, human IgG1 includes a substitution (for example, a substitution from Asn to Ala) at position 297 according to the Eu numbering. In one embodiment, human IgG1 includes a substitution (for example, a substitution for Asp for Ala) at position 265 according to Eu numbering, a substitution (for example, a Pro to Ala substitution) at position 329 according to Eu numbering, or both. In one embodiment, human IgG1 includes a substitution (for example, a Leu to Ala substitution) at position 234 according to Eu numbering, a substitution (for example, a Leu to Ala substitution) at position 235 according to the numbering of Me, or both. In one embodiment, the heavy chain constant region comprises an amino acid sequence shown in Table 3, or a substantially identical sequence (for example, which is at least about 80%, 85%, 90%, 92%, 95 %, 97%, 98%, or 99% sequence identity) with it.
[00367] [00367] In yet another embodiment, the anti-CD73 antibody molecule includes a kappa light chain constant region, for example,
[00368] [00368] In another embodiment, the anti-CD73 antibody molecule includes an IgG4 heavy chain constant region, for example, a human IgG4 and a kappa light chain constant region, for example, a light chain constant region of human kappa, for example, a heavy and light chain constant region comprising an amino acid sequence shown in Table 3, or a substantially identical sequence (for example, which is at least about 80%, 85%, 90 %, 92%, 95%, 97%, 98%, or 99% sequence identity) with the same. In yet another mode, the anti-CD73 antibody molecule includes an IgG1 heavy chain constant region, for example, a human IgG1 and a kappa light chain constant region, for example, a constant region of human kappa light chain, for example, a heavy and light chain constant region comprising an amino acid sequence shown in Table 3, or a substantially identical sequence (for example, which is at least about 80%, 85% , 90%, 92%, 95%, 97%, 98%, or 99% sequence identity) with it. In one embodiment, human IgG1 includes a substitution at position 297 according to the Eu numbering (for example, a substitution from Asn to Ala). In one embodiment, human IgG1 includes a substitution at position 265 according to Eu numbering, a substitution at position 329 according to Eu numbering, or both (for example, an Asp to Ala substitution at position 265 and / or the replacement of Pro to Ala in the
[00369] [00369] In another embodiment, the anti-CD73 antibody molecule includes a heavy chain variable region and a constant region, a light chain variable region and a constant region, or both, which comprise the 918, 350 amino acid sequence , 356, 358, 930, 373, 374, 376, 377, 379, 363, 366, 407, 893, 939, 430, or 398; or as described in Table 1; or encoded by a sequence of nucleotides in Table 1; or a substantially identical sequence (for example, which has at least about 80%, 85%, 90%, 92%, 95%, 97%, 98%, or 99% sequence identity) with any of the aforementioned sequences .
[00370] [00370] In some embodiments, the anti-CD73 antibody molecule includes at least one, two or three complementarity determining regions (CDRs) of a variable heavy chain region of an antibody described herein, for example, an antibody chosen from 918, 350, 356, 358, 930, 373, 374, 376, 377, 379, 363, 366, 407, 893, 939, 430, or 398; or as described in Table 1, or encoded by a nucleotide sequence in Table 1; or a substantially identical sequence (for example, that has at least about 80%, 85%, 90%, 92%, 95%, 97%, 98%, or 99% sequence identity) with any of the su sequences - pracitadas.
[00371] [00371] In some embodiments, the anti-CD73 antibody molecule comprises at least one, two or three complementarity determining regions (CDRs) from a variable heavy chain region comprising an amino acid sequence shown
[00372] [00372] In some embodiments, the anti-CD73 antibody molecule includes at least one, two or three complementarity determining regions (CDRs) of a variable light chain region of an antibody described herein, for example, an anti - body chosen from 918, 350, 356, 358, 930, 373, 374, 376, 377, 379, 363, 366, 407, 893, 939, 430, or 398; or as described in Table 1, or encoded by a nucleotide sequence in Table 1; or a substantially identical sequence (for example, which has at least about 80%, 85%, 90%, 92%, 95%, 97%, 98%, or 99% sequence identity) with any of the above sequences - tadas.
[00373] [00373] In certain embodiments, the anti-CD73 antibody molecule includes at least one, two or three CDRs (or collectively all CDRs) from a light chain variable region comprising an amino acid sequence shown in Table 1, or encoded by a nucleotide sequence shown in Table 1. In some embodiments, one or more of the CDRs (or collectively all of the CDRs) have one, two, three, four, five, six or more changes, for example, amino acid substitutions, insertions or deletions, in relation to the CDRs shown in Table 1, or encoded by a sequence
[00374] [00374] In some embodiments, the anti-CD73 antibody molecule includes at least one, two, three, four, five or six CDRs (or collectively all CDRs) from a variable region of heavy and light chain comprising a sequence of amino acids shown in Table 1, or encoded by a nucleotide sequence shown in Table 1. In some embodiments, one or more of the CDRs (or collectively all of the CDRs) have one, two, three, four, five, six or more changes, for example, amino acid substitutions, insertions or deletions, in relation to the CDRs shown in Table 1, or encoded by a nucleotide sequence shown in Table 1.
[00375] [00375] In certain embodiments, the anti-CD73 antibody molecule includes all six CDRs of an antibody described in this document, for example, an antibody chosen from 918, 350, 356, 358, 930, 373, 374, 376, 377, 379, 363, 366, 407, 893, 939, 430, or 398; or as described in Table 1; or encoded by a nucleotide sequence in Table 1, or approximately related CDRs, for example, CDRs that are identical or that have at least one amino acid change, but no more than two, three or four changes (e.g., substitutions, deletions or insertions, for example, conservative substitutions). In certain embodiments, the anti-CD73 antibody molecule can include any CDR described herein. In certain embodiments, the anti-CD73 antibody molecule includes a substitution on a CDR heavy chain, for example, one or more substitutions on a CDR1, CDR2 and / or CDR3 heavy chain.
[00376] [00376] In some embodiments, the anti-CD73 antibody molecule includes at least one, two or three CDRs according to Kabat et al. (for example, at least one, two or three CDRs according to the definition of Kabat as shown in Table 1) of a variable heavy chain region of an antibody described in this document, for example, an antibody chosen from 918, 350, 356, 358, 930, 373, 374, 376, 377, 379, 363, 366, 407, 893, 939, 430, or 398; or as described in Table 1; or encoded by a nucleotide sequence in Table 1; or a substantially identical sequence (for example, that has at least about 80%, 85%, 90%, 92%, 95%, 97%, 98%, or 99% sequence identity) with any of the aforementioned sequences; or that has at least one amino acid change, but no more than two, three or four changes (for example, substitutions, deletions or insertions, for example, conservative substitutions) in relation to one, two or three CDRs according to Kabat et al. shown in Table 1.
[00377] [00377] In certain embodiments, the anti-CD73 antibody molecule includes at least one, two or three CDRs according to Kabat et al. (for example, at least one, two or three CDRs according to the definition of Kabat as shown in Table 1) from a light chain variable region of an antibody described in this document, for example, an antibody chosen from 918, 350, 356, 358, 930, 373, 374, 376, 377, 379, 363, 366, 407, 893, 939, 430, or 398; or encoded by a nucleotide sequence in Table 1; or a substantially identical sequence (for example, that has at least about 80%, 85%, 90%, 92%, 95%, 97%, 98%, or 99% sequence identity) with any one of the aforementioned strings; or that has at least one amino acid change, but no more than two, three or four changes (for example, substitutions, deletions or insertions, for example, conservative substitutions) in relation to one, two or three CDRs according to Kabat et al. shown in Table 1.
[00378] [00378] In certain embodiments, the anti-CD73 antibody molecule includes at least one, two, three, four, five or six CDRs according to Kabat et al. (for example, at least one, two, three, four, five or six CDRs according to the definition of Kabat as shown in Table 1) of the variable heavy and light chain regions of an antibody described in this document, for example, an antibody chosen from 918, 350, 356, 358, 930, 373, 374, 376, 377, 379, 363, 366, 407, 893, 939, 430, or 398; or as described in Table 1; or encoded by a nucleotide sequence in Table 1; or a substantially identical sequence (for example, which has at least about 80%, 85%, 90%, 92%, 95%, 97%, 98%, or 99% sequence identity) with any of the above sequences pracitadas; or that has at least one amino acid change, however, no more than two, three or four changes (for example, substitutions, deletions or insertions, for example, conservative substitutions) in relation to one, two, three, four, five or six CDRs according to Kabat et al. shown in Table 1.
[00379] [00379] In some embodiments, the anti-CD73 antibody molecule includes all six CDRs according to Kabat et al. (for example, all six CDRs according to the Kabat definition as shown in Table 1) of the variable heavy and light chain regions of an antibody described in this document, for example, an antibody chosen from 918, 350, 356, 358, 930, 373, 374, 376, 377, 379, 363, 366, 407, 893, 939, 430, or 398; or as described in Table 1; or encoded by a nucleotide sequence in Table 1; or a substantially identical sequence (for example, which has at least about 80%, 85%, 90%, 92%, 95%, 97%, 98%, or 99% sequence identity) with any of the aforementioned sequences ; or that has at least one amino acid change, but no more than two, three or four changes (for example, substitutions, deletions or insertions, for example, conservative substitutions) in relation to all six CDRs according to Kabat et al. shown in Table 1. In one embodiment, the anti-CD73 antibody molecule can include any CDR described in the present document.
[00380] [00380] In some embodiments, the anti-CD73 antibody molecule includes at least one, two or three hypervariable Chothia loops (for example, at least one, two or three hypervariable loops according to the Chothia definition as shown) in Table 1) of a heavy chain variable region of an antibody described in this document, for example, an antibody chosen from 918, 350, 356, 358, 930, 373, 374, 376, 377, 379, 363, 366, 407, 893, 939, 430, or 398; or as described in Table 1; or encoded by a nucleotide sequence in Table 1; or at least the amino acids of such hypervariable bonds that come in contact with CD73; or that have at least one amino acid change, but no more than two, three or four changes (for example, substitutions, deletions or insertions, for example, conservative substitutions) in relation to one, two or three loops hypervariables according to Chothia et al. shown in Table 1.
[00381] [00381] In certain embodiments, the anti-CD73 antibody molecule includes at least one, two or three hypervariable Chothia loops (for example, at least one, two or three hypervariable loops according to the Chothia definition as shown in the Table 1) a light chain variable region of an antibody described herein, for example, an antibody chosen from 918, 350, 356, 358, 930, 373, 374, 376, 377, 379, 363, 366, 407, 893, 939, 430, or 398; or as described in Table 1; or encoded by a nucleotide sequence in Table 1; or at least the amino acids of such hypervariable bonds that come in contact with CD73; or that has at least one amino acid change, but no more than two, three or four changes (for example, substitutions, deletions or insertions, for example, conservative substitutions) in relation to one, two or three hypervariable bonds of according to Chothia et al. shown in Table 1.
[00382] [00382] In certain embodiments, the anti-CD73 antibody molecule includes at least one, two, three, four, five or six hypervariable loops (for example, at least one, two, three, four, five or six la - hypervariable services according to Chothia's definition as shown in Table 1) of the variable regions of heavy and light chain of an antibody described in this document, for example, an antibody chosen from 918, 350, 356, 358 , 930, 373, 374, 376, 377, 379, 363, 366, 407, 893, 939, 430, or 398; or as described in Table 1; or encoded by a nucleotide sequence in Table 1; or at least the amino acids of those hypervariable bonds that come in contact with CD73; or who have at least one amino acid change, but no more than two, three or four changes (for example, substitutions, deletions or insertions, for example, conservative substitutions) in relation to one, two, three, four, five or six hypervariable ties according to Chothia et al. shown in Table 1.
[00383] [00383] In some embodiments, the anti-CD73 antibody molecule includes all six hypervariable loops (for example, all six hypervariable loops according to the Chothia definition as shown in Table 1) of an antibody described in this document. for example, an antibody chosen from 918, 350, 356, 358, 930, 373, 374, 376, 377, 379, 363, 366, 407, 893, 939, 430, or 398; or approximately related hypervariable loops, for example, hypervariable loops that are identical or have at least one amino acid change, but no more than two, three or four changes (for example, substitutions, deletions or insertions, for example, conservative substitutions); or who have at least one amino acid change, but no more than two, three or four changes (for example, substitutions, deletions or insertions, for example, conservative substitutions) in relation to all six hypervariable bonds of according to Chothia et al. shown in Table 1. In one embodiment, the anti-CD73 antibody molecule can include any hypervariable loop described herein.
[00384] [00384] In yet another embodiment, an anti-CD73 antibody molecule includes at least one, two or three hypervariable loops that have the same canonical structures as the corresponding hypervariable loop of an antibody described in this document, for example - plo, an antibody chosen from 918, 350, 356, 358, 930, 373, 374, 376, 377, 379, 363, 366, 407, 893, 939, 430, or 398, for example, the same structures that at least loop 1 and / or loop 2 of the heavy and / or light chain variable domains of an antibody described herein. See, for example, Chothia et al., (1992) J. Mol. Biol. 227: 799 to 817; Tomlinson et al., (1992) J. Mol. Biol. 227: 776 to 798 for descriptions of canonical hypervariable loop structures. These structures can be determined by inspecting the tables described in these references.
[00385] [00385] In certain embodiments, the anti-CD73 antibody molecule includes a combination of CDRs or hypervariable loops defined according to Kabat et al. and Chothia et al.
[00386] [00386] In one embodiment, the anti-CD73 antibody molecule includes at least one, two or three CDRs or hypervariable loops from a variable heavy chain region of an antibody described herein, for example, an antibody chosen from 918, 350, 356, 358, 930, 373, 374, 376, 377, 379, 363, 366, 407, 893, 939, 430, or 398, as defined by Kabat and Chothia (for example, at least one, two or three CDRs or hypervariable loops according to the definition of Kabat and Chothia as shown in Table 1); or encoded by a nucleotide sequence in Table 1; or a substantially identical sequence (for example, that has at least about 80%, 85%, 90%, 92%, 95%, 97%, 98%, or 99% sequence identity) with any of the su sequences - pracitadas; or that has at least one amino acid change, however, no more than two, three or four changes (for example, substitutions, deletions or insertions, for example, conservative substitutions) in relation to one, two or three CDRs or hypervariable loops according to Kabat and / or Chothia shown in Table 1.
[00387] [00387] In one embodiment, the anti-CD73 antibody molecule includes at least one, two or three CDRs or hypervariable loops from a variable light chain region of an antibody described in this document, for example, an antibody chosen from from 918, 350, 356, 358, 930, 373, 374, 376, 377, 379, 363, 366, 407, 893, 939, 430, or 398 according to the definition of Kabat and Chothia (for example , at least one, two or three CDRs or hypervariable loops according to the definition of Kabat and Chothia as presented in Table 1); or encoded by a nucleotide sequence in Table 1; or a substantially identical sequence (for example, that has at least about 80%, 85%, 90%, 92%, 95%, 97%, 98%, or 99% sequence identity) with any of the su sequences - pracitadas; or that has at least one amino acid change, however, no more than two, three or four changes (for example, substitutions, deletions or insertions, for example, conservative substitutions) in relation to one, two or three CDRs or hypervariable loops according to Kabat and / or Chothia shown in Table 1.
[00388] [00388] In some embodiments, the anti-CD73 antibody molecule includes at least one, two or three CDRs according to the definition of IMGT (for example, at least one, two or three CDRs according to the definition of IMGT as shown in Table 1) of a heavy chain variable region of an antibody described in this document, for example, an antibody chosen from 918, 350, 356, 358, 930, 373, 374, 376, 377, 379 , 363, 366, 407, 893, 939, 430, or 398; or as described in Table 1; or encoded by a nucleotide sequence in Table 1; or a substantially identical sequence (for example, that has at least about 80%, 85%, 90%, 92%, 95%, 97%, 98%, or 99% sequence identity) with any of the aforementioned sequences; or that has at least one amino acid change, however, no more than two, three or four changes (for example, substitutions, deletions or insertions, for example, conservative substitutions) in relation to one, two or three CDRs according with the definition of IMGT shown in Table 1.
[00389] [00389] In certain embodiments, the anti-CD73 antibody molecule includes at least one, two or three CDRs according to the IMGT definition (for example, at least one, two or three CDRs according to the definition of IMGT as shown in Table 1) of a light chain variable region of an antibody described in this document, for example, an antibody chosen from 918, 350, 356, 358, 930, 373, 374, 376, 377, 379 , 363, 366, 407, 893, 939, 430,
[00390] [00390] In certain embodiments, the anti-CD73 antibody molecule includes at least one, two, three, four, five or six CDRs according to the IMGT definition (for example, at least one, two, three, four, five or six CDRs according to the definition of IMGT as presented in Table 1) of the heavy and light chain variable regions of an antibody described in this document, for example, an antibody chosen from 918, 350, 356, 358, 930, 373, 374, 376, 377, 379, 363, 366, 407, 893, 939, 430, or 398; or as described in Table 1; or encoded by a nucleotide sequence in Table 1; or a substantially identical sequence (for example, which has at least about 80%, 85%, 90%, 92%, 95%, 97%, 98%, or 99% sequence identity) with any of the aforementioned sequences ; or that has at least one amino acid change, but no more than two, three or four changes (for example, substitutions, deletions or insertions, for example, conservative substitutions) in relation to one, two, three, four, five or six CDRs according to the definition of IMGT shown in Table 1.
[00391] [00391] In some embodiments, the anti-CD73 antibody molecule includes all six CDRs according to the IMGT definition (for example, all six CDRs according to the IMGT definition as shown in Table 1) from the variable regions heavy and light chain of an antibody described in this document, for example, an antibody chosen from 918, 350, 356, 358, 930, 373, 374, 376, 377, 379, 363, 366, 407, 893, 939, 430, or 398; or as described in Table 1; or encoded by a nucleotide sequence in Table 1; or a substantially identical sequence (for example, which has at least about 80%, 85%, 90%, 92%, 95%, 97%, 98%, or 99% sequence identity) with any of the aforementioned sequences ; or that has at least one amino acid change, but no more than two, three or four changes (for example, substitutions, deletions or insertions, for example, conservative substitutions) in relation to all six CDRs according to the definition of IMGT shown in Table 1. In one modality, the anti-CD73 antibody molecule can include any CDR described in this document.
[00392] [00392] The anti-CD73 antibody molecule can contain any combination of CDRs or hypervariable loops according to the definitions of Kabat and Chothia.
[00393] [00393] In some embodiments, the anti-CD73 antibody molecule includes at least one, two or three hypervariable Chothia loops from a variable heavy chain region of an antibody described herein, for example, an antibody from Table 1, or at least the amino acids of such hypervariable bonds that come in contact with CD73.
[00394] [00394] In some embodiments, the anti-CD73 antibody molecule includes at least one, two or three hypervariable Chothia loops from a variable light chain region of an antibody described herein, for example, an antibody from the Table 1, or at least the amino acids of such hypervariable bonds that come in contact with CD73.
[00395] [00395] In some embodiments, the anti-CD73 antibody molecule includes at least one, two or three hypervariable Kabat loops from a variable heavy chain region of an antibody described herein, for example, an antibody from Table 1, or at least the amino acids from such hypervariable bonds that come into contact with CD73.
[00396] [00396] In some embodiments, the anti-CD73 antibody molecule includes at least one, two or three hypervariable Kabat loops from a variable light chain region of an antibody described herein, for example, an antibody from Table 1, or at least the amino acids from such hypervariable bonds that come into contact with CD73.
[00397] [00397] In some embodiments, the anti-CD73 antibody molecule includes at least one, two or three hypervariable IMGT loops from a variable heavy chain region of an antibody described herein, for example, an antibody from Table 1, or at least the amino acids from such hypervariable bonds that come into contact with CD73.
[00398] [00398] In some embodiments, the anti-CD73 antibody molecule includes at least one, two or three hypervariable IMGT loops from a variable light chain region of an antibody described herein, for example, an antibody from Table 1, or at least the amino acids from such hypervariable bonds that come into contact with CD73.
[00399] [00399] In certain embodiments, the anti-CD73 antibody molecule includes at least one, two, three, four, five or six hypervariable loops of the variable regions of heavy and light chain of an antibody described in this document, for example , an antibody from Table 1, or at least the amino acids from such hypervariable bonds that come into contact with CD73.
[00400] [00400] In certain embodiments, the anti-CD73 antibody molecule includes all six hypervariable loops of the heavy and light chain variable regions of an antibody described in this document, for example, an antibody from Table 1, or at least the amino acids of such hypervariable loops that come into contact with CD73, or at least the amino acids of those hypervariable loops that come into contact with CD73, or hypervariable loops that are closely related, for example, hypervariable loops that are identical or have hair least one amino acid change, but no more than two, three or four changes (for example, substitutions, for example, conservative substitutions, deletions or insertions).
[00401] [00401] In some embodiments, the anti-CD73 antibody molecule includes at least one, two or three hypervariable loops that have the same canonical structures as the corresponding hypervariable loop of an antibody described in this document, for example, an antibody from Table 1, for example, the same canonical structures as at least loop 1 and / or loop 2 of the heavy and / or light chain variable domains of an antibody described in this document. See, for example, Chothia et al., (1992) J. Mol. Biol. 227: 799 to 817; Tomlinson et al., (1992) J. Mol. Biol. 227: 776 to 798 for descriptions of canonical hypervariable loop structures. These structures can be determined by inspecting the tables described in these references. In a modality, for example, a modality that comprises a variable region, CDR (for example, Chothia, Kabat or IMGT CDR), or another sequence mentioned in this document, for example, in Table 1, the antisera molecule body is a monospecific antibody molecule, a bispecific antibody molecule or is an antibody molecule comprising an antigen binding fragment of an antibody, for example, an antibody medium or antigen binding fragment of an antibody medium.
[00402] [00402] In some embodiments, the heavy or light chain variable domain or both of the anti-CD73 antibody molecule includes an amino acid sequence, which is substantially identical to an amino acid disclosed in this document, for example, which has at least about 80%, 85%, 90%, 92%, 95%, 97%, 98%, or 99% sequence identity to a variable region of an antibody described herein, for example, an antibody chosen from 918, 350, 356, 358, 930, 373, 374, 376, 377, 379, 363, 366, 407, 893, 939, 430, or 398; or as described in Table 1; or encoded by a nucleotide sequence in Table 1; or that differs at least 1 or 5 residues, but less than 40, 30, 20 or 10 residues, from a variable region of an antibody described in this document.
[00403] [00403] In certain embodiments, the heavy or light chain variable region or both of the anti-CD73 antibody molecule includes an amino acid sequence encoded by a nucleic acid sequence described herein or a nucleic acid that hybridizes to a nucleic acid sequence described in the present document (for example, a nucleic acid sequence as shown in Table 1) or its complement, for example, under low stringency, medium stringency or high stringency , or other hybridization condition described in this document.
[00404] [00404] In certain embodiments, the anti-CD73 antibody molecule comprises at least one, two, three or four antigen binding regions, for example, variable regions, which have an amino acid sequence as shown in Table 1, or one sequence substantially identical to the same (for example, a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity to the same, or that differs by no more than 1, 2, 5, 10 or 15 amino acid residues of the sequences shown in Table 1. In certain embodiments, the anti-CD73 antibody molecule includes a VH and / or VL domain encoded by a nucleic acid that has a nucleotide sequence that encodes an antibody in the Table 1, or a sequence substantially identical to any of the nucleotide sequences (for example, a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity to it, or that differs by no more than 3, 6, 15, 30 or 45 nucleotides from sequences shown in Table 1).
[00405] [00405] In certain embodiments, the anti-CD73 antibody molecule comprises at least one, two or three (for example, all) CDRs from a heavy chain variable region that has an amino acid sequence as shown in Table 1, or a sequence substantially homologous to the same (for example, a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with the same and / or that has one, two, three or more substitutions, insertions or deletions, for example, preserved substitutions). In some embodiments, the anti-CD73 antibody molecule comprises at least one, two or three (for example, all) CDRs from a light chain variable region that has an amino acid sequence as shown in Table 1, or a substantially homologous sequence to it (for example, a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with the same and / or that has one, two, three or more substitutions, insertions or deletions , for example, preserved substitutions). In certain embodiments, the anti-CD73 antibody molecule comprises at least one, two, three, four, five or six (for example, all) CDRs from the heavy and light chain variable regions that have an amino acid sequence as shown in the Table 1, or a sequence substantially homologous to the same (for example, a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity to the same and / or that has one, two, three or more substitutions, insertions or deletions, for example, preserved substitutions).
[00406] [00406] In some embodiments, the anti-CD73 antibody molecule comprises at least one, two or three (for example, all) CDRs and / or hypervariable loops of a heavy chain variable region that has an amino acid sequence of an antibody described herein, for example, an antibody chosen from 918, 350, 356, 358, 930, 373, 374, 376, 377, 379, 363, 366, 407, 893, 939, 430, or 398, as summarized in Table 1, or a sequence substantially identical to the same (for example, a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with the same and / or having one, two, three or more substitutions, insertions or deletions, for example, preserved substitutions). In certain embodiments, the anti-CD73 antibody molecule comprises at least one, two or three (for example, all) CDRs and / or hypervariable loops from a light chain variable region that has an amino acid sequence of a described antibody in this document, for example, an antibody chosen from 918, 350, 356, 358, 930, 373, 374, 376, 377, 379, 363, 366, 407, 893, 939, 430, or 398, as summarized in Table 1, or a sequence substantially identical to the same (for example, a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with the same and / or that has one, two, three or more substitutions, insertions or deletions, for example, conserved substitutions). In some embodiments, the anti-CD73 antibody molecule comprises all six CDRs and / or hypervariable loops described in this document, for example, described in Table 1.
[00407] [00407] In some embodiments, the antibody molecule has a variable region that is identical in sequence, or that differs by 1, 2, 3 or 4 amino acids from a variable region described in the present document (for example, a region of FR disclosed in this document).
[00408] [00408] In one embodiment, the anti-CD73 antibody molecule comprises a heavy chain variable region (VH) comprising a VHCDR3 amino acid sequence of GGLYGSGSYLS-DFDL (SEQ ID NO: 37). In one embodiment, the anti-CD73 antibody molecule comprises a heavy chain variable region (VH) comprising an amino acid sequence VHCDR3 of ES-QESPYNNWFDP (SEQ ID NO: 3).
[00409] [00409] In one embodiment, the anti-CD73 antibody molecule comprises a heavy chain variable region (VH) comprising a VHCDR1 amino acid sequence of SEQ ID NO: 88, a VÓHCDR 2 amino acid sequence of SEQ ID NO: 89 and a VÓHCDR3 amino acid sequence of SEQ ID NO: 37 and a light chain variable region (VL) comprising a VLCDR1 amino acid sequence of SEQ ID NO: 48, a amino acid sequence VLCDR2 noacids of SEQ ID NO: 49 and a VLCDR3 amino acid sequence of SEQ ID NO: 50, each disclosed in Table
[00410] [00410] In one embodiment, the anti-CD73 antibody molecule comprises a variable heavy chain (VH) region comprising a VHCDR1 amino acid sequence of SEQ ID NO: 122, a VHCDR2 amino acid sequence of SEQ ID NO: 123 and a VHCDR3 amino acid sequence of SEQ ID NO: 37 and a light chain variable region (VL) comprising a VLCDR1 amino acid sequence of SEQ ID NO: 51, a VLCDR2 amino acid sequence of SEQ ID NO: 52 and a VLCDR3 amino acid sequence of SEQ ID NO: 53, each disclosed in Table 2. In one embodiment, the anti-CD73 antibody molecule comprises a heavy chain variable region (VH ) comprising a VHCDR1 amino acid sequence of SEQ ID NO: 124, a VHCDR2 amino acid sequence of SEQ ID NO: 125 and a VÓHCDR3 amino acid sequence of SEQ ID NO: 3 and a light chain variable region (VL) comprising a VLCDR1 amino acid sequence of SEQ ID NO: 17, a sequence of amino VLCDR2 amino acids of SEQ ID NO: 18 and a VLCDR3 amino acid sequence of SEQ ID NO: 19, each disclosed in Table 2.
[00411] [00411] In one embodiment, the anti-CD73 antibody molecule comprises a heavy chain variable region (VH) comprising a VHCDR1 amino acid sequence of SEQ ID NO: 126, a VÓHCDR 2 amino acid sequence of SEQ ID NO: 89 and a VÓHCDR3 amino acid sequence of SEQ ID NO: 37 and a light chain variable region (VL) comprising a VLCDR1 amino acid sequence of SEQ ID NO: 48, a amino acid sequence VLCDR2 noacids of SEQ ID NO: 49 and a VLCDR3 amino acid sequence of SEQ ID NO: 50, each disclosed in Table 2. In one embodiment, the anti-CD73 antibody molecule comprises a heavy chain variable region (VH) comprising a VHCDR1 amino acid sequence of SEQ ID NO: 127, a sequence
[00412] [00412] In one embodiment, the anti-CD73 antibody molecule comprises a heavy chain variable region (VH) comprising a VÓHCDR1 amino acid sequence of SEQ ID NO: 128, a VÓMCDR2 amino acid sequence of SEQ ID NO: 129, and a VÓHCDR3 amino acid sequence of SEQ ID NO: 43 and a light chain variable region (VL) comprising a VLCDR1 amino acid sequence of SEQ ID NO: 54, a VLCDR2 amino acid sequence of SEQ ID NO: 52 and a VLCDR3 amino acid sequence of SEQ ID NO: 50, each disclosed in Table 2. In one embodiment, the anti-CD73 antibody molecule comprises a heavy chain variable region ( VH) comprising a VÓHCDR1 amino acid sequence of SEQ ID NO: 130, a VHCDR2 amino acid sequence of SEQ ID NO: 131, and a VÓHCDR3 amino acid sequence of SEQ ID NO: 9 and a variable region of light chain (VL) comprising a VLCDR1 amino acid sequence of SEQ ID NO: 20, a sequence of VLCDR2 amino acids of SEQ ID NO: 18 and a VLCDR3 amino acid sequence of SEQ ID NO: 16, each disclosed in Table 2.
[00413] [00413] In one embodiment, the anti-CD73 antibody molecule comprises a heavy chain variable region (VH) comprising a VÓHCDR1 amino acid sequence of SEQ ID NO: 189, a VÓHCDR2 amino acid sequence of SEQ ID NO: 89 and a VÓHCDR3 amino acid sequence of SEQ ID NO: 43 and a light chain variable region (VL) comprising a VLCDR1 amino acid sequence of SEQ ID NO: 48, a amino acid sequence of VLCDR2 of SEQ ID NO: 49 and an amino acid sequence of VLCDR3 of SEQ ID NO: 50, each disclosed in Table 2. In one embodiment, the anti-CD73 antibody molecule comprises a heavy chain variable region (VH ) comprising a VHCDR1 amino acid sequence of SEQ ID NO: 196, a VHCDR2 amino acid sequence of SEQ ID NO: 91 and a VÓHCDR3 amino acid sequence of SEQ ID NO: 9 and a region light chain variable (VL) comprising a VLCDR1 amino acid sequence of SEQ ID NO: 14, a sequence that of VLCDR2 amino acids of SEQ ID NO: 15 and a sequence of VLCDR3 amino acids of SEQ ID NO: 16, each disclosed in Table 2.
[00414] [00414] In one embodiment, the anti-CD73 antibody molecule comprises a VH comprising a VHCDR1 amino acid sequence of SEQ ID NO: 38, a VHCDR 2 amino acid sequence of SEQ ID NO: 36 and an amino acid sequence of VHCDR3 of SEQ ID NO: 37; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 48, a VLCDR2 amino acid sequence of SEQ ID NO: 49 and a VLCDR3 amino acid sequence of SEQ ID NO: 50. In one In this embodiment, the anti-CD73 antibody molecule comprises a VH comprising an amino acid sequence of VHCDR1 of SEQ ID NO: 72, an amino acid sequence of VHCDR 2 of SEQ ID NO: 71 and an amino acid sequence of VÓHCDR3 of SEQ ID NO: 37; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 48, a VLCDR2 amino acid sequence of SEQ ID NO: 49 and a VLCDR3 amino acid sequence of SEQ ID NO: 50. In one embodiment, the molecule An anti-CD73 antibody comprises a VH comprising a VÓHCDR1 amino acid sequence of SEQ ID NO: 38, a VÓHCDR2 amino acid sequence of
[00415] [00415] In one embodiment, the anti-CD73 antibody molecule comprises a VH comprising an amino acid sequence of VHCDR1 of SEQ ID NO: 39, an amino acid sequence of
[00416] [00416] In one embodiment, the anti-CD73 antibody molecule comprises a VH comprising a VHCDR1 amino acid sequence of SEQ ID NO: 35, a VHCDR 2 amino acid sequence of SEQ ID NO: 36 and an amino acid sequence of VHCDR3 of SEQ ID NO: 37; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 48, a VLCDR2 amino acid sequence of SEQ ID NO: 49 and a VLCDR3 amino acid sequence of SEQ ID NO: 50. In one In this embodiment, the anti-CD73 antibody molecule comprises a VH comprising an amino acid sequence of VHCDR1 of SEQ ID NO: 70, an amino acid sequence of VHCDR 2 of SEQ ID NO: 71 and an amino acid sequence of VÓHCDR3 of SEQ ID NO: 37; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 48, a VLCDR2 amino acid sequence of SEQ ID NO: 49 and a VLCDR3 amino acid sequence of SEQ ID NO: 50. In one embodiment, the molecule An anti-CD73 antibody comprises a VH comprising a VÓHCDR1 amino acid sequence of SEQ ID NO: 81, a VÓHCDR2 amino acid sequence of
[00417] [00417] In one embodiment, the anti-CD73 antibody molecule comprises a VH comprising an amino acid sequence of VHCDR1 of SEQ ID NO: 41, an amino acid sequence of
[00418] [00418] In one embodiment, the anti-CD73 antibody molecule comprises a VH comprising a VHCDR1 amino acid sequence of SEQ ID NO: 190, a VHCDR 2 amino acid sequence of SEQ ID NO: 36 and an amino acid sequence of VHCDR3 of SEQ ID NO: 43; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 48, a VLCDR2 amino acid sequence of SEQ ID NO: 49 and a VLCDR3 amino acid sequence of SEQ ID NO: 50. In one In this embodiment, the anti-CD73 antibody molecule comprises a VH comprising an amino acid sequence of VÓHCDR1 of SEQ ID NO: 191, an amino acid sequence of VHCDR 2 of SEQ ID NO: 71 and an amino acid sequence of VÓHCDR3 of SEQ ID NO: 43; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 48, a VLCDR2 amino acid sequence of SEQ ID NO: 49 and a VLCDR3 amino acid sequence of SEQ ID NO: 50. In one embodiment, the molecule An anti-CD73 antibody comprises a VH comprising a VÓHCDR1 amino acid sequence of SEQ ID NO: 192, a VHCDR2 amino acid sequence of
[00419] [00419] In other embodiments, the aforementioned antibodies comprise a variable region of heavy chain comprising an amino acid sequence that is at least about 85%
[00420] [00420] In other embodiments, the aforementioned antibodies comprise a variable light chain region comprising an amino acid sequence that is at least about 85% (for example, at least 90%, 95%, 97%, 98% , 99%, or 100%) of sequence identity with any of SEQ ID NOs: 55 or 21.
[00421] [00421] In other embodiments, the aforementioned antibodies comprise a heavy chain comprising an amino acid sequence that is at least about 85% (for example, at least 90%, 95%, 97%, 98%, 99% , or 100%) of sequence identity with any of SEQ ID NOs: 46, 79, 86, 114, 116 or
[00422] [00422] In other embodiments, the aforementioned antibodies comprise a light chain comprising an amino acid sequence that is at least about 85% (for example, at least 90%, 95%, 97%, 98%, 99%, or 100%) sequence identity with any of SEQ ID NOs: 57 or 23.
[00423] [00423] In other embodiments, the antibody molecule comprises a variable region of heavy chain that comprises a
[00424] [00424] In other embodiments, the antibody molecule comprises a heavy chain comprising an amino acid sequence that is at least about 85% (for example, at least 90%, 95%, 97%, 98% , 99%, or 100%) sequence identity with SEQ ID NO: 46; and a light chain comprising an amino acid sequence that has at least about 85% (for example, at least 90%, 95%, 97%, 98%, 99%, or 100%) sequence with SEQ ID NO: 57. In other embodiments, the antibody molecule comprises a heavy chain comprising an amino acid sequence that is at least about 85% (for example, at least 90%, 95%, 97% , 98%, 99%, or 100%) sequence identity with SEQ ID NO: 79; and a light chain comprising an amino acid sequence that has at least about 85% (e.g., at least 90%, 95%, 97%, 98%, 99%, or 100%) of sequence identity with SEQ ID NO: 57. In other embodiments, the antibody molecule comprises a heavy chain comprising an amino acid sequence that is at least about 85% (for example, at least 90%, 95%, 97%, 98 %, 99%, or 100%) sequence identity with SEQ ID NO: 86; and a light chain comprising an amino acid sequence that has at least about 85% (e.g., at least 90%, 95%, 97%, 98%, 99%, or 100%) of sequence identity with SEQ ID NO: 57. In other embodiments, the antibody molecule comprises a heavy chain comprising an amino acid sequence that is at least about 85% (for example, at least 90%, 95%, 97%, 98%, 99% , or 100%) sequence identity with SEQ ID NO: 114; and a light chain comprising an amino acid sequence that has at least about 85% (for example, at least 90%, 95%, 97%, 98%, 99%, or 100%) of sequence identity with SEQ ID NO: 57. In other embodiments, the antibody molecule comprises a heavy chain comprising an amino acid sequence that is at least about 85% (for example, at least 90%, 95%, 97%, 98 %, 99%, or 100%) of sequence identity with SEQ ID NO: 116; and a light chain comprising an amino acid sequence that has at least about 85% (for example, at least 90%, 95%, 97%, 98%, 99%, or 100%) of sequence identity with SEQ ID NO: 57. In other embodiments, the antibody molecule comprises a heavy chain comprising an amino acid sequence that is at least about 85% (for example, at least 90%, 95%, 97% , 98%, 99%, or 100%) sequence identity with SEQ ID NO: 117; and a light chain comprising an amino acid sequence that is at least about 85% (e.g., at least 90%, 95%,
[00425] [00425] In other embodiments, the antibody molecule comprises a VHCDR1, a VHCDR2, a VHCDR3, a VLCDRI1, a VLCDR2 and a VLCDR3 which comprise the amino acid sequences of SEQ ID NOs: 81, 201, 37, 205, 206 and 207, respectively. In other embodiments, the antibody molecule comprises a VHCDR1, a VHCDR2, a VHCDR3, a VLCDR1, a VLCDR 2 and a VLCDR3 that comprise the amino acid sequences of SEQ ID NOs: 38, 201, 37, 205 , 206 and 207, respectively. In other embodiments, the antibody molecule comprises a VHCDR1, a VHCDR2, a VHCDR3, a VLCDR1, a VLCDR2 and a VLCDR3 that comprise the amino acid sequences of SEQ ID NOs: 82, 202, 37, 208, 209 and 228, respectively . In other embodiments, the antibody molecule comprises a VHCDRI1, a VHCDR2, a VHCDR3, a VLCDR1, a VLCDR2 and a VLCDR3 that comprise the amino acid sequences of SEQ ID
[00426] [00426] In other embodiments, the antibody molecule comprises a VHCDR1, a VHCDR2, a VHCDR3, a VLCDRI1, a VLCDR2 and a VLCDR3 which comprise the amino acid sequences of SEQ ID NOs: 81, 231, 37, 205, 206 and 207, respectively. In other embodiments, the antibody molecule comprises a VHCDR1, a VHCDR2, a VHCDR3, a VLCDR1, a VLCDR 2 and a VLCDR3 that comprise the amino acid sequences of SEQ ID NOs: 38, 231, 37, 205 , 206 and 207, respectively. In other embodiments, the antibody molecule comprises a VHCDR1, a VHCDR2, a VHCDR3, a VLCDR1, a VLCDR2 and a VLCDR3 that comprise the amino acid sequences of SEQ ID NOs: 82, 232, 37, 208, 209 and 228, respectively . In other embodiments, the antibody molecule comprises a VHCDR1, a VHCDR2, a VHCDR3, a VLCDR1, a VLCDR2 and a VLCDR3 that comprise the amino acid sequences of SEQ ID NOs: 83, 233, 43, 229, 209 and 207, respectively . In other embodiments, the antibody molecule comprises a VHCDR1, a VHCDR2, a VHCDR3, a VLCDR1, a VLCDR2 and a VLCDR3 that comprise the amino acid sequences of SEQ ID NOs: 192, 231, 43, 205, 206 and 207 , respectively. In other fashion-
[00427] [00427] In other embodiments, the antibody molecule comprises a VHCDR1, a VHCDR2, a VHCDR3, a VLCDRI1, a VLCDR2 and a VLCDR3 that comprise the amino acid sequences of SEQ ID NOs: 235, 236, 237, 246, 15 and 247, respectively. In other embodiments, the antibody molecule comprises a VHCDR1, a VHCDR2, a VHCDR3, a VLCDR1, a VLCDR 2 and a VLCDR3 that comprise the amino acid sequences of SEQ ID NOs: 238, 236, 237, 246 , 15 and 247, respectively. In other embodiments, the antibody molecule comprises a VHCDR1, a VHCDR2, a VHCDR3, a VLCDR1, a VLCDR2 and a VLCDR3 comprising the amino acid sequences of SEQ ID NOs: 239, 240, 237, 248, 18 and 249, respectively . In other embodiments, the antibody molecule comprises a VHCDR1, a VHCDR2, a VHCDR3, a VLCDR1, a VLCDR2 and a VLCDR3 comprising the amino acid sequences of SEQ ID NOs: 241, 242, 243, 250, 18 and 247 , respectively. In other ways, the antibody molecule comprises a VHCDR1, a VHCDR2, a VHCDR3, a VLCDR1, a VLCDR2 and a VLCDR3 comprising the amino acid sequences of SEQ ID NOs: 244, 236, 243, 246, 15 and 247 , respectively. In other ways, the antibody molecule comprises a VH and VL that comprises the amino acid sequences of SEQ ID NOs: 245 and 251, respectively (or an amino acid sequence that is at least about 85% (for example , at least 90%, 95%, 97%, 98%, 99%, or 100%) sequence identity therewith).
[00428] [00428] In other embodiments, the antibody molecule comprises a VHCDR1, a VHCDR2, a VHCDR3, a VLCDRI1, a VLCDR2 and a VLCDR3 that comprise the amino acid sequences of SEQ ID NOs: 252, 253, 254, 262, 263 and 264, respectively. In other embodiments, the antibody molecule comprises a VHCDR1, a VHCDR2, a VHCDR3, a VLCDR1, a VLCDR 2 and a VLCDR3 that comprise the amino acid sequences of SEQ ID NOs: 255, 253, 254, 262 , 263 and 264, respectively. In other embodiments, the antibody molecule comprises a VHCDR1, a VHCDR2, a VHCDR3, a VLCDR1, a VLCDR2 and a VLCDR3 that comprise the amino acid sequences of SEQ ID NOs: 155, 256, 254, 265, 266 and 267, respectively . In other embodiments, the antibody molecule comprises a VHCDR1, a VHCDR2, a VHCDR3, a VLCDR1, a VLCDR2 and a VLCDR3 comprising the amino acid sequences of SEQ ID NOs: 257, 258, 259, 268, 266 and 264 , respectively. In other modalities, the antibody molecule comprises a VHCDR1, a VHCDR2, a VHCDR3, a VLCDR1, a VLCDR2 and a VLCDR3 which comprise the amino acid sequences of SEQ ID NOs: 260, 253, 259, 262, 263 and 264 , respectively. In other modalities, the antibody molecule comprises a VH and VL that comprises the amino acid sequences of SEQ ID NOs: 261 and 269, respectively (or an amino acid sequence that is at least about 85% (for example , at least 90%, 95%, 97%, 98%, 99%, or 100%) sequence identity therewith).
[00429] [00429] In other embodiments, the antibody molecule comprises a VHCDR1, a VHCDR2, a VHCDR3, a VLCDRI1, a VLCDR2 and a VLCDR3 comprising the amino acid sequences of SEQ ID NOs: 287, 288, 289, 298, 49 and 299, respectively. In other embodiments, the antibody molecule comprises
[00430] [00430] In other embodiments, the antibody molecule comprises a VHCDR1, a VHCDR2, a VHCDR3, a VLCDRI1, a VLCDR2 and a VLCDR3 that comprise the amino acid sequences of SEQ ID NOs: 304, 305, 306, 14, 15 and 314, respectively. In other embodiments, the antibody molecule comprises a VHCDR1, a VHCDR2, a VHCDR3, a VLCDR1, a VLCDR 2 and a VLCDR3 that comprise the amino acid sequences of SEQ ID NOs: 61, 305, 306, 14 , 15 and 314, respectively. In other embodiments, the antibody molecule comprises a VHCDR1, a VHCDR2, a VHCDR3, a VLCDR1, a VLCDR2 and a VLCDR3 which comprise the amino acid sequences of SEQ ID NOs: 307, 308, 306, 17, 18 and 315, respectively . In other embodiments, the antibody molecule comprises a VHCDR1, a VHCDR2, a VHCDR3, a VLCDR1, a VLCDR2 and a VLCDR3 that comprise the amino acid sequences of SEQ ID NOs: 309, 310, 311, 20, 18 and 314, respectively . In other embodiments, the antibody molecule comprises a VHCDR1, a VHCDR2, a VHCDR3, a VLCDR1, a VLCDR2 and a VLCDR3 that comprise the amino acid sequences of SEQ ID NOs: 312, 305, 311, 14, 15 and 314 , respectively. In other modalities, the antibody molecule comprises a VH and VL that comprises the amino acid sequences of SEQ ID NOs: 313 and 316, respectively (or an amino acid sequence that is at least about 85% (for example , at least 90%, 95%, 97%, 98%, 99%, or 100%) sequence identity therewith).
[00431] [00431] In other embodiments, the antibody molecule comprises a VHCDR1, a VHCDR2, a VHCDR3, a VLCDRI1, a VLCDR2 and a VLCDR3 that comprise the amino acid sequences of SEQ ID NOs: 317, 318, 319, 14, 15 and 328, respectively. In other embodiments, the antibody molecule comprises a VHCDR1, a VHCDR2, a VHCDR3, a VLCDR1, a VLCDR 2 and a VLCDR3 that comprise the amino acid sequences of SEQ ID NOs: 320, 318, 319, 14 , 15 and 328, respectively. In other embodiments, the antibody molecule comprises a VHCDR1, a VHCDR2, a VHCDR3, a VLCDR1, a VLCDR2 and a VLCDR3 that comprise the amino acid sequences of SEQ ID NOs: 321, 322, 319, 17, 18 and 329, respectively . In other embodiments, the antibody molecule comprises a VHCDRI1, a VHCDR2, a VHCDR3, a VLCDR1, a VLCDR2 and a VLCDR3 that comprise the amino acid sequences of SEQ ID
[00432] [00432] In other embodiments, the aforementioned antibody molecules are chosen from a complete antibody, a bispecific antibody, Fab, F (ab ') 2, Fv or a single chain Fv fragment (scFv).
[00433] [00433] In other embodiments, the antibody molecules mentioned above comprise a heavy chain constant region selected from I9gG1, IgG2, IgG3 and I9G4.
[00434] [00434] In other embodiments, the antibody molecules mentioned above comprise a light chain constant region chosen from the kappa or lambda light chain constant regions.
[00435] [00435] In some embodiments, the anti-CD73 antibody molecule comprises a heavy chain variable region, a light chain variable region, a heavy chain constant region and / or a light chain constant region disclosed in Table 1. In in some embodiments, the anti-CD73 antibody molecule comprises a heavy chain constant region and / or a light chain constant region disclosed in Table 3. In some embodiments, the anti-CD73 antibody molecule comprises a constant region heavy chain comprising a sequence of selected amino acids
[00436] Exemplary anti-CD73 antibody sequences are described in Tables 1 and 2 below.
[00437] [00437] In other embodiments, the aforementioned antibody molecules are capable of binding to human CD73 with a dissociation constant (Kp) of less than about 1 x 10º M, 1 x 10º M, 1 x 106 M, 1 x 107 M, 1 x 108 M, 1 x 10º M, for example, as measured by Biacore, Octet, flow cytometry or ELISA.
[00438] [00438] In some embodiments, the antibody molecule binds to a mammalian CD73, for example, human or cinomolg. For example, the antibody molecule binds to an epitope, for example, linear or conformational epitope, (for example, an epitope as described herein), on CD73. In certain aspects, it is advantageous to identify an antibody that binds with high affinity to human and cinomolgous counterparts of a protein of interest. The desirable cross-reactivity allows the same antibody (or two
[00439] [00439] In some embodiments, an isolated antibody molecule that competes for binding to human CD73 with the aforementioned anti-CD73 antibody molecules is disclosed in the present document.
[00440] [00440] In some embodiments, an isolated antibody molecule that binds to the same epitope as, substantially the same epitope as, an epitope that overlaps with, or an epitope that substantially overlaps with the epitope of the aforementioned anti-CD73 antibody molecules.
[00441] [00441] In some embodiments, the antibody molecule binds to the N-terminal domain of human CD73. In some embodiments, the antibody molecule binds to human CD73 loop A and / or loop B. In some embodiments, the antibody molecule, when bound to human CD73, induces a conformational exchange in the C loop of human CD73.
[00442] [00442] In some embodiments, the antibody molecule reduces the hydrogen-deuterium exchange in one or more regions of a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein that consists of the amino acid sequence of SEQ ID NO: 171) when linked to it, in which one or more regions are selected from the group consisting of residues 158 to 172, residues 206 to 215, residues 368 to 387 and residues 87 to 104 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry, for example, the method used to Figures 14 and 15. Representative data are shown in Figures 14 and 15. In some embodiments, the hydrogen-deuterium exchange is reduced by at least 10%, 20%, 30%, 40%, 50%, 60 %, 70%, 80%, or 90%.
[00443] [00443] In some embodiments, the antibody molecule reduces the hydrogen-deuterium exchange in one or more regions of a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 106 when bound to it, where to one or more regions are selected from the group consisting of residues 158 to 172, residues 206 to 215, residues 368 to 387 and residues 87 to 104 of SEQ ID NO: 106, for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry.
[00444] [00444] In some embodiments, the antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO : 171), leads to a greater reduction in hydrogen-deuterium exchange in residues 368 to 387 of SEQ ID NO: 105 than in residues 297 to 309 of SEQ ID NO: 105, for example, as measured by a method described in the present document, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7.5 and room temperature, for example, the method used for Figures 14 and 15. Representative data are shown in Figures 14 and 15. In some embodiments, the reduction in hydrogen-deuterium exchange in residues 368 to 387 of SEQ ID NO: 105 is at least 1.2 times, 1.3 times, 1.4 times, 1.5 times, 1.6 times, 1.7 times, 1.8 times, 1.9 times, 2 times, 2.5 times, 3 times, 3.5 times or 4 times of the reduction in hydrogen exchange iodine in residues 297 to 309 of SEQ ID NO: 105.
[00445] [00445] In some embodiments, the anti-CD73 antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 106, leads to a greater reduction in hydrogen-deuterium exchange in residues 368 to 387 of SEQ ID NO: 106 than in residues 297 to 309 of SEQ ID NO: 106, for example, as measured by a method described in the present document, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7.5 and room temperature.
[00446] [00446] In some embodiments, the anti-CD73 antibody molecule, when bound to human CD73, reduces the hydrogen-deuterium exchange in loop C of human CD73 to a greater extent than in loop A, loop B or loop D of CD73 human, for example, as measured by a method described in this document, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7.5 and room temperature, for example, the all used for Figures 14 and 15. Representative data are shown in Figures 14 and 15. In some embodiments, the reduction in hydrogen-deuterium exchange in loop C of human CD73 is at least 1.2 times, 1.3 times , 1.4 times, 1.5 times, 1.6 times, 1.7 times, 1.8 times, 1.9 times, 2 times, 2.5 times, 3 times, 3.5 times or 4 times of reduction in exchange for hydrogen-deuterium in loop A, loop B or loop D of human CD73.
[00447] [00447] In some embodiments, the anti-CD73 antibody molecule, when linked to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the hydrogen-deuterium exchange in residues 368 to 387 of SEQ ID NO: 105 to a greater extent than in residues 158 to 172, residues 206 to 215 or residues 297 to 309 of SEQ ID NO: 105, for example, as measured by a method described in this document, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7.5 and room temperature, for example, the method used for Figures 14 and 15. Representative data are shown in Figures 14 and 15. In some embodiments, the reduction in hydrogen-deuterium exchange in residues 368 to 387 of SEQ ID NO: 105 is at least 1, 2 times, 1.3 times, 14 times, 1.5 times, 1.6 times, 1.7 times, 1.8 times, 1.9 times, 2 times, 2.5 times, 3 times, 3.5 times or 4 times the reduction in hydrogen-deuterium exchange in residues 158 to 172, residues 206 to 215 or residues 297 to 309 of SEQ ID NO: 105.
[00448] [00448] In some embodiments, the anti-CD73 antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 106, reduces the hydrogen-deuterium exchange in residues 368 to 387 of SEQ ID NO: 106 to a greater extent than in residues 158 to 172, residues 206 to 215 or residues 297 to 309 of SEQ ID NO: 106, for example, as measured by a method described in this document, for example , mass spectrometry of hydrogen deuterium exchange conducted for 1 minute in exchange at pH 7.5 and room temperature.
[00449] [00449] In some embodiments, the anti-CD73 antibody molecule, when bound to human CD73, reduces the hydrogen-deuterium exchange in loop A of human CD73 to a greater extent than in loop B, loop C or loop D of human CD73, for example, as measured by a method described in this document, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7.5 and room temperature, for example, method used for Figures 14 and 15. Representative data are shown in Figures 14 and 15. In some embodiments, the reduction in hydrogen-deuterium exchange in loop A of human CD73 is at least 1.2 times, 1, 3 times, 1.4 times, 1.5 times, 1.6 times, 1.7 times, 1.8 times, 1.9 times, 2 times, 2.5 times, 3 times, 3.5 times or 4 reduction in hydrogen-deuterium exchange in loop B, loop C or loop D of human CD73.
[00450] [00450] In some embodiments, the anti-CD73 antibody molecule, when linked to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the hydrogen-deuterium exchange in residues 158 to 172 of SEQ ID NO: 105 to a greater extent than in residues 206 to 215, residues 368 to 387 or residues 297 to 309 of SEQ ID NO: 105, for example, as measured by a method described in this document, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7.5 and room temperature, for example, the method used for Figures 14 and 15. Representative data are shown in Figures 14 and 15. In some embodiments, the reduction in hydrogen-deuterium exchange in residues 158 to 172 of SEQ ID NO: 105 is at least 1, 2 times, 1.3 times, 14 times, 1.5 times, 1.6 times, 1.7 times, 1.8 times, 1.9 times, 2 times, 2.5 times, 3 times, 3.5 times or 4 times the reduction in hydrogen-deuterium exchange in residues 206 to 215, residues 368 to 387 or residues 297 to 309 of SEQ ID NO: 105.
[00451] [00451] In some embodiments, the anti-CD73 antibody molecule, when attached to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), reduces the hydrogen-deuterium exchange in residues 158 to 172 of SEQ ID NO: 106 to a greater extent than in residues 206 to 215, residues 368 to 387 or residues 297 to 309 of SEQ ID NO: 106, for example, as measured by a method described in the present document, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7.5 and room temperature.
[00452] [00452] In some embodiments, the anti-HIV antibody molecule
[00453] [00453] In some embodiments, the anti-CD73 antibody molecule, when bound to a protein that comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 106, is able to reduce the hydrogen-deuterium exchange in a or more regions of human CD73, in which one or more regions are selected from the group consisting of residues 158 to 172, residues 206 to 215, residues 368 to 387 and residues 87 to 104 of SEQ ID NO: 106, in that the region that has the greatest reduction among one or more regions does not consist of residues 206 to 215 of SEQ ID NO: 106, for example, as measured by a method described in this document, for example, mass exchange spectrometry of hydrogen deuterium conducted for 1 minute in exchange at pH 7.5 and room temperature.
[00454] [00454] In some embodiments, the anti-CD73 antibody molecule, when attached to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID
[00455] [00455] In some embodiments, the anti-CD73 antibody molecule, when bound to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO: 171), leads to a reduction in hydrogen-deuterium exchange less than, for example, 0.05, 0.04, 0.03 or 0.02 Da per residue in residues 206 to 215 of SEQ ID NO: 105, for example, as measured by a method described in this document, for example, hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7.5 and room temperature, for example, the method used for Figures 14 and 15. Representative data are shown in Figures 14 and 15.
[00456] [00456] In some embodiments, the antibody molecule that binds to a human CD73 dimer, said dimer consisting of a first CD73 monomer and a second CD73 monomer, wherein, when the antibody molecule comprises a first antigen binding domain and a second antigen binding domain, the first antigen binding domain binds to the first CD73 monomer and the second antigen binding domain binds to the second CD73 monomer, for example, when tested with the use of size exclusion chromatography, for example, when tested using a method described in this document
[00457] [00457] In some embodiments, a composition comprising a plurality of antibody molecules that binds to a human CD73 dimer is disclosed in the present document, wherein said dimer consisting of a first CD73 monomer and a second CD73 monomer, each monomer comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer consisting of the amino acid sequence of SEQ ID NO: 171), wherein, when the antibody molecules in plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at least 30%, 35%, 40%, 45% , 50%, 55%, 60%, 65%, 70%, 75%, or 80% of the antibody molecules in that composition binds to the CD73 dimer to form a complex, in which each of the said complex consists of an antibody molecule and a CD73 dimer, for example, when measured using the size exclusion chromatography ho and the percentage value is obtained by determining the amount of antibody molecules in the complex in relation to the total amount of antibody binding molecules to CD73 (excluding unbound antibody molecules), for example, when tested using a method described in this document, for example, the method used for Figures 17A, 17B, 18A and 18B. Representative data are shown in Figures 17A, 17B, 18A and 18B.
[00458] [00458] In some embodiments, a plurality of antibody molecules that bind to a human CD73 dimer is disclosed in the present document, wherein said dimer consists of a first CD73 monomer and a second CD73 monomer, each monomer comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, each monomer that contains
[00459] [00459] In some embodiments, the anti-CD73 antibody molecule, by binding to human CD73, does not lead to the oligomerization of human CD73.
[00460] [00460] In some embodiments, the antibody molecule preferably binds to an open conformation, for example, a catalytically inactive conformation, of CD73 over a closed conformation, for example, a catalytically active conformation, CD73, for example, does not bind or bind to the closed conformation, for example, the catalytically active conformation, CD73 with lower affinity, for example, 50%, 60%, 70%, 80%, 90%, 95%, or 99% lower affinity than when the antibody molecule binds to the open conformation, for example, the catalytically inactive conformation of CD73.
[00461] [00461] In some embodiments, the anti-HIV antibody molecule
[00462] [00462] In some embodiments, the anti-CD73 antibody molecule contacts, for example, directly or indirectly, at least one residue within residues 158 to 172 of SEQ ID NO: 105. In some embodiments, the antibody molecule anti-CD73 contacts, for example, directly or indirectly, at least one residue within residues 206 to 215 of SEQ ID NO: 105. In some embodiments, the anti-CD73 antibody molecule contacts, for example, directly or indirectly , at least one residue within residues 368 to 387 of SEQ ID NO: 105. In some embodiments, the anti-CD73 antibody molecule contacts, for example, directly or indirectly, at least one residue within residues 87 to 104 of SEQ ID NO: 105. In some embodiments, the anti-CD73 antibody molecule contacts, for example, directly or indirectly, at least one residue within residues 368 to 387 of SEQ ID NO: 106.
[00463] [00463] In some embodiments, the anti-CD73 antibody molecule inhibits or reduces the enzymatic activity of CD73 (e.g., soluble human CD73 or membrane-bound human CD73), for example, human CD73-mediated conversion of adenosine monophosphate ( AMP) in adenosine, for example, as measured by a method described herein, for example, a malachite green phosphate (MG) assay or a modified Cell Titration (CTG) assay, for example, the method used in the Figures. 2B,
[00464] [00464] In some embodiments, the anti-CD73 antibody molecule increases the proliferation of stimulated anti-CD3 / anti-CD28 T cells, for example, CD4 + T cells, in the presence of adenosine monophosphate (AMP), for example, as measured by a method described herein, for example, a CellTrace Violet (CTV) cell proliferation assay, for example, the method used in Figures 12A or 12B. Representative data are shown in Figures 12A and 12B. In some embodiments, the anti-CD3 / anti-CD28 stimulated T cell, for example, CD4 + T cell, proliferation is increased at least 2 times, 3 times, 4 times, 5 times, 6 times, 7 times, 8 times, 9 times or 10 times.
[00465] [00465] In another aspect, the invention provides an isolated nucleic acid that encodes any of the aforementioned antibody molecules, vectors and host cells thereof. The nucleic acid molecule includes, but is not limited to, RNA, genomic DNA and cDNA.
[00466] [00466] In some embodiments, the isolated nucleic acid encodes the antibody heavy chain variable region, light chain variable region, heavy chain and / or light chain of any of the aforementioned antibody molecules.
[00467] [00467] In some embodiments, the isolated nucleic acid encodes a variable region of heavy chain, where the nucleic acid comprises the nucleotide sequence of SEQ ID NO: 45, 78, 85, 143, 152, 160, 67, 32, 11, or 169, or a nucleotide sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 45, 78, 85, 143, 152, 160, 67, 32, 11, or
[00468] [00468] In some embodiments, the isolated nucleic acid encodes a heavy chain, wherein the nucleic acid comprises the nucleotide sequence of SEQ ID NO: 47, 80, 87, 69, 34, or 13, or a sequence of nucleotides that have at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 47, 80, 87, 69, 34, or 13.
[00469] [00469] In some embodiments, the isolated nucleic acid encodes a light chain variable region, where the nucleic acid comprises the nucleotide sequence of SEQ ID NO: 56, 144, 22 or 170, or a sequence of nucleotides that it has at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 56, 144, 22 or 170.
[00470] [00470] In some embodiments, the isolated nucleic acid encodes a light chain, wherein the nucleic acid comprises the nucleotide sequence of SEQ ID NO: 58 or 24, or a nucleotide sequence that is at least about 85% , 90%, 95%, or 99% sequence identity with SEQ ID NO: 58 or 24. Pharmaceutical Compositions and Kits
[00471] [00471] In some respects, such disclosure provides compositions, for example, pharmaceutically acceptable compositions, which include an anti-CD73 antibody molecule described herein, formulated together with a pharmaceutically acceptable carrier. As used herein, "pharmaceutically acceptable carrier" includes any and all solvents, dispersion media, isotonic or absorption retarding agents and the like that are physiologically compatible. The carrier may be suitable for intravenous, intramuscular, subcutaneous, parenteral,
[00472] [00472] The compositions presented in this document can be in a variety of forms. These include, for example, liquid, semi-solid and solid dosage forms, such as liquid solutions (for example, injectable and infusible solutions), dispersions or suspensions, liposomes and suppositories. An appropriate form depends on the intended mode of administration and therapeutic application. Typical suitable compositions are in the form of injectable or infusible solutions. A suitable mode of administration is parenteral (for example, intravenous, subcutaneous, intraperitoneal, intramuscular). In some embodiments, the antibody molecule is administered by infusion or intravenous injection. In certain modalities, the antibody is administered by intramuscular or subcutaneous injection.
[00473] [00473] The phrases "parenteral administration" and "administered parenterally" as used here mean different modes of administration from enteral and topical administration, usually by injection, and include, without limitation, intravenous, intramuscular, intra injection and infusion. -arterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intra-articular, subcapsular, subarachnoid, intraspinal, epidural and intrasternal.
[00474] [00474] Therapeutic compositions should typically be sterile and stable under conditions of manufacture and storage. The composition can be formulated as a solution, microemulsion, dispersion, liposome or other ordered structure suitable for high antibody concentration. Sterile injectable solutions can be prepared by incorporating the active compound (i.e., antibody or antibody portion) in the required amount in a suitable solvent with one or a combination of ingredients listed above, as necessary, followed by sterilization filtered. Generally
[00475] [00475] Antibody molecules can be administered by a variety of methods. Several are known in the art, and for many therapeutic applications, a suitable route / mode of administration is intravenous injection or infusion. In one embodiment, the antibody molecules can be administered by intravenous infusion at a rate of more than 20 mg / min, for example, 20 to 40 mg / min. In one embodiment, the antibody molecules can be administered by intravenous infusion at a rate greater than or equal to 40 mg / min to achieve a dose of about 35 to 440 mg / m , about 70 to 310 mg / m or about 110 to 130 mg / m2. In one embodiment, can the antibody molecules be administered by intravenous infusion at a rate of less than 10 mg / min, for example, less than or equal to 5 mg / min to achieve a dose of about 1 to 100 mg / m , about 5 to 50 mg / m , about 7 to 25 mg / m or about 10 mg / m . As will be noted by the skilled technician, the route and / or mode of administration will vary depending on the desired results. In certain modalities
[00476] [00476] In certain embodiments, an antibody molecule can be administered orally, for example, with an inert diluent or an assimilable edible carrier. The antibody molecule (and other ingredients, if desired) can also be wrapped in a hard or soft shelled gelatin capsule, compressed into tablets or incorporated directly into the individual's diet. For oral therapeutic administration, the antibody molecule can be incorporated with excipients and used in the form of digestible tablets, oral tablets, pills, capsules, elixirs, suspensions, syrups, lozenges and the like. In order to administer an antibody molecule by administration other than parenteral administration, it may be necessary to coat the compound with, or co-administer the compound with, a material to prevent its inactivation. Therapeutic compositions can also be administered with medical devices known in the art.
[00477] [00477] Dosage regimes are adjusted to provide the desired response (for example, a therapeutic response). For example, a single bolus may be administered, several divided doses may be administered over time, or the dose may be proportionate.
[00478] [00478] In some embodiments, an anti-CD73 antibody molecule disclosed in this document is administered by injection (eg, subcutaneously or intravenously) at a dose (eg, a flat dose) of about 60 mg at 2400 mg, for example, about 100 mg to 2400 mg, about 100 mg to 2200 mg, about 100 mg to 2000 mg, about 100 mg to 1800 mg, about 100 mg to 1600 mg, about 100 mg to 1400 mg, about 100 mg to 1200 mg, about 100 mg to 1000 mg, about 100 mg to 800 mg, about 100 mg to 600 mg, about 100 mg to 400 mg, about 100 mg to 200 mg or about 100 mg, about 180 mg or about 200 mg. The dosing schedule (eg, flat dosing schedule) can vary from, for example, once a week to once every 2, 3 or 4 weeks. In one embodiment, an anti-CD73 antibody molecule disclosed in this document is administered at a dose of about 100 mg to 200 mg once a week, once every two weeks or once every three weeks
[00479] [00479] In some embodiments, an anti-CD73 antibody molecule disclosed in this document is administered by injection (eg, subcutaneously or intravenously) at a dose (eg, a flat dose) of about 5 mg at 100 mg, about 100 mg to 500 mg, about 500 mg to 1000 mg, about 1000 mg to 1500 mg, about 1500 mg to 2000 mg, about 2000 mg to 2500 mg, about 2500 mg at 3000 mg, about 3000 mg to 3500 mg or about 3500 mg to 4000 mg, for example, once a week (QW), once every two weeks (Q2W) or once every four weeks ( Q4W). In certain embodiments, the antibody molecule is administered, for example, intravenously, at a dose of about 6 mg, about 20 mg, about 60 mg, about 200 mg, about 600 mg, about 1200 mg, about 2400 mg, about 3000 mg or about 3600 mg, for example, QW, Q2W or Q4W. In certain embodiments, the antibody molecule is administered, for example, intravenously, at a dose of about 60 mg Q2W. In certain embodiments, the antibody molecule is administered, for example, intravenously, at a dose of about 200 mg Q2W. In certain embodiments, the antibody molecule is administered, for example, intravenously, at a dose of about 600 mg Q2W. In certain embodiments, the antibody molecule is administered, for example, intravenously, at a dose of about 1200 mg Q2W. In certain embodiments, the antibody molecule is administered, for example, intravenously, at a dose of about 2400 mg Q2W. In certain embodiments, the antibody molecule is administered, for example, intravenously, at a dose of about 3000 mg Q2W. In certain embodiments, the antibody molecule is administered, for example, intravenously, at a dose of about 3600 mg Q2W.
[00480] [00480] In one embodiment, an anti-CD73 antibody molecule disclosed in this document is administered, for example, by infusion, over a period of 30 minutes, a period of 1 hour or a period of up to 2 hours.
[00481] [00481] A non-limiting range, exemplary for a therapeutically or prophylactically effective amount of an antibody molecule is 0.1-30 mg / kg, more preferably 1-25 mg / kg. Dosages and therapeutic regimens of the anti-CD73 antibody molecule can be determined by a skilled person. In certain embodiments, the anti-CD73 antibody molecule is administered by injection (for example, subcutaneously or intravenously) at a dose of about 1 to 40 mg / kg, for example, 1 to 30 mg / kg, for example about 5 to 25 mg / kg, about 10 to 20 mg / kg, about 1 to 5 mg / kg, 1 to 10 mg / kg, 5 to 15 mg / kg, 10 to 20 mg / kg, 15 to 25 mg / kg or about 3 mg / kg. The dosing schedule can vary, for example, once a week to once every 2, 3 or 4 weeks. In one embodiment, the anti-CD73 antibody molecule is administered at a dose of about 10 to 20 mg / kg every two weeks. The antibody molecule can be administered by intravenous infusion at a rate of more than 20 mg / min, for example, 20 to 40 mg / min, for example, greater than or equal to 40 mg / min to achieve a dose of about 35 at 440 mg / m , about 70 to 310 mg / m or about 110 to 130 mg / m2. In modalities, the infusion rate of about 110 to 130 mg / m reaches a level of about 3 mg / kg. In other embodiments, the antibody molecule can be administered by intravenous infusion at a rate of less than 10 mg / min, for example, less than or equal to 5 mg / min to achieve a dose of about 1 to 100 mg / m , for example, about 50 mg / m , about 7 to 25 mg / m or about 10 mg / m . In some embodiments, the antibody is infused over a period of about 30 min. It is to be noted that dosage values may vary with the type and severity of the condition being relieved. It is to be further understood that, for any particular subject, the specific dosage regimens must be adjusted over time according to the individual need and the professional assessment of the person administering or supervising the administration of the compositions and that the Dosage ranges presented here are exemplary only and are not intended to limit the scope or practice of the claimed composition.
[00482] [00482] The pharmaceutical compositions of the invention may include a "therapeutically effective amount" or a "prophylactically effective amount" of an antibody molecule of the invention. A "therapeutically effective amount" refers to an effective amount, at the dosages and for periods of time necessary, to achieve the desired therapeutic result. A therapeutically effective amount of the modified antibody or antibody fragment can vary according to factors such as the condition of the disease, age, sex and weight of the subject and the ability of the antibody or portion of antibody to elicit a desired response in the individual. A therapeutically effective amount is also one in which any toxic or harmful effects of the antibody molecule are outweighed by the therapeutically beneficial effects. A "therapeutically effective dosage" preferably inhibits a measurable parameter by at least about 20%, more preferably by at least about 40%, even more preferably by at least about 60% and even more preferably by at least about 80% in relation to untreated individuals. The measurable parameter can be, for example, tumor growth rate or pathogen growth rate. The ability of an antibody molecule to inhibit a measurable parameter can be assessed in an animal model system predictive of efficacy in the corresponding human disease. Alternatively, that property of a composition can be assessed by examining the ability of the compound to inhibit such an inhibition in vitro by assays known to the skilled clinician.
[00483] [00483] A "prophylactically effective amount" refers to an effective amount, in the dosages and for periods of time necessary, to achieve the desired prophylactic result. Typically, since a prophylactic dose is used in subjects before or at an earlier stage of the disease, the prophylactically effective amount will be less than the therapeutically effective amount.
[00484] [00484] A kit comprising an antibody molecule described here is also within the scope of the invention. The kit can include one or more other elements including: instructions for use; other reagents, for example, an identification, a therapeutic agent or an agent useful for chelation or otherwise coupling an antibody to an identification or therapeutic agent, or a radioprotective composition; devices or other materials for preparing the antibody molecule for administration; pharmaceutically acceptable carriers; and devices or other materials for administration to an individual. Uses of Anti-CD73 Antibody Molecules
[00485] [00485] The anti-CD73 antibody molecules disclosed in this document are diagnosed in vitro and in vivo, as well as therapeutic and prophylactic uses. For example, these molecules can be administered to cells in culture, in vitro or ex vivo, or to an individual, for example, a human individual, to treat, prevent and / or diagnose a variety of disorders, such as cancers and infectious disorders.
[00486] [00486] “Consequently, in one aspect, the invention provides a method for modifying an immune response in an individual which comprises administering to the individual the antibody molecule described herein, so that the immune response in the individual is modified. In one embodiment, the immune response is intensified, stimulated or over-regulated. In one embodiment, the antibody molecules enhance an immune response in an individual by blocking CD73.
[00487] [00487] “As used herein, the term" individual "is intended to include human and non-human animals. In a fashion, the individual is a human individual, for example, a human patient who has a disorder or condition characterized by abnormal CD73 functioning. The term "non-human animals" includes mammals and non-mammals, such as non-human primates. In a modality, the subject is a human. In one modality, the individual is a human patient who needs an intensified immune response. In one embodiment, the individual is immunocompromised, for example, the individual is subjected to or has undergone chemotherapy or radiation therapy. Alternatively, or in combination, the subject is, or is at risk of, being immunocompromised as a result of an infection. The methods and compositions described in this document are suitable for treating human patients who have a disorder that can be treated by increasing the T cell-mediated immune response. For example, the methods and compositions described in this document can enhance numerous activities. immune. In one embodiment, the individual has an increased number or activity of tumor-infiltrating T lymphocytes (TILs). Therapeutic Uses Cancer
[00488] [00488] In one aspect, the invention relates to the treatment of an individual in vivo with the use of an anti-CD73 antibody molecule so that the growth of cancerous tumors is inhibited or re-
[00489] [00489] — Consequently, in one embodiment, the invention provides a method for inhibiting the growth of tumor cells in an individual, which comprises administering to the individual a therapeutically effective amount of an anti-CD73 antibody molecule described in the this document.
[00490] [00490] In one embodiment, the methods are suitable for the treatment of cancer in vivo. To achieve antigen-specific immune enhancement, the anti-CD73 antibody molecule can be administered together with an antigen of interest. When antibodies to CD73 are administered in combination with one or more agents, the combination can be administered in order or simultaneously. Types of cancer
[00491] [00491] In another aspect, a method to treat an individual, for example, to reduce or ameliorate, a hyperproliferative disorder or disorder (for example, a cancer), for example, solid tumor, a hematological cancer, soft tissue tumor or a metastatic lesion in an individual is provided. The method includes administering to the individual one or more anti-CD73 antibody molecules described in this document, alone or in combination with other therapeutic agents or modalities.
[00492] [00492] “As used here, the term" cancer "is intended to include all types of cancerous growths or oncogenic processes, metastatic tissues or malignant-transformed cells, tissues or organs, regardless of the histopathological type or stage of the invasion. Examples of cancerous disorders include, but are not limited to, solid tumors, hematological cancers, soft tissue tumors and metastatic lesions. Examples of solid tumors include malignancies, for example, sarcomas and carcinomas (including adenocarcinomas and squamous cell carcinomas), of the various organ systems, such as those that affect the liver, lung, breast, lymphoid, gastrointestinal (eg colon), gastrourinary tract (for example, kidney, urothelial cells), prostate and pharynx. Adenocarcinomas include malignancies such as most colon cancers, rectal cancer, renal cell carcinoma, liver cancer, non-small cell lung cancer, small intestine cancer and esophageal cancer. Squamous cell carcinomas include malignancies, for example, in the region of the lung, esophagus, skin, head and neck, oral cavity, anus and cervix. In a fashion, cancer is a melanoma, for example, an advanced stage melanoma. Metastatic lesions from the aforementioned cancers can also be treated or prevented using the methods and compositions of the invention.
[00493] [00493] Exemplary cancers whose growth can be inhibited with the use of the antibody molecules disclosed in this document include cancers typically responsive to immunotherapy. Non-limiting examples of preferred cancers for treatment include melanoma (eg, metastatic malignant melanoma), kidney cancer (eg, clear cell carcinoma), prostate cancer (eg, hormonal refractory prostate adenocarcinoma) nal), breast cancer, colon cancer and lung cancer (eg, non-small cell lung cancer). Additionally,
[00494] [00494] Examples of other cancers that can be treated include bone cancer, pancreatic cancer, skin cancer, head or neck cancer, malignant cutaneous or intraocular melanoma, uterine cancer, ovarian cancer, rectal cancer, anal cancer, cancer gastroesophageal cancer, stomach cancer, testicular cancer, uterine cancer, fallopian tube carcinoma, endometrial carcinoma, cervical carcinoma, vagina carcinoma, vulva carcinoma, Merkel cell cancer, Hodgkin's lymphoma , non-Hodgkin's lymphoma, cancer of the esophagus, cancer of the small intestine, cancer of the endocrine system, cancer of the thyroid gland, cancer of the parathyroid gland, cancer of the adrenal gland, soft tissue sarcoma, cancer of the urethra, cancer of the penis, chronic leukemias or acute including acute myeloid leukemia, chronic myeloid leukemia, acute lymphoblastic leukemia, chronic lymphocytic leukemia, solid childhood tumors, lymphocytic lymphoma, bladder cancer, multiple myeloma, myelodysplastic syndromes, cancer of the kidney or ureter, carcinoma of the renal pelvis, neoplasm of the central nervous system (CNS), primary CNS lymphoma, tumor angiogenesis, spinal axis tumor, brain stem glioma, pituitary adenoma, sarcoma of Kaposi, epidermoid cancer, squamous cell cancer, T cell lymphoma, environmentally induced cancers including those induced by asbestos (eg, mesothelioma) and combinations of said cancers.
[00495] [00495] In some modalities, the therapies here can be used to treat a patient who has (or is identified as having) a cancer associated with an infection, for example, a viral or bacterial infection. Exemplary cancers include cervical cancer, anal cancer, HPV-associated head and neck squamous cell cancer, HPV-associated esophageal papillomas, associated lymphomas
[00496] [00496] In other modalities, cancer is a malignancy or hematological cancer including, but not limited to, leukemia or lymphoma. For example, the anti-CD73 antibody molecule can be used to treat cancers and malignancies including, but not limited to, for example, acute leukemias including, but not limited to, for example, acute B-cell lymphoid leukemia ("BALL" ), acute T-cell lymphoid leukemia ("TALL"), acute lymphoid leukemia (ALL); one or more chronic leukemias including, but not limited to, for example, chronic myelogenous leukemia (CML), chronic lymphocytic leukemia (CLL); additional haematological cancers or haematological disorders including, but not limited to, for example, B-cell prolinoflitic leukemia, blast plasmacytoid dendritic cell neoplasm, Burkitt's lymphoma, large diffuse B-cell lymphoma, follicular lymphoma, hair cell leukemia , small cell or large cell follicular lymphoma, malignant lymphoproliferative disorders, MALT lymphoma, mantle cell lymphoma, marginal zone lymphoma, multiple myeloma, myelodysplasia and myelodysplastic syndrome, non-Hodgkin's lymphoma, plasmablastic lymphoma, dendritic cell neoplasm , Waldenstrom's macroglobulinemia and "pre-leukemia" which are a diverse collection of hematological disorders linked by ineffective production (or dysplasia) of myeloid blood cells and the like.
[00497] [00497] In one embodiment, the cancer is chosen from a lung cancer (for example, a non-small cell lung cancer (NSCLC) (for example, an NSCLC with squamous and / or non-squamous histology or an adenocarcinoma NSCLC)), a melanoma (eg, advanced melanoma), kidney cancer (eg, renal cell carcinoma, eg, clear cell renal cell carcinoma), liver cancer, myeloma (for example, example, a multiple myeloma), a prostate cancer, a breast cancer (for example, a breast cancer that does not express one, two or all of the estrogen receptor, progesterone receptor or Her2 / neu, for example , triple negative breast cancer), colorectal cancer (eg, microsatellite stable colorectal cancer (MSS)), ovarian cancer, pancreatic cancer, head and neck cancer (eg, carcinoma of head and neck squamous cell (HNSCC), anal cancer, gastroesophageal cancer, thyroid cancer, cancer r cervical, lymphoproliferative disease (eg, post-transplant lymphoproliferative disease) or hematological cancer, T-cell lymphoma, non-Hodgkin's lymphoma or leukemia (for example, myeloid leukemia).
[00498] [00498] In one embodiment, cancer is chosen from lung cancer (for example, non-small cell lung cancer), pancreatic cancer (for example, pancreatic ductal adenocarcinoma), cancer of breast (for example, triple negative breast cancer), melanoma, head and neck cancer (for example, scaly head and neck cancer), colorectal cancer (for example, microsatellite stable colorectal cancer (MSS)), cancer ovarian or kidney cancer (eg kidney cell carcinoma).
[00499] [00499] In one embodiment, the cancer is chosen from bladder cancer, leukemia, lymphoma, glioma, glioblastoma, ovarian cancer, thyroid cancer, esophageal cancer, prostate cancer, uterine / cervical cancer, testicular cancer , esophageal cancer, gastrointestinal cancer, colon cancer, kidney cancer, stomach cancer, germ cell cancer, bone cancer, liver cancer, skin cancer, central nervous system neoplasm, myeloma, sarcoma and related cancer - linked to viruses. Combination of Anti-CD73 Antibody Molecules
[00500] [00500] The anti-CD73 antibody molecules can be used in combination with other therapies. For example, combination therapy may include a composition of the present invention co-formulated with, and / or co-administered with, one or more additional therapeutic agents, for example, one or more anticancer agents, cytotoxic or cytostatic agents, hormonal treatment, vaccines and / or other immunotherapies. In other modalities, the antibody molecules are administered in combination with other therapeutic treatment modalities, including surgery, radiation, cryosurgery and / or thermotherapy. Such combination therapies can advantageously use the lower dosages of the administered therapeutic agents, thus avoiding possible toxicities or complications associated with the various monotherapies.
[00501] [00501] The term "in combination with" is not intended to imply that the therapy or therapeutic agents must be administered at the same time and / or formulated for delivery together, although these delivery methods are within the scope described in the present document. The anti-CD73 antibody molecules can be administered simultaneously with, before or subsequent to one or more other therapies or additional therapeutic agents. The anti-CD73 antibody molecule and the other therapeutic agent or protocol can be administered in any order. In general, each agent will be administered at a dose and / or on a specific time schedule for that agent. It will be further appreciated that the therapeutic agent used in this combination can be administered together in a single composition or administered separately in different compositions. In general, additional therapeutic agents used in combination are expected to be used at levels that do not exceed the levels at which they are used individually. In some embodiments, the levels used in combination will be lower than those used individually. Exemplary A2A Adenosine Receptor Antagonists
[00502] [00502] In certain embodiments, the anti-CD73 molecules described herein are administered in combination with an adenosine A2A receptor antagonist (A2AR). Exemplary A2AR antagonists include, for example, PBF509 (Palobio-pharma / Novartis), CPI444 / V81444 (Corvus / Genentech), AZD4635 / HTL-1071 (Astrazeneca / Heptares), Vipadenante (Redox / Juno), GBV- 2034 (Globavir), AB928 (Arcus Biosciences), Theophylline, Istradefiline (Kyowa Hakko Kogyo), Tozadenante / SYN-115 (Wake up), KW-6356 (Kyowa Hakko Kogyo), ST-4206 (Leadiant Biosciences) and Preladenante / SCH 420 (Merck / Schering).
[00503] [00503] In certain embodiments, the APAR antagonist is PBF509. PBF509 and other A2AR antagonists are disclosed in U.S. No. 8,796,284 and WO 2017/025918, hereby incorporated by reference in their entirety. PBF509 refers to 5-bromo-2,6-di- (1H-pyrazol-1-yl) pyrimidine-4-amine with the following structure: it is JN to JJ nº
[00504] [00504] In certain embodiments, the A2AR antagonist is CPI444 / V81444. CPI-444 and other A2AR antagonists are disclosed in WO 2009/156737, incorporated in this document as a reference in its entirety. In certain embodiments, the A2AR antagonist is (S) -7- (5-methylfuran-2-yl) -3 - ((6 - ((((tetrahydrofuran-3-yl) oxy) Methyl) pyridin- 2-yl) methyl) -3H- [1,2,3] triazolo [4,5-d] pyrimidin-5-amine. In certain embodiments, the A2AR antagonist is (R) -7- (S-methylfuran-2-yl) -3 - ((6 - ((((tetrahydrofuran-3-yl) oxy) Methyl) pyridin-2-
[00505] [00505] In certain embodiments, the A2AR antagonist is AZDA4635 / HTL-1071. A2AR antagonists are disclosed in the document WO 2011/095625, incorporated in this document as a reference in its entirety. In certain embodiments, the AZAR antagonist is 6- (2-chloro-6-methylpyridin-4-i1) -5- (4-fluorophenyl) -1,2,4-triazin-3-amine. In certain embodiments, the A2AR antagonist has the following structure: Steel AN IT IS
[00506] [00506] In certain embodiments, the APAR antagonist is ST-4206 (Leadiant Biosciences). In certain embodiments, the AZAR antagonist is an A2AR antagonist described in U.S. No. 9,133,197, hereby incorporated by reference in its entirety for reference. In certain embodiments, the APAR antagonist has the following structure: NH,
[00507] [00507] In certain embodiments, the APAR antagonist is an A2AR antagonist described in documents No. U.S. 8114845, U.S. 9029393, U.S. 20170015758 or U.S. 20160129108, incorporated herein by reference in its entirety.
[00508] [00508] In certain embodiments, the A2AR antagonist is istradefilline (CAS Registry Number: 155270-99-8). Istradefiline is also known as KW-6002 or 8 - [(E) -2- (3,4-dimethoxyphenyl) vinyl] -1,3-diethyl-7-methyl-3,7-dihydro-1H-purine- 2,6-dione. Istradefiline is disclosed, for example, in LeWitt et al. (2008) Annals of Neurology 63 (3): 295 to 302).
[00509] [00509] In certain modalities, the A2aR antagonist is toxic (Biotie). Tozadenant is also known as SYN115 or 4-hydroxy-N- (4-methoxy-7-morpholin-4-yl-1,3-benzothiazol-2-i1) -4-methylpiperidine-1-carboxamide. Tozadenant blocks the effect of endogenous adenosine on A2a receptors, resulting in potentiation of the dopamine effect on the D2 receptor and inhibition of the glutamate effect on the MGIuR5 receptor. In some modalities, the A2aR antagonist is preladenant (CAS Registry Number: 377727-871-2). Preladenant is also known as SCH 420814 or 2- (2-Furanyl) -7- [2- [4- [4- (2-methoxyethoxy) phenyl] -1-piperazinyl] ethyl] 7H-pyrazole [4,3- and] [1,2,4] triazolo [1,5-c] pyrimidine-5-amine. The Prelate was developed as a drug that acted as a potent and selective antagonist at the A2A adenosine receptor.
[00510] [00510] In certain modalities, the A2aR antagonist is vipade-
[00511] [00511] Other exemplary A2aR antagonists include, for example, ATL-444, MSX-3, SCH-58261, SCH-412,348, SCH-442,416, VER-6623, VER-6947, VER-7835, CGS-15943 or ZM -241,385.
[00512] [00512] In some embodiments, the A2aR antagonist is an A2aR pathway antagonist (for example, a CD-73 inhibitor, for example, an anti-CD73 antibody) is MEDI9447. MEDI9447 is a monoclonal antibody specific for CD73. Targeting extracellular adenosine production by CD73 can reduce the immunosuppressive effects of adenosine. MEDI9447 has been reported to have a range of activities, for example, inhibition of CD73 ectonucleotidase activity, AMP-mediated lymphocyte suppression relief and inhibition of singene tumor growth. MEDI9447 can drive changes in both myeloid and lymphoid infiltrating leukocyte populations within the tumor microenvironment. These changes include, for example, an increase in CD8 effector cells and activated macrophages, as well as a reduction in myeloid-derived suppressor cell (MDSC) and regulatory T lymphocyte propositions. Exemplary PD-1 inhibitors
[00513] [00513] In certain embodiments, the anti-CD73 antibody molecule described herein is administered in combination with a PD-1 inhibitor. The PD-1 inhibitor can be an antibody, an antigen binding fragment thereof, an immunoadhesin, a fusion protein or an oligopeptide. In some embodiments, the PD-1 inhibitor is chosen from PDROO1 (Novartis), Nivolumabe (Bristol-Myers Squibb), Pembrolizumab (Merck & Co), Pidilizumabe (CureTech), MEDIO680 (Medimmune), REGN2810 (Regeneron), TSR-042 (Tesaro), PF-O06801591 (Pfizer), BGB-A317 (Beigene), BGB-
[00514] [00514] In one embodiment, the PD-1 inhibitor is an anti-PD-1 antibody molecule. In one embodiment, the PD-1 inhibitor is an anti-PD-1 antibody molecule as described in US 2015/0210769, published on July 30, 2015, entitled "Antibody Molecules to PD- 1 and Uses Thereof,", incorporated by reference in its entirety.
[00515] [00515] In one embodiment, the anti-PD-1 antibody molecule comprises at least one, two, three, four, five or six complementarity determining regions (CDRs) (or collectively all CDRs) of a variable chain region heavy and light chain comprising an amino acid sequence shown in Table 5 (for example, the heavy and light chain variable region sequences of BAPO49-Clone-E or BAPO49-Clone-B disclosed in Table 5), or encoded by a nucleotide sequence shown in Table 5. In some embodiments, CDRs conform to the definition of Kabat (for example, as shown in Table 5). In some modalities, CDRs are in accordance with the definition of Chothia (for example, as shown in Table 5). In some modalities, CDRs conform to the combined CDR definitions of both Kabat and Chothia (for example, as shown in Table 5). In one embodiment, the combination of Kabat's CDR and VH CDR1's Chothia comprises the GYTFTTYWMH amino acid sequence (SEQ ID NO: 541). In one embodiment, one or more of the CDRs (or collectively all of the CDRs) have one, two, three, four, five, six or more changes, for example, amino acid substitutions (for example, conservative amino acid substitutions) or deletions, with respect to an amino acid sequence shown in Table 5, or encoded by a nucleotide sequence shown in Table 5.
[00516] [00516] In one embodiment, the anti-PD-1 antibody molecule comprises a heavy chain variable region (VH) comprising a VÓHCDR1 amino acid sequence of SEQ ID NO: 501, a VHCDR2 amino acid sequence of SEQ ID NO: 502 and a VOMCDR3 amino acid sequence of SEQ ID NO: 503 and a light chain variable region (VL) comprising a VLCDR1 amino acid sequence of SEQ ID NO: 510, a sequence of VLCDR2 amino acids of SEQ ID NO: 511 and a VLCDR3 amino acid sequence of SEQ ID NO: 512, each disclosed in Table 5.
[00517] [00517] In one embodiment, the antibody molecule comprises a VH comprising a VHCDR1 encoded by the nucleotide sequence of SEQ ID NO: 524, a VHCDR 2 encoded by the nucleotide sequence of SEQ ID NO: 525 and a VHCDR3 encoded by the nucleotide sequence of SEQ ID NO: 526; and a VL comprising a VLCDR1 encoded by the nucleotide sequence of SEQ ID NO: 529, a VLCDR 2 encoded by the nucleotide sequence of SEQ ID NO: 530 and a VLCDR3 encoded by the nucleotide sequence of SEQ ID NO: 530 : 531, each disclosed in Table 5.
[00518] [00518] In one embodiment, the anti-PD-1 antibody molecule comprises a VH that comprises the amino acid sequence of SEQ ID NO: 506, or an amino acid sequence that is at least about 85%, 90%, 95 %, or 99% sequence identity or more with SEQ ID NO: 506. In one embodiment, the anti-PD-1 antibody molecule comprises a VL comprising the amino acid sequence of SEQ ID NO: 520, or a amino acid sequence that has at least about 85%, 90%, 95%, or 99% sequence identity or greater with SEQ ID NO: 520. In one embodiment, the anti-PD-1 antibody molecule comprises a VL that understand
[00519] [00519] In one embodiment, the antibody molecule comprises a VH encoded by the nucleotide sequence of SEQ ID NO: 507, or a nucleotide sequence that is at least about 85%, 90%, 95%, or 99% sequence identity or more with SEQ ID NO: 507. In one embodiment, the antibody molecule comprises a VL encoded by the nucleotide sequence of SEQ ID NO: 521 or 517, or a nucleotide sequence that is at least about 85%, 90%, 95%, or 99% sequence identity or more with SEQ ID NO: 521 or 517. In one embodiment, the antibody molecule comprises a VH encoded by the nucleotide sequence of SEQ ID NO : 507 and a VL encoded by the nucleotide sequence of SEQ ID NO: 521 or 517.
[00520] [00520] In one embodiment, the anti-PD-1 antibody molecule comprises a heavy chain that comprises the amino acid sequence of SEQ ID NO: 508, or an amino acid sequence that is at least about 85%, 90%, 95%, or 99% sequence identity or greater with SEQ ID NO: 508. In one embodiment, the anti-PD-1 antibody molecule comprises a light chain comprising the amino acid sequence of SEQ ID NO: 522, or a amino acid sequence that is at least about 85%, 90%,
[00521] [00521] In one embodiment, the antibody molecule comprises a heavy chain encoded by the nucleotide sequence of SEQ ID NO: 509, or a nucleotide sequence that is at least about 85%, 90%, 95%, or 99% of sequence identity or more with SEQ ID NO: 509. In one embodiment, the antibody molecule comprises a light chain encoded by the nucleotide sequence of SEQ ID NO: 523 or 519, or a nucleotide sequence that has at least about 85%, 90%, 95%, or 99% sequence identity or more with SEQ ID NO: 523 or 519. In one embodiment, the antibody molecule comprises a heavy chain encoded by the nucleotide sequence of SEQ ID NO: 509 and a light chain encoded by the nucleotide sequence of SEQ ID NO: 523 or 519.
[00522] [00522] The antibody molecules described in this document can be produced by vectors, host cells and methods described in document No. US 2015/0210769, incorporated by reference in their entirety.
[00523] [00523] In one embodiment, the anti-PD-1 antibody molecule is Nivolumab (Bristol-Myers Squibb), also known as MDX-1106, MDX-1106-04, ONO-4538, BMS-936558 or OPDIVOG. Nivolumab (clone 5C4) and other anti-PD-1 antibodies are disclosed in US
[00524] [00524] In one embodiment, the anti-PD-1 antibody molecule is Pembrolizumab (Merck & Co), also known as Lambrolizumab, MK-3475, MK03475, SCH-900475 or KEYTRUDAG. Pembroli- zumab and other anti-PD-1 antibodies are disclosed in Hamid, O. et al. (2013) New England Journal of Medicine 369 (2): 13444, No. US. 8,354,509 and WO 2009/114335, incorporated by reference in their entirety. In one embodiment, the anti-PD-1 antibody molecule comprises one or more of the CDR sequences (or collectively all of the CDR sequences), the heavy chain or light chain variable region sequence or the heavy chain sequence or Pembrolizumab light chain, for example, as disclosed in Table 6.
[00525] [00525] In one embodiment, the anti-PD-1 antibody molecule is Pidilizumab (CureTech), also known as CT-011. Pidilizumab and other anti-PD-1 antibodies are disclosed in Rosenblatt, J. et al. (2011) J Immunotherapy 34 (5): 409-18, U.S. No. 7,695,715, U.S. No. 7,332,582 and U.S. No. 8,686,119, incorporated by reference in their entirety. In one embodiment, the anti-PD-1 antibody molecule comprises one or more of the CDR sequences (or collectively all of the CDR sequences), the heavy chain or light chain variable region sequence or the heavy chain sequence or Pidilizumab light chain, for example, as disclosed in Table 6.
[00526] [00526] In one embodiment, the anti-PD-1 antibody molecule is MEDIO680 (Medimmune), also known as AMP-514. ME-DIO0680 and other anti-PD-1 antibodies are disclosed in US 9,205,148 and WO 2012/145493, incorporated by reference in their entirety. In one embodiment, the anti-PD-1 antibody molecule comprises one or more of the CDR sequences (or collectively all of the CDR sequences), the heavy chain or light chain variable region sequence, or the DNA sequence. heavy chain or light chain of ME-DI0680.
[00527] [00527] In one embodiment, the anti-PD-1 antibody molecule is REGN2810 (Regeneron). In one embodiment, the anti-PD-1 antibody molecule comprises one or more of the CDR sequences (or collectively all of the CDR sequences), the heavy chain or light chain variable region sequence or the heavy chain sequence or REGN2810 light chain.
[00528] [00528] In one embodiment, the anti-PD-1 antibody molecule is PF-06801591 (Pfizer). In one embodiment, the anti-PD-1 antibody molecule comprises one or more of the CDR sequences (or collectively all of the CDR sequences), the heavy chain or light chain variable region sequence, or the heavy chain or light chain of PF-06801591.
[00529] [00529] In one embodiment, the anti-PD-1 antibody molecule is BGB-A317 or BGB-108 (Beigene). In one embodiment, the anti-PD-1 antibody molecule comprises one or more of the CDR sequences (or collectively all of the CDR sequences), the heavy chain or light chain variable region sequence or the heavy chain or chain sequence lightweight BGB-A317 or BGB-108.
[00530] [00530] In one embodiment, the anti-PD-1 antibody molecule is INCSHR1210 (Incyte), also known as INCSHRO01210 or SHR-1210. In one embodiment, the anti-PD-1 antibody molecule comprises one or more of the CDR sequences (or collectively all of the CDR sequences), the heavy chain or light chain variable region sequence or the heavy chain sequence or light chain of INCSHR1210.
[00531] [00531] In one embodiment, the anti-PD-1 antibody molecule is TSR-042 (Tesaro), also known as ANBO11. In one fashion, the anti-PD-1 antibody molecule comprises one or more of the CDR sequences (or collectively all of the CDR sequences), the heavy chain or light chain variable region sequence or the heavy chain sequence or TSR-042 light chain.
[00532] [00532] Other anti-PD-1 antibodies include those described, for example, in WO 2015/112800, WO 2016/092419, WO 2015/085847, WO 2014/179664, WO 2014/194302, WO 2014/209804, WO 2015 / 200119, US 8,735,553, US 7,488,802, US 8,927,697, US 8,993,731 and US 9,102,727, incorporated by reference in their entirety.
[00533] [00533] In one embodiment, the anti-PD-1 antibody is an antibody that competes for binding and / or binds to the same epitope on PD-1 as one of the anti-PD-1 antibodies described herein.
[00534] [00534] In one embodiment, the PD-1 inhibitor is a peptide that inhibits the PD-1 signaling pathway, for example, as described in U.S. No. 8,907,053, incorporated by reference in its entirety. In one embodiment, the PD-1 inhibitor is an immunoadhesin (for example, an immunoadhesin comprising an extracellular binding or PD-1 portion of PD-L1 or PD-L2 fused to a constant region (for example , an Fc region of an immunoglobulin sequence). In one embodiment, the PD-1 inhibitor is AMP-224 (B7-DClIg (Amplimmune), for example, disclosed in WO 2010/027827 and WO 2011/066342, incorporated by reference in their entirety).
[00535] [00535] In certain embodiments, the anti-CD73 antibody molecule described herein is administered in combination with a PD-L1 inhibitor. The PD-L1 inhibitor can be an antibody, an antigen binding fragment thereof, an immunoadhesin, a fusion protein or an oligopeptide. In some embodiments, the PD-L1 inhibitor is chosen from FAZO53 (Novartis), Atezolizumab (Genentech / Roche), Avelumab (Merck Serono and Pfizer), Durvalumabe (MedlImmune / AstraZeneca) or BMS-936559 (Bristol- Myers Squibb). Exemplary anti-PD-L1 Antibody Molecules
[00536] [00536] In one embodiment, the PD-L1 inhibitor is an anti-PD-L1 antibody molecule. In one embodiment, the PD-L1 inhibitor is an anti-PD-L1 antibody molecule as disclosed in document No. US 2016/0108123, published on April 21, 2016, entitled "Antibody Molecules to PD-L1 and Uses Thereof ' , incorporated by reference in its entirety.
[00537] [00537] In one embodiment, the anti-PD-L1 antibody molecule comprises at least one, two, three, four, five or six complementarity determining regions (CDRs) (or collectively all CDRs) of a variable chain region heavy and light chain comprising an amino acid sequence shown in Table 7 (for example, the heavy and light chain variable region sequences of BAPOS8-Clone O or BAPOS58-Clone N disclosed in Table 7), or encoded by a sequence of nucleotides shown in Table 7. In some embodiments, CDRs conform to the Kabat definition (for example, as shown in Table 7). In some modalities, CDRs are in accordance with the definition of Chothia (for example, as shown in Table 7). In some modalities, CDRs conform to the combined CDR definitions of both Kabat and Chothia (for example, as shown in Table 7). In one embodiment, the combination of Kabat's CDR and VH CDR1's Chothia comprises the GYTFTSYWMY amino acid sequence (SEQ ID NO: 647). In one embodiment, one or more of the CDRs (or collectively all of the CDRs) have one, two, three, four, five, six or more changes, for example, amino acid substitutions (for example, conservative amino acid substitutions) or deletions, with respect to an amino acid sequence shown in Table 7, or encoded by a nucleotide sequence shown in Table 7.
[00538] [00538] In one embodiment, the anti-PD-L1 antibody molecule comprises a variable heavy chain (VH) region comprising a VÓHCDR1 amino acid sequence of SEQ ID NO: 601, a VHCDR2 amino acid sequence of SEQ ID NO: 602 and a VÓMCDR3 amino acid sequence of SEQ ID NO: 603 and a light chain variable region (VL) comprising a VLCDR1 amino acid sequence of SEQ ID NO: 609, a sequence of VLCDR2 amino acids of SEQ ID NO: 610 and a VLCDR3 amino acid sequence of SEQ ID NO: 611, each disclosed in Table 7.
[00539] [00539] In one embodiment, the anti-PD-L1 antibody molecule comprises a VH comprising a VHCDR1 encoded by the nucleotide sequence of SEQ ID NO: 628, a VHCDR 2 encoded by the nucleotide sequence of SEQ ID NO : 629, and a VHCDR3 encoded by the nucleotide sequence of SEQ ID NO: 630; and a VL comprising a VLCDR1 encoded by the nucleotide sequence of SEQ ID NO: 633, a VLCDR 2 encoded by the nucleotide sequence of SEQ ID NO: 634 and a VLCDR3 encoded by the nucleotide sequence of SEQ ID NO: 635 , each disclosed in Table 7.
[00540] [00540] In one embodiment, the anti-PD-L1 antibody molecule comprises a VH that comprises the amino acid sequence of SEQ ID NO: 606, or an amino acid sequence that is at least about 85%, 90%, 95 %, or 99% sequence identity with SEQ ID NO: 606. In one embodiment, the anti-PD-L1 antibody molecule comprises a VL that comprises the amino acid sequence of SEQ ID NO: 616, or an amino acid sequence that it has at least about 85%, 90%, 95%, or 99% sequence identity or more with SEQ ID NO: 616. In one embodiment, the anti-PD-L1 antibody molecule comprises a VH comprising the amino acid sequence of SEQ ID NO: 620, or an amino acid sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 620. In one embodiment, the anti-PD-L1 antibody molecule comprises a VL that comprises the amino acid sequence of SEQ ID NO: 624, or an amino acid sequence that is at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 624. In one embodiment, the anti-PD-L1 antibody molecule comprises a VH comprising the amino acid sequence of SEQ ID NO: 606 and a VL comprising the amino acid sequence of SEQ ID NO:
[00541] [00541] In one embodiment, the antibody molecule comprises a VH encoded by the nucleotide sequence of SEQ ID NO: 607, or a nucleotide sequence that has at least about 85%, 90%, 95%, or 99% identity with SEQ ID NO:
[00542] [00542] In one embodiment, the anti-PD-L1 antibody molecule comprises a heavy chain that comprises the amino acid sequence of SEQ ID NO: 608, or an amino acid sequence that is at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 608. In one embodiment, the anti-PD-L1 antibody molecule comprises a light chain comprising the amino acid sequence of SEQ ID NO: 618, or a sequence of amino acids that have at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 618. In one embodiment, the anti-PD-L1 antibody molecule comprises a heavy chain comprising the sequence amino acid sequence of SEQ ID NO: 622, or an amino acid sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 622. In one embodiment, the antibody molecule anti-PD-L1 comprises a light chain that comprises the amino acid sequence of SEQ ID NO: 626, or an amino acid sequence that has pe it minus about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 626. In one embodiment, the anti-PD-L1 antibody molecule comprises a heavy chain that comprises the sequence of amino acids of SEQ ID NO: 608 and a light chain comprising the amino acid sequence of SEQ ID NO: 618. In one embodiment, the anti-PD-L1 antibody molecule comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 622 and a light chain comprising the amino acid sequence of SEQ ID NO: 626.
[00543] [00543] In one embodiment, the antibody molecule comprises a heavy chain encoded by the nucleotide sequence of SEQ ID NO: 615, or a nucleotide sequence that is at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 615. In one embodiment, the antibody molecule comprises a light chain encoded by the nucleotide sequence of SEQ ID NO: 619, or a nucleotide sequence that is at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 619. In one embodiment, the antibody molecule comprises a heavy chain encoded by the nucleotide sequence of SEQ ID NO: 623, or a nucleotide sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 623. In one embodiment, the antibody molecule comprises a light chain encoded by the nucleotide sequence of SEQ ID NO: 627, or a nucleotide sequence that is at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 627. In one embodiment, the antibody molecule comprises a heavy chain encoded by the nucleotide sequence of SEQ ID NO: 615 and a light chain encoded by the nucleotide sequence of SEQ ID NO: 619. In one embodiment, the antibody molecule comprises a heavy chain encoded by the nucleotide sequence of SEQ ID NO: 623 and a light chain encoded by the nucleotide sequence of SEQ ID NO: 627.
[00544] [00544] The antibody molecules described in this document can be produced by vectors, host cells and methods described in document No. US 2016/0108123, incorporated by reference in their entirety.
[00545] [00545] In one embodiment, the anti-PD-L1 antibody molecule is Atezolizumab - (Genentech / Roche), also known as MPDL3280A, RG7446, RO5541267, YW243.55.870 or TECEN-TRIQTY, Atezolizumab and other anti-PD- antibodies L1 are disclosed in document No. US 8,217,149, which is incorporated by reference in its entirety. In one embodiment, the anti-PD-L1 antibody molecule comprises one or more of the CDR sequences (or collectively all of the CDR sequences), the heavy chain or light chain variable region sequence or the heavy chain sequence or Atezolizumab light chain, for example, as disclosed in Table 8.
[00546] [00546] In one embodiment, the anti-PD-L1 antibody molecule is Avelumab (Merck Serono and Pfizer), also known as MSBO0010718C. Avelumab and other anti-PD-L1 antibodies are disclosed in document No. WO 2013/079174, incorporated by reference in its entirety. In one embodiment, the anti-PD-L1 antibody molecule comprises one or more of the CDR sequences (or collectively all of the CDR sequences), the heavy chain or light chain variable region sequence or the heavy chain sequence or Avelumab light chain, for example, as disclosed in Table 8.
[00547] [00547] In one embodiment, the anti-PD-L1 antibody molecule is Durvalumabe (MediImmune / AstraZeneca), also known as MEDI4736. Durvalumab and other anti-PD-L1 antibodies are disclosed in US No. 8,779,108, which is incorporated by reference in its entirety. In one embodiment, the anti-HIV antibody molecule
[00548] [00548] In one embodiment, the anti-PD-L1 antibody molecule is BMS-936559 (Bristol-Myers Squibb), also known as MDX-1105 or 12A4. BMS-936559 and other anti-PD-L1 antibodies are disclosed in US 7,943,743 and WO 2015/081158, incorporated by reference in their entirety. In one embodiment, the anti-PD-L1 antibody molecule comprises one or more of the CDR sequences (or collectively all of the CDR sequences), the heavy chain or light chain variable region sequence, or the heavy chain or light chain of BMS-936559, for example, as disclosed in Table 8.
[00549] [00549] The additionally known anti-PD-L1 antibodies include those described, for example, in WO 2015/181342, WO 2014/100079, WO 2016/000619, WO 2014/022758, WO 2014/055897, WO 2015 / 061668, WO 2013/079174, WO 2012/145493, WO 2015/112805, WO 2015/109124, WO 2015/195163, US 8,168,179, US 8,552,154, US 8,460,927 and US 9,175,082, incorporated by reference in their entirety .
[00550] [00550] In one embodiment, the anti-PD-L1 antibody is an antibody that competes for binding and / or binds to the same epitope on PD-L1 as one of the anti-PD-L1 antibodies described herein. Table 8. Amino acid sequences of other exemplary anti-PD-L1 antibody molecules "Tue KIHTCPPCPAPELLGGPSVELFPPKPKDTLMISRTPEVTOWVDVSHED-. PEVKFNWYVDGVEVHNAKTKPREE- JAYASTYRVVSVLTVLHODWLNGKEYKCKVSNKALPAPIEKTISKA- KGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESN- / GAQPENNYKTTPPVLDSDGSFFLYSKLTVDKS
[00551] [00551] In certain embodiments, the anti-CD73 molecule described herein is administered in combination with a LAG-3 inhibitor known in the art. The LAG-3 inhibitor can be an antibody, an antigen binding fragment thereof, an immunoadhesin, a fusion protein or oligopeptide. In some embodiments, the LAG-3 inhibitor is chosen from LAG525 (Novartis), BMS-986016 (Bristol-Myers Squibb), TSR-033 (Tesaro), MK-4280 (Merck & Co) or REGN3767 ( Regeneron).
[00552] [00552] In one embodiment, the LAG-3 inhibitor is an anti-LAG-3 antibody molecule. In one embodiment, the LAG-3 inhibitor is an anti-LAG-3 antibody molecule as disclosed in document No. US 2015/0259420, published on September 17, 2015, entitled "Antibody Molecules to LAG-3 and Uses Thereof ", incorporated by reference in its entirety.
[00553] [00553] In one embodiment, the anti-LAG-3 antibody molecule comprises at least one, two, three, four, five or six complementarity determining regions (CDRs) (or collectively all CDRs) of a variable chain region heavy and light chain comprising an amino acid sequence shown in Table 9 (for example, from the BAPOSO-Clone | or BAPOSO-CIone J variable region sequences disclosed in Table 9), or encoded by a sequence of nucleotides shown in Table 9. In some embodiments, CDRs conform to the Kabat definition (for example, as shown in Table 9). In some modalities, CDRs are in accordance with the definition of Chothia (for example, as shown in Table 5). In some modalities, CDRs conform to the combined CDR definitions of both Kabat and Chothia (for example, as shown in Table 9). In one embodiment, the combination of Kabat's CDR and VH CDR1's Chothia comprises the GFTLTNYGMN amino acid sequence (SEQ ID NO: 766). In one embodiment, one or more of the CDRs (or collectively all of the CDRs) have one, two, three, four, five, six or more changes, for example, amino acid substitutions (for example, conservative amino acid substitutions) or deletions, with respect to an amino acid sequence shown in Table 9, or encoded by a nucleotide sequence shown in Table 9.
[00554] [00554] In one embodiment, the anti-LAG-3 antibody molecule comprises a heavy chain variable region (VH) comprising a VÓHCDR1 amino acid sequence of SEQ ID NO: 701, a VHCDR2 amino acid sequence of SEQ ID NO: 702 and a VÓMCDR3 amino acid sequence of SEQ ID NO: 703 and a light chain variable region (VL) comprising a VLCDR1 amino acid sequence of SEQ ID NO: 710, a sequence of VLCDR2 amino acids of SEQ ID NO: 711 and a VLCDR3 amino acid sequence of SEQ ID NO: 712, each disclosed in Table 9.
[00555] [00555] In one embodiment, the anti-LAG-3 antibody molecule comprises a VH comprising a VHCDR1 encoded by the nucleotide sequence of SEQ ID NO: 736 or 737, a VMHCDR2 encoded by the nucleotide sequence of SEQ ID NO: 738 or 739, and a VHCDR3 encoded by the nucleotide sequence of SEQ ID NO: 740 or 741; and a VL comprising a VLCDR1 encoded by the nucleotide sequence of SEQ ID NO: 746 or 747, a VLCDR 2 encoded by the nucleotide sequence of SEQ ID NO: 748 or 749, and a VLCDR3 encoded by the nucleotide sequence of SEQ ID NO: 750 or 751, each disclosed in Table 9. In one embodiment, the anti-LAG-3 antibody molecule comprises a VH comprising a VHCDR1 encoded by the nucleotide sequence of SEQ ID NO: 758 or 737, a VHCDR 2 encoded by the nucleotide sequence of SEQ ID NO: 759 or 739, and a VHCDR3 encoded by the nucleotide sequence of SEQ ID NO: 760 or 741; and a VL comprising a VLCDR1 encoded by the nucleotide sequence of SEQ ID NO: 746 or 747, a VLCDR2 encoded by the nucleotide sequence of SEQ ID NO: 748 or 749, and a VLCDR3 encoded by the nucleotide sequence of SEQ ID NO: 750 or 751, each disclosed in Table 9.
[00556] [00556] In one embodiment, the anti-LAG-3 antibody molecule comprises a VH that comprises the amino acid sequence of SEQ ID NO: 706, or an amino acid sequence that is at least about 85%, 90%, 95 %, or 99% sequence identity with SEQ ID NO: 706. In one embodiment, the anti-LAG-3 antibody molecule comprises a VL that comprises the amino acid sequence of SEQ ID NO: 718, or an amino acid sequence that it has at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 718. In one embodiment, the anti-LAG-3 antibody molecule comprises a VH that comprises the sequence of amino acids of SEQ ID NO: 724, or an amino acid sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 724. In one embodiment, the anti-antibody molecule -LAG-3 comprises a VL that comprises the amino acid sequence of SEQ ID NO: 730, or an amino acid sequence that has at least about 85%, 90%, 95%, or u 99% sequence identity with SEQ ID NO: 730. In one embodiment, the anti-LAG-3 antibody molecule comprises a VH comprising the amino acid sequence of SEQ ID NO: 706 and a VL comprising the amino acid sequence SEQ ID NO:
[00557] [00557] In one embodiment, the antibody molecule comprises a VH encoded by the nucleotide sequence of SEQ ID NO: 707 or 708, or a nucleotide sequence that is at least about 85%, 90%, 95%, or 99 % sequence identity with SEQ ID NO: 707 or 708. In one embodiment, the antibody molecule comprises a VL encoded by the nucleotide sequence of
[00558] [00558] In one embodiment, the anti-LAG-3 antibody molecule comprises a heavy chain that comprises the amino acid sequence of SEQ ID NO: 709, or an amino acid sequence that is at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 709. In one embodiment, the anti-LAG-3 antibody molecule comprises a light chain comprising the amino acid sequence of SEQ ID NO: 721, or a sequence of amino acids that have at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 721. In one embodiment, the anti-LAG-3 antibody molecule comprises a heavy chain that comprises the sequence amino acids of SEQ ID NO: 727, or an amino acid sequence that is at least about 85%,
[00559] [00559] In one embodiment, the antibody molecule comprises a heavy chain encoded by the nucleotide sequence of SEQ ID NO: 716 or 717, or a nucleotide sequence that is at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 716 or 717. In one embodiment, the antibody molecule comprises a light chain encoded by the nucleotide sequence of SEQ ID NO: 722 or 723, or a sequence of nucleotides that it has at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 722 or 723. In one embodiment, the antibody molecule comprises a heavy chain encoded by the sequence nucleotides of SEQ ID NO: 728 or 729, or a nucleotide sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 728 or
[00560] [00560] The antibody molecules described in this document can be produced by vectors, host cells and methods described in document No. US 2015/0259420, incorporated by reference in its entirety. Table 9. Amino acid and nucleotide sequences of exemplary anti-LAG-3 antibody molecules HE of Basa re SEQD NO: 701 (Kabat SEQD NO: 702 (Kabat) SEQ1D NO: 703 (Kabat SEQ ID NO: 704 (Chothia 'SEQD NO: 705 (Chothia) SEQ ID NO: 703 (Chothia HCDR3 NPPYYYGTNNAEAMDY SEQ ID NO: 706 VvH QVQALVOSGAEVKKPGASVKVSCKASGFTLTNYGMNWVROQAR- GQORLEWIGWINTDTGEVYYDD
[00561] [00561] In one embodiment, the anti-LAG-3 antibody molecule is BMS-986016 (Bristol-Myers Squibb)) also known as BMSS986016. BMS-986016 and other anti-LAG-3 antibodies are disclosed in documents No. WO 2015/116539 and US 9,505,839, incorporated by reference in their entirety. In one embodiment, the anti-LAG-3 antibody molecule comprises one or more of the CDR sequences (or collectively all of the CDR sequences), the heavy chain or light chain variable region sequence or the heavy chain sequence or BMS-986016 light chain, for example, as disclosed in Table 10.
[00562] [00562] In one embodiment, the anti-LAG-3 antibody molecule is TSR-033 (Tesaro). In one embodiment, the anti-LAG-3 antibody molecule comprises one or more of the CDR sequences (or collectively all of the CDR sequences), the heavy chain or light chain variable region sequence or the heavy chain sequence or TSR-033 light chain.
[00563] [00563] In one embodiment, the anti-LAG-3 antibody molecule is MK-4280 (Merck & Co). In one embodiment, the anti-LAG-3 antibody molecule comprises one or more of the CDR (or co-sequences)
[00564] [00564] In one embodiment, the anti-LAG-3 antibody molecule is REGN3767 (Regeneron). In one embodiment, the anti-LAG-3 antibody molecule comprises one or more of the CDR sequences (or collectively all of the CDR sequences), the heavy chain or light chain variable region sequence or the heavy chain sequence or REGN3767 light chain.
[00565] [00565] In one embodiment, the anti-LAG-3 antibody molecule is IMP731 or GSK2831781 (GSK and Prima BioMed). IMP731 and other anti-LAG-3 antibodies are disclosed in documents No. WO 2008/132601 and US 9,244,059, incorporated by reference in their entirety. In one embodiment, the anti-LAG-3 antibody molecule comprises one or more of the CDR sequences (or collectively all of the CDR sequences), the heavy chain or light chain variable region sequence or the heavy chain sequence or IMP731 light chain, for example, as disclosed in Table
[00566] [00566] In one embodiment, the anti-LAG-3 antibody molecule is IMP761 (Prima BioMed). In one embodiment, the anti-LAG-3 antibody molecule comprises one or more of the CDR sequences (or collectively all of the CDR sequences), the heavy chain or light chain variable region sequence, or the heavy chain or IMP761 light chain.
[00567] [00567] Other known anti-LAG-3 antibodies include those described, for example, in documents No. WO 2008/132601, WO 2010/019570, WO 2014/140180, WO 2015/116539, WO 2015/200119, WO 2016/028672 , US 9,244,059, US 9,505,839, incorporated by reference in their entirety.
[00568] [00568] In one embodiment, the anti-LAG-3 antibody is an antibody that competes for binding and / or binds to the same epitope in LAG-3 as one of the anti-LAG-3 antibodies described herein.
[00569] [00569] In one embodiment, the anti-LAG-3 inhibitor is a soluble LAG-3 protein, for example, IMP321 (Prima BioMed), for example, as disclosed in document No. WO 2009/044273, incorporated by way of reference in its entirety.
[00570] [00570] In certain embodiments, the anti-CD73 antibody molecule described herein is administered in combination with a TIM-3 inhibitor. The TIM-3 inhibitor can be an antibody, an antigen binding fragment thereof, an immunoadhesin, a fusion protein or an oligopeptide. In some embodiments, the TIM-3 inhibitor is chosen from MGB453 (Novartis), TSR-022 (Tesaro) or LY3321367 (Eli Lilly). Exemplary anti-TIM-3 Antibody Molecules
[00571] [00571] In one embodiment, the TIM-3 inhibitor is an anti-TIM-3 antibody molecule. In one embodiment, the TIM-3 inhibitor is an anti-TIM-3 antibody molecule as disclosed in document No. US 2015/0218274, published on August 6, 2015, entitled "Antibody Molecules to TIM-3 and Uses Thereof" , incorporated by reference in its entirety.
[00572] [00572] In one embodiment, the anti-TIM-3 antibody molecule comprises at least one, two, three, four, five or six complementarity determining regions (CDRs) (or collectively all CDRs) in a variable chain region heavy and light that comprises an amino acid sequence shown in Table 11 (for example, the ABTIM3-hum11 or ABTIM3-humO3 variable region sequences disclosed in Table 11), or encoded by a nucleotide sequence shown in Table 11. In some modalities, CDRs are in accordance with the definition of Kabat (for example, as shown in Table 11). In some modalities, CDRs are in accordance with the Chothia definition (for example, as shown in Table 11). In one embodiment, one or more of the CDRs (or collectively all of the CDRs) have one, two, three, four, five, six or more changes, for example, amino acid substitutions (for example, conservative amino acid substitutions) or deletions, with respect to an amino acid sequence shown in Table 11, or encoded by a nucleotide sequence shown in Table 11.
[00573] [00573] In one embodiment, the anti-TIM-3 antibody molecule comprises a heavy chain variable region (VH) comprising a VÓHCDR1 amino acid sequence of SEQ ID NO: 801, a VHCDR2 amino acid sequence of SEQ ID NO: 802 and a VÓMCDR3 amino acid sequence of SEQ ID NO: 803 and a light chain variable region (VL) comprising a VLCDR1 amino acid sequence of SEQ ID NO: 810, a sequence of VLCDR2 amino acids of SEQ ID NO: 811 and a VLCDR3 amino acid sequence of SEQ ID NO: 812, each disclosed in Table 11. In one embodiment, the anti-TIM-3 antibody molecule comprises a variable region of heavy chain (VH) comprising a VHCDR1 amino acid sequence of SEQ ID NO: 801, a VOMCDR2 amino acid sequence of SEQ ID NO: 820 and a VÓMCDCD3 amino acid sequence of SEQ ID NO: 803 and a variable region of light chain (VL) comprising a VLCDR1 amino acid sequence of SEQ ID NO: 810, a sequence VLCDR2 amino acid sequence of SEQ ID NO: 811 and a VLCDR3 amino acid sequence of SEQ ID NO: 812, each disclosed in Table 11.
[00574] [00574] In one embodiment, the anti-TIM-3 antibody molecule comprises a VH that comprises the amino acid sequence of SEQ ID NO: 806, or an amino acid sequence that is at least about 85%, 90%, 95 %, or 99% sequence identity with SEQ ID NO: 806. In one embodiment, the anti-TIM-3 antibody molecule comprises a VL comprising the amino acid sequence of SEQ ID NO: 816, or a sequence of amino acids that have at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 816. In one embodiment, the anti-TIM-3 antibody molecule comprises a VH that comprises the amino acid sequence of SEQ ID NO: 822, or an amino acid sequence
[00575] [00575] In one embodiment, the antibody molecule comprises a VH encoded by the nucleotide sequence of SEQ ID NO: 807, or a nucleotide sequence that has at least about 85%, 90%, 95%, or 99% identity with SEQ ID NO:
[00576] [00576] In one embodiment, the anti-TIM-3 antibody molecule comprises a heavy chain that comprises the amino acid sequence of SEQ ID NO: 808, or an amino acid sequence that is at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 808. In one embodiment, the anti-TIM-3 antibody molecule comprises a light chain comprising the amino acid sequence of SEQ ID NO: 818, or a sequence of amino acids that have at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 818. In one embodiment, the anti-TIM-3 antibody molecule comprises a heavy chain that comprises the sequence amino acid sequence of SEQ ID NO: 824, or an amino acid sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 824. In one embodiment, the antibody molecule anti-TIM-3 comprises a light chain that comprises the amino acid sequence of SEQ ID NO: 828, or an amino acid sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 828. In one embodiment, the anti-TIM-3 antibody molecule comprises a heavy chain comprising the sequence of amino acids of SEQ ID NO: 808 and a light chain comprising the amino acid sequence of SEQ ID NO: 818. In one embodiment, the anti-TIM-3 antibody molecule comprises a heavy chain comprising the sequence of amino acids of SEQ ID NO: 824 and a light chain comprising the amino acid sequence of SEQ ID NO: 828.
[00577] [00577] In one embodiment, the antibody molecule comprises a heavy chain encoded by the nucleotide sequence of SEQ ID NO: 809, or a nucleotide sequence that is at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 809. In one embodiment, the antibody molecule comprises a light chain encoded by the nucleotide sequence of SEQ ID NO: 819, or a nucleotide sequence that is at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 819. In one embodiment, the antibody molecule comprises a heavy chain encoded by the nucleotide sequence of SEQ ID NO: 825, or a nucleotide sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 825. In one embodiment, the antibody molecule comprises a light chain encoded by the nucleotide sequence of SEQ ID NO: 829, or a nucleotide sequence that is at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 829. In one embodiment, the antibody molecule comprises a heavy chain encoded by the nucleotide sequence of SEQ ID NO: 809 and a light chain encoded by the nucleotide sequence of SEQ ID NO: 819. In one embodiment, the antibody molecule comprises a heavy chain encoded by the nucleotide sequence of SEQ ID NO: 825 and a light chain encoded by the nucleotide sequence of SEQ ID NO: 829.
[00578] [00578] The antibody molecules described in this document can be produced by vectors, host cells and methods described in document No. US 2015/0218274, incorporated by reference in their entirety.
[00579] [00579] In one embodiment, the anti-TIM-3 antibody molecule is
[00580] [00580] In one embodiment, the anti-TIM-3 antibody molecule is LY3321367 (Eli Lilly). In one embodiment, the anti-TIM-3 antibody molecule comprises one or more of the CDR sequences (or collectively all of the CDR sequences), the heavy chain or light chain variable region sequence, or the heavy chain or light chain of LY3321367.
[00581] [00581] In one embodiment, the anti-TIM-3 antibody molecule is the F38-2E2 antibody clone. In one embodiment, the anti-TIM-3 antibody molecule comprises one or more of the CDR sequences (or collectively all of the CDR sequences), the heavy chain or light chain variable region sequence, or the heavy chain or light chain of F38-2E2.
[00582] [00582] - Other known anti-TIM-3 antibodies include those described, for example, in documents No. WO 2016/111947, WO 2016/071448, WO 2016/144803, US No. 8,552,156, US No. 8,841,418 and US No. 9,163 .087, incorporated by reference in their entirety.
[00583] [00583] In one embodiment, the anti-TIM-3 antibody is an antibody that competes for binding and / or binds to the same epitope in TIM-3 as one of the anti-TIM-3 antibodies described herein.
[00584] [00584] In certain embodiments, the anti-CD73 antibody molecule described herein is administered in combination with a CTLA-4 inhibitor. The CTLA-4 inhibitor can be an antibody, an antigen-binding fragment thereof, an immunomandine, a fusion protein or an oligopeptide. In some modalities, the CTLA-4 inhibitor is Ipilimumab (Yervoyº, Bristol-Myers Squibb) or Tremelimumab (Pfizer). The Ipilimumab antibody and other anti-cCTLA-4 antibodies are disclosed in document No. UJS.
[00585] [00585] In certain embodiments, the anti-CD73 antibody molecule described herein is administered in combination with a GITR agonist. The GITR agonist can be an antibody, an antigen binding fragment thereof, an immunoadhesin, a fusion protein or an oligopeptide. In some embodiments, the GITR agonist is GWN323 (Novartis), BMS-986156 (BMS), MK-4166 or MK-1248 (Merck), TRX518 (Leap Therapeutics), IN-CAGN1876 (Incyte / Agenus), AMG 228 ( Amgen) or INBRX-110 (Inhi- brx). Exemplary Anti-GITR Antibody Molecules
[005868] [005868] In one embodiment, the GITR agonist is an anti-GITR antibody molecule. In one embodiment, the GITR agonist is an anti-GITR antibody molecule as described in WO 2016/057846, published on April 14, 2016, entitled "Compositions and Methods of Use for Augmented Immune Response and Cancer Therapy" , incorporated by reference in its entirety.
[00587] [00587] In one embodiment, the anti-GITR antibody molecule comprises at least one, two, three, four, five or six complementarity determining regions (CDRs) (or collectively all CDRs) of a variable region of heavy and light chain comprising an amino acid sequence shown in Table 13 (for example, from the MAB7 heavy and light chain variable region sequences disclosed in Table 13), or encoded by a nucleotide sequence shown in Table 13. In some modalities, CDRs are in accordance with the definition of Kabat (for example, as shown in Table 13). In some modalities, CDRs are in accordance with the Chothia definition (for example, as shown in Table 13). In one embodiment, one or more of the CDRs (or collectively all of the CDRs) have one, two, three, four, five, six or more changes, for example, amino acid substitutions (for example, conservative amino acid substitutions) or deletions, relative to an amino acid sequence shown in Table 13, or encoded by a nucleotide sequence shown in Table 13.
[00588] [00588] In one embodiment, the anti-GITR antibody molecule comprises a heavy chain variable region (VH) comprising a VHCDR1 amino acid sequence of SEQ ID NO: 909, a VHCDR 2 amino acid sequence of SEQ ID NO: 911 and a VÓHCDR3 amino acid sequence of SEQ ID NO: 913 and a light chain variable region (VL) comprising a VLCDR1 amino acid sequence of SEQ ID NO: 914, a sequence of VLCDR2 amino acids of SEQ ID NO: 916 and a VLCDR3 amino acid sequence of SEQ ID NO: 918, each disclosed in Table 13.
[00589] [00589] In one embodiment, the anti-GITR antibody molecule comprises a VH comprising the amino acid sequence of SEQ ID NO: 901, or an amino acid sequence of at least 85%, 90%, 95%, or 99% identical or more SEQ ID NO: 901. In one mode, the anti-GITR antibody molecule comprises a VL comprising the amino acid sequence of SEQ ID NO: 902, or an amino acid sequence of at least 85%, 90% , 95%, or 99% identical or more to SEQ ID NO: 902. In one embodiment, the anti-GITR antibody molecule comprises a VH comprising the amino acid sequence of SEQ ID NO: 901 and a VL comprising the amino acid sequence of SEQ ID NO: 902.
[00590] [00590] In one embodiment, the antibody molecule comprises a VH encoded by the nucleotide sequence of SEQ ID NO: 905, or a nucleotide sequence of at least 85%, 90%, 95%, or 99% identical or more to SEQ ID NO: 905. In one embodiment, the antibody molecule comprises a VL encoded by the nucleotide sequence of SEQ ID NO: 906, or a nucleotide sequence of at least 85%, 90%, 95%, or 99% identical or more SEQ ID NO: 906. In one embodiment, the antibody molecule comprises a VH encoded by the nucleotide sequence of SEQ ID NO: 905 and a VL encoded by the nucleotide sequence of SEQ ID
[00591] [00591] In one embodiment, the anti-GITR antibody molecule comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 903, or an amino acid sequence of at least 85%, 90%, 95%, or 99% identical or more to SEQ ID NO:
[00592] [00592] In one embodiment, the antibody molecule comprises a heavy chain encoded by the nucleotide sequence of SEQ ID NO: 907, or a nucleotide sequence of at least 85%, 90%, 95%, or 99% identical or more to SEQ ID NO: 907. In one mode, the antibody molecule comprises a light chain encoded by the nucleotide sequence of SEQ ID NO: 908, or a nucleotide sequence of at least 85%, 90%, 95 %, or 99% identical or more to SEQ ID NO: 908. In one embodiment, the antibody molecule comprises a heavy chain encoded by the nucleotide sequence of SEQ ID NO: 907 and a light chain encoded by the nucleotide sequence of SEQ ID NO: 908.
[00593] [00593] The antibody molecules described in this document can be produced by vectors, host cells and methods described in document No. WO 2016/0578486, incorporated by reference in its entirety.
[00594] [00594] In one embodiment, the anti-GITR antibody molecule is BMS-986156 (Bristol-Myers Squibb), also known as BMS 986156 or BMS986156. BMS-986156 and other anti-GITR antibodies are disclosed in, for example, documents U.S. No. 9,228,016 and WO 2016/196792, incorporated by reference in their entirety. In one embodiment, the anti-GITR antibody molecule comprises one or more of the CDR sequences (or collectively all of the CDR sequences), the heavy chain or light chain variable region sequence or the heavy chain sequence or BMS-986156 light chain, for example, as disclosed in Table 14.
[00595] [00595] In one embodiment, the anti-GITR antibody molecule is MK-4166 or MK-1248 (Merck). MK-4166, MK-1248 and other anti-GITR antibodies are disclosed, for example, in U.S. No. documents
[00596] [00596] In one embodiment, the anti-GITR antibody molecule is TRX518 (Leap Therapeutics). TRX518 and other anti-GITR antibodies are disclosed, for example, in U.S. No. 7,812,135, U.S. No. 8,388,967, U.S. No. 9,028,823, WO 2006/105021, and Ponte J et al. (2010) Clinical Immunology; 135: 896, incorporated by reference in their entirety. In one embodiment, the anti-GITR antibody molecule comprises one or more of the CDR sequences (or collectively all of the CDR sequences), the heavy chain or light chain variable region sequence, or the heavy chain sequence or TRX518 light chain.
[00597] [00597] In one embodiment, the anti-GITR antibody molecule is INCAGN1876 (Incyte / Agenus). INCAGN1876 and other anti-GITR antibodies are disclosed, for example, in documents No. US 2015/0368349 and WO 2015/184099, incorporated by reference in their entirety. In one embodiment, the anti-GITR antibody molecule comprises one or more of the CDR sequences (or collectively all of the CDR sequences), the heavy chain or light chain variable region sequence or the heavy chain or chain sequence INCAGN1876 light weight.
[00598] [00598] In one embodiment, the anti-GITR antibody molecule is AMG 228 (Amgen). AMG 228 and other anti-GITR antibodies are disclosed in, for example, U.S. Documents 9,464,139 and WO 2015/031667, incorporated by reference in their entirety. In one embodiment, the anti-GITR antibody molecule comprises one or more of the CDR sequences (or collectively all of the CDR sequences), the heavy chain or light chain variable region sequence, or the heavy chain or light chain sequence of AMG 228.
[00599] [00599] In one embodiment, the anti-GITR antibody molecule is INBRX-110 (Inhibrx). INBRX-110 and other anti-GITR antibodies are disclosed, for example, in documents No. US 2017/0022284 and WO 2017/015623, incorporated by reference in their entirety. In one embodiment, the GITR agonist comprises one or more of the CDR sequences (or collectively all of the CDR sequences), the heavy chain or light chain variable region sequence or the INBRX-110 heavy chain or light chain sequence .
[00600] [00600] In one embodiment, the GITR agonist (for example, a fusion protein) is MEDI 1873 (Medimmune), also known as MEDI1873. MEDI 1873 and other GITR agonists are disclosed, for example, in U.S. No. 2017/0073386, WO 2017/025610, and Ross et al. Cancer Res 2016; 76 (14 Suppl): Abstract nr 561, incorporated by reference in its entirety. In one embodiment, the GITR agonist comprises one or more of an IgG Fc domain, a functional multimerization domain, and a receptor binding domain of a MEDI glucocorticoid-induced TNF receptor ligand (GITRL) 1873.
[00601] [00601] Other known GITR agonists (for example, anti-GITR antibodies) include those described, for example, in document No. WO 2016/054638, incorporated by reference in their entirety.
[00602] [00602] In one embodiment, the anti-GITR antibody is an antibody that competes for binding and / or binds to the same epitope in GITR as one of the anti-GITR antibodies described herein.
[00603] [00603] In one embodiment, the GITR agonist is a peptide that activates the GITR signaling pathway. In one embodiment, the GITR agonist is an immunoadhesin binding fragment (for example, an immunoadhesin binding fragment comprising a GITR or extracellular GITRL binding portion) fused to a constant region (for example, an Fc region immunoglobulin sequence). Table 14. Amino acid sequences of other exemplary anti-GITR antibody molecules
[00604] [00604] In certain embodiments, the anti-CD73 antibody molecule described herein is administered in combination with an anti-CD3 multispecific antibody molecule (for example, a bispecific anti-cCD3 antibody molecule). In one embodiment, the multispecific anti-CD3 antibody molecule binds to CD3 and a tumor target antigen (TTA). In one embodiment, TTA is chosen from CD19, CD20, CD38 or CD123. In one embodiment, the anti-CD3 multispecific antibody molecule is in a format disclosed in Figures 1A, 1B, 1C and 125 of WO 2016/182751, incorporated in this document as a reference in its entirety.
[00605] [00605] In one embodiment, the anti-CD3 multispecific antibody molecule is an anti-CD3 x anti-CD123 bispecific antibody molecule, for example, XENP14045 (for example, as shown in Table 15) or an antibody molecule bispecific anti-CD3 x anti-CD123 disclosed in WO 2016/086189 or WO 2016/182751, incorporated herein by reference in its entirety. In one embodiment, the bispecific anti-CD3 x anti-CD123 antibody molecule comprises one or more of the CDR sequences (or collectively all of the CDR sequences), the heavy chain or light chain variable region sequence, or the sequence heavy chain or light chain of XENP14045, or an amino acid sequence substantially identical to the same (e.g., a sequence that has at least about 85%, 90%, or 95% sequence identity to it).
[00606] [00606] In one embodiment, the anti-CD3 multispecific antibody molecule is a bispecific anti-CD3 x anti-CD20 antibody, for example, XENP13676 (for example, as shown in Table 15) or a bispecific anti-antibody molecule -CD3 x anti-CD20 published in WO 2016/086189 or WO 2016/182751, incorporated in this document as a reference in its entirety. In one embodiment, the bispecific anti-CD3 x anti-CD20 antibody molecule comprises one or more of the CDR sequences (or collectively all the CDR sequences), the heavy chain or light chain variable region sequence, or the sequence heavy chain or light chain of XENP13676, or an amino acid sequence substantially identical to the same (for example, a sequence that has at least about 85%, 90%, or 95% sequence identity to the same) . Table 15. Amino acid sequences of exemplary anti-CD3 bispecific antibody molecules XENP14045 (Fab-) scFv-Fc anti-CD123 x anti-CD3 SEQ ID NO: 177 Heavy chain 1 / QVOLQAOSGAEVKKPGASVKVSCKASGYT- (Fab-Fc anti-CD123 ) | FTDYYMKWVKQASHGKSLEWMGDIIPSNGATFYNQKFKGKATLTV- DRSTSTAYMELSSLRSEDTAVYYCARSHLLRASWFAYWGQG- TLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVK- DYFPEPVTVSWNSGALTSGVHTFPAVLOSSGLYSLSSVVTVPSSS- LGTQTYICNVNHKPSDTKVDKKVEPKSCDKTHTCPPCPAPPVAG- PSVFLFPPKPKDTLMISRTPEVTCVVVDVKHEDPEVKFNWYVD- GVEVHNAKTKPREEEYNSTYRVVSVLTVLHQDWLNGKEYK- 'CKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVS- LTCDVSGFYPSDIAVEWESDGQPENNYKTTPPVLDSDGSFFLYS-
[00607] [00607] In certain embodiments, the anti-CD73 antibody molecule described herein is administered in combination with an IL-15 / IL-15Ra complex. In some embodiments, the IL-15 / IL-15Ra complex is chosen from NIZ985 (Novartis), ATL-803 (Altor) or CYPO0150 (Cytune).
[00608] [00608] In one embodiment, the IL-15 / IL-15Ra complex comprises human IL-15 complexed with a soluble form of human IL-15Ra. The complex can comprise IL-15 covalently or non-covalently linked to a soluble form of IL-15Ra. In a particular embodiment, human IL-15 is non-covalently linked to a soluble form of IL-15Ra. In a particular embodiment, the human I1L-15 of the composition comprises an amino acid sequence of SEQ ID NO: 183 in Table 16 and the soluble form of human IL-15Ra comprises an amino acid sequence of SEQ ID NO: 184 in Table 16, as described in WO 2014/066527, incorporated by reference in its entirety. The molecules described in this document can be produced by vectors, host cells and methods described in document No. WO 2007/084342, incorporated by reference in its entirety. Table 16. Exemplary amino acid and nucleotide sequences of IL-15 / IL-15Ra complexes NIZ985 LESGDASIHDTVENLIILANNSLSSNGNVTESGCKECEELEEKNIKEFLOQS:
[00609] [00609] In one embodiment, the IL-15 / IL-15Ra complex is ALT-803, an IL-15 / IL-15Ra Fc fusion protein (soluble IL-15N72D complex: IL-15RaSu / Fc). ALT-803 is disclosed in WO 2008/143794, incorporated by reference in its entirety. In one embodiment, the IL-15 / IL-15Ra Fc fusion protein comprises the sequences as disclosed in Table 17.
[00610] [00610] In one embodiment, the IL-15 / IL-15Ra complex comprises IL-15 fused to the sushi domain of IL-15Ra (CYP0150, Cytan). The IL-15Ra sushi domain refers to a domain that begins at the first cysteine residue after the | IL- signaling peptide
[00611] [00611] In certain embodiments, the anti-CD73 antibody molecule described herein is administered in combination with a STING agonist. In some embodiments, the combination is used to treat a cancer, for example, a cancer described in this document, for example, a solid tumor (for example, a breast cancer, a squamous cell carcinoma, a melanoma, a ovarian cancer, fallopian tube carcinoma, peritoneal carcinoma, soft tissue sarcoma, esophageal cancer, head and neck cancer, endometrial cancer, cervical cancer or basal cell carcinoma), for example , a haematological malignancy (eg, leukemia (eg, chronic lymphocytic leukemia (CLL) or lymphoma (eg, marginal zone B cell lymphoma, small lymphocytic lymphoma, follicular lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma))). In some modalities, cancer is chosen from head and neck cancer (eg, head and neck squamous cell carcinoma (HNSCC)), skin cancer (eg melanoma) ), lung cancer (for example, non-small cell lung cancer (NSCLC)), or breast cancer (for example, triple negative breast cancer (TNBC)).
[00612] [00612] In some embodiments, the STING agonist is cyclic dinucleotide, for example, a cyclic dinucleotide comprising purine or pyrimidine nucleobases (for example, adenosine, guanine, uracil, thymine or cytosine nucleobases). In some modalities, the cyclic dinucleotide nucleobases comprise the same or different nucleobases.
[00613] [00613] In some embodiments, the STING agonist comprises an adenosine or guanosine nucleobase. In some modalities, the STING agonist comprises an adenosine nucleobase and a guanosine nucleobase. In some embodiments, the STING agonist comprises two adenosine nucleobases or two guanosine nucleobases.
[00614] [00614] In some embodiments, the STING agonist comprises a modified cyclic dinucleotide, for example, which comprises a modified nucleobase, a modified ribose or a modified phosphate bond. In some embodiments, the modified cyclic dinucleotide comprises a modified phosphate bond, for example, a thiophosphate.
[00615] [00615] In some embodiments, the STING agonist comprises a cyclic dinucleotide (for example, a modified cyclic dinucleotide) with 2 ', 5' or 3 ', 5' phosphate bonds. In some modalities, the STING agonist comprises a cyclic dinucleotide
[00616] [00616] In some embodiments, the STING agonist is Rp, Rp dithio 2 ', 3' c-di-AMP (for example, Rp, Rp-dithio c- [A (2 ', 5') DA (3 ' , 5 ') P]), or a cyclic dinucleotide analog thereof. In some embodiments, the STING agonist is a compound represented in U.S. Patent Publication No. 2015/0056224 (for example, a compound in Figure 2c, for example, compound 21 or compound 22). In some modalities, the STING agonist is c- [G (2 ', 5') DG (3 ', 5') p], an O- substituted dithio ribose derivative of the same or a compound shown in Figure 4 PCT Publications No. WO 2014/189805 and WO 2014/189806. In some embodiments, the STING agonist is c- [A (2 ', 5') DA (3 ', 5') D] or an O-substituted dithio ribose derivative thereof, or is a compound shown in Figure 5 PCT Publications No. WO 2014/189805 and WO 2014/189806. In some modalities, the STING agonist is c- [G (2 ', 5') DA (3 ', 5') p] or an O-substituted dithio ribose derivative thereof, or is a compound represented in Figure 5 of PCT Publications No. WO 2014/189805 and WO 2014/189806. In some embodiments, the STING agonist is 2'-0- propargyl-cyclic- [A (2 ', 5') DA (3 ', 5') D] (2-O-propargyl-ML-CDA) or a compound represented in Figure 7 of PCT Publication No. WO 2014/189806.
[00617] [00617] Other exemplary STING agonists are disclosed, for example, in PCT Publications nº WO 2014/189805 and WO 2014/189806 and U.S. Publication No. 2015/0056225. Exemplary CSF-1 / 1R Binding Agents
[00618] [00618] In certain embodiments, the anti-CD73 antibody molecule described herein is administered in combination with a CSF-1 / 1IR binding agent. In some modalities, the combination is used to treat cancer, for example,
[00619] [00619] In some embodiments, the CSF-1 / 1R binding agent is an inhibitor of macrophage colony stimulating factor (M-CSF). M-CSF is also sometimes known as CSF-1.
[00620] [00620] In another embodiment, the CSF-1 / IR binding agent is a CSF-1R tyrosine kinase inhibitor, 4 - ((2 - ((((1R, 2R) -2-hydroxycyclohexyl) amino) benzo [d] thiazol-6-yl) oxy) -N-methylpicolinamide (Compound A15), or a compound disclosed in PCT Publication No. WO 2005/073224. In some modalities, cancer is chosen from brain cancer (eg glioblastoma multiforme (GBM)), pancreatic cancer or breast cancer (eg triple negative breast cancer (TNBC)).
[00621] [00621] In some embodiments, the CSF-1 / 1R binding agent (for example, a CSF-1R tyrosine kinase inhibitor), 4 - ((2- (((1R, 2R) -2-hydroxycyclo- hexyl) amino) benzo [dltiazol-6-yl) oxy) -N-methylpicolinamide (Compound A15), or a compound disclosed in PCT Publication No. WO 2005/073224, is administered in combination with a CD73 inhibitor (for example example, an anti-CD73 antibody molecule).
[00622] [00622] In certain embodiments, the CSF-1 / IR binding agent (for example, a CSF-1R tyrosine kinase inhibitor), 4 - ((2- (((1R, 2R) -2-hydroxycyclo- hexyl) amino) benzo [dltiazol-6-yl) Oxy) -N-methylpicolinamide (Compound A15), or a compound disclosed in PCT Publication No. WO 2005/073224, is administered in combination with the CD73 inhibitor (for example, the anti-CD73 antibody molecule) to treat a cancer, for example, a solid tumor (for example, an advanced solid tumor), for example, a brain cancer (for example, glioblastoma multiforme (GBM), for example, glioblas-
[00623] [00623] In some embodiments, the CSF-1 / 1R binding agent is an inhibitor of M-CSF, Compound A33 or a CSF-1 binding agent disclosed in PCT Publication No. WO 2004/045532 or Publication PCT No. WO 2005/068503 including RXK1 or 5H4 (for example, an antibody molecule or Fab fragment against M-CSF). In some modalities, cancer is chosen from endometrial cancer, skin cancer (eg, melanoma), pancreatic cancer or breast cancer (eg, triple negative breast cancer (TNBC)) .
[00624] [00624] In some embodiments, the CSF-1 / IR binding agent is a CSFIR or 4- (2 - ((1R, 2R) -2-hydroxycyclohexylamino) benzothiazole-6-yloxy) -N- methylpicolinamide. - 4- (2 - ((1R, 2R) -2-hydroxycyclohexylamino) benzothiazol-6-yloxy) -N-methylpicolinamide is disclosed as example 157 on page 117 of PCT Publication No. WO 2007/121484.
[00625] [00625] In some embodiments, the CSF-1 / IR binding agent is pexidartinib (CAS Registry Number 1029044-16-3). Pexidarithinib is also known as PLX3397 or 5 - ((5-chloro-1H-pyrrolo [2,3-b] pyridin-3-yl)] methyl) - N - ((6- (trifluoromethyl) pyridin-3- iNmethyl) pyridin-2-amine Pexidartinib is a small molecule tyrosine kinase receptor (RTK) inhibitor of KIT, CSFIR and FLT3 In some embodiments, the CSF-1 / IR binding agent, for example, pexidartinib, is used in combination with a CD73 inhibitor, for example, an anti-CD73 antibody molecule described herein.
[00626] [00626] In some embodiments, the CSF-1 / IR binding agent is emactuzumab. Emactuzumab is also known as
[00627] [00627] In some embodiments, the CSF-1 / IR binding agent is FPAOO8. FPAOO8 is a humanized mAb that inhibits CSFIR. In some embodiments, the CSF-1 / IR binding agent, for example, FPAOO8, is used in combination with a CD73 inhibitor, for example, an anti-CD73 antibody molecule described herein. Exemplary IDO / TDO inhibitors
[00628] [00628] In certain embodiments, the anti-CD73 antibody molecule described herein is administered in combination with an indolamine 2,3-dioxigenase (IDO) and / or 2,3-dioxigenase tryptophan (TDO) inhibitor. In some embodiments, the combination is used to treat a cancer, for example, a cancer described in this document, for example, a solid tumor (for example, melanoma, non-small cell lung cancer, colon cancer, cell cancer scaly head and neck, ovarian cancer, peritoneal cancer, fallopian tube cancer, breast cancer (for example, metastatic or HER2 negative breast cancer)), for example, hematological malignancy (for example, a lymphoma, for example, a non-Hodgkin's lymphoma or a Hodgkin's lymphoma (for example, a large diffuse B-cell lymphoma (DLBCL))).
[00629] [00629] In some embodiments, the IDO / TDO inhibitor is chosen from (4E) -4 - [(3-chloro-4-fluoroanilino) -nitrosomethylidene] - 1,2,5-0xadiazole-3- amine (also known as INCB24360), indo-ximod (1-methyl-D-tryptophan) or a-cyclohexyl-5SH-Imidazo [5,1-alisoin-dol-5-ethanol (also known as NLG919).
[00630] [00630] In some modalities, the IDO / TDO inhibitor is epacadostate (CAS Registry Number: 1204669-58-8). Epacadostate is also known as INCB24360 or INCB024360 (Incyte). Epacadostate is a powerful and selective indolamine 2,3-dioxigenase (IDO1) inhibitor with an IC50 of 10 nM, highly selective in relation to other related enzymes such as IDO 2 or tryptophan 2,3-dioxigenase (TDO).
[00631] [00631] In some modalities, the IDO / TDO inhibitor is indoxi- mode (New Link Genetics). Indoximod, the D-isomer of 1-methyltryptophan, is a small molecule indolamine 2,3-dioxigenase (IDO) pathway inhibitor that disrupts the mechanisms through which tumors escape immune-mediated destruction.
[00632] [00632] In some embodiments, the IDO / TDO inhibitor is NLG919 (New Link Genetics). NLG919 is a powerful pathway inhibitor of IDO (indolamine- (2,3) -dioxigenase) with Ki / EC50 of 7 nM / 75 NM in cell free assays.
[00633] [00633] In some embodiments, the IDO / TDO inhibitor is FO001287 (Flexus / BMS). FO01287 is a small molecule indolamine 2,3-dioxigenase 1 (IDO1) inhibitor. Exemplary TGF-B inhibitors
[00634] [00634] In certain embodiments, the anti-CD73 antibody molecule described herein is administered in combination with a beta transforming growth factor inhibitor (TGF-B). In some embodiments, the combination is used to treat a cancer, for example, a cancer described in this document, for example, a solid tumor (for example, a brain cancer (for example, a glioma), a melanoma, a kidney cancer (eg, renal cell carcinoma), malignant pleural mesothelioma (eg, recurrent malignant pleural mesothelioma) or breast cancer (eg, metastatic breast cancer)). In certain embodiments, the cancer is chosen from a colorectal cancer (for example, a microsatellite stable colorectal cancer (MSS CRC), a liver cancer (for example, a hepatocellular carcinoma), a lung cancer (for example , non-small cell lung cancer (HSCLC)), breast cancer (e.g. triple negative breast cancer (TNBC)), TGF-B expression cancer, pancreatic cancer, prostate cancer or kidney cancer (for example, kidney cell carcinoma).
[00635] [00635] TGF-B belongs to a large family of structurally related cytokines including, for example, bone morphogenetic proteins (BMPs), growth and differentiation factors, activins and inhibins. In some embodiments, the TGF-B inhibitors described in this document may bind and / or inhibit one or more isoforms of TGF-B (for example, one, two or all TGF-B1, TGF-B2 or TGF-B3 ).
[00636] [00636] In some modalities, the TGF-B inhibitor is fresolimumab (CAS Registry Number: 948564-73-6). Fresolimumab is also known as GC1008. Fresolimumab is a human monoclonal antibody that binds to and inhibits TGF-beta isoforms 1, 2 and 3.
[00637] [00637] The fresolimumab heavy chain has the amino acid sequence of: QVQLVASGAEVKKPGSSVKVSCKASGYTFSSNVIS WVRQAPGQGLEWMGGVIPIVDIANYAQRFKGRVTITADESTSTTYM ELSSLRSEDTAVYYCASTLGLVLDAMDYWGQGTLVTVSSASTKG- PSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGAL- TSGVHTFPAVLOSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNT- KVDKRVESKYGPPCPSCPAPEFLGGPSVFLFPPKPKDTLMIS- RTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEQFNS- TYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISKA-
[00638] [00638] The light chain has fresolimumabe ami- noácidos sequence: ETVLTAOSPGTLSLSPGERATLSCRASQSLGSSYLAWY QQAKPGQAPRLLIYGASSRAPGIPDRFSGSGSGTDFTLTISR- LEPEDFAVYYCQQYADSPITFGQAGTRLEIKRTVAAPSVFIFPPS- DEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTE- QDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNR- GEC (SEQ ID NO: 173).
[00639] [00639] —Fresolimumab is disclosed, for example, in WO 2006/086469, U.S. No. 8,383,780 and U.S. No. 8,591,901.
[00640] [00640] In some embodiments, the TGF-B inhibitor is XOMA 089. XOMA 089 is also known as XPA.42.089. XOMA 089 is a fully human monoclonal antibody that binds and neutralizes ligands 1 and 2 of TG] GF-beta.
[00641] [00641] The XOMA 089 heavy chain variable region has the amino acid sequence of: QVQLVOSGAEVKKPGSSVKVSC KASGGTFSSYAISWVRQAPGQGLEWMGGIIPIFGTANYAQK- FQGRVTITADESTSTAYMELSSLRSEDTAVYRQ (NO.
[00642] [00642] The XOMA 089 light chain variable region has the amino acid sequence of: SYELTQPPSVSVAPGQTARITCGAN- DIGSKSVHWYQQKAGQAPVLVVEDEDIIRPSGIPERISGSNSGNTATL- TISRVEAGDEADYGQTDR (2016).
[00643] [00643] In certain embodiments, the combination includes a CD73 inhibitor (for example, an anti-CD73 antibody molecule described herein) and a TGF-B inhibitor (for example, a TGF-B inhibitor described herein) document).
[00644] [00644] In one embodiment, the combination includes a TGF-B inhibitor, XOMA 089 or a compound disclosed in PCT Publication No. WO 2012/167143, and a CD73 inhibitor (for example, an anti-CD73 antibody described in this document).
[00645] [00645] In one embodiment, the TGF-B inhibitor, XOMA 089, or a compound disclosed in PCT Publication No. WO 2012/167143, is administered in combination with a CD73 inhibitor (for example, an anti- CD73) to treat pancreatic cancer, colorectal cancer (for example, microsatellite stable colorectal cancer (MSS-CRC)), lung cancer (for example, non-small cell lung cancer), cancer of breast (for example, triple negative breast cancer), liver cancer (for example, hepatocellular carcinoma), prostate cancer or kidney cancer (for example, clear cell renal cell carcinoma). Exemplary VEGFR inhibitors
[00646] [00646] In certain embodiments, the anti-CD73 antibody molecule described herein is administered in combination with a vascular endothelial growth factor receptor (VEGF) inhibitor (for example, an inhibitor of one or more among VEGFR (for example, example, VEGFR-1, VEGFR-2 or VEGFR-3) or VEGF). In some embodiments, the combination is used to treat a cancer, for example, a cancer described in this document, for example, a solid tumor (eg, a melanoma, a breast cancer, a colon cancer, esophageal cancer, gastrointestinal stromal tumor (GIST), kidney cancer (eg, renal cell cancer), liver cancer, non-small cell lung cancer (NSCLC), ovarian cancer, pancreatic cancer, prostate cancer or stomach cancer), for example, a malignancy
[00647] [00647] In some embodiments, the VEGFR inhibitor is vatalanib succinate (Compound A47) or a compound disclosed in EP 296122.
[00648] [00648] In some embodiment, the VEGFR inhibitor is an inhibitor of one or more of VEGFR-2, PDGFRbeta, KIT or Raf kinase C, 1-methyl-5 - ((2- (5- (trifluoromethyl) -1H- imidazo | -2-yl) pyridin-4-yl) oxy) -N- (4- (trifluoromethyl) phenyl) - 1 H-benzo [d] imidazol-2-amine (Compound A37) or a compound disclosed in the Publication of PCT No. WO 2007/030377.
[00649] [00649] - Other exemplary VEGFR pathway inhibitors that can be used in the combinations disclosed in this document include, for example, bevacizumab (AVASTINO), axitinib (INLYTAG); brivanib alaninate (BMS-582664, (S) - ((R) -1- (4- (4-Fluoro-2-methyl-1H-indol-5-yloxy) -S-methylpyrrole propanoate [2,1- fI [1,2, A4] triazin-6-yloxy) propan-2-i1) 2-amino); sorafenib (NEXAVARO); pazopanib (VOTRIENTO); sunitinib malate (SUTENT); cedira-nibe (AZD2171, CAS 288383-20-1); vargatef (BIBF1120, CAS 928326-83-4); Foretinib (GSK1363089); telatinib (BAY57-9352, CAS 332012-40-5); apatinib (YN968D1, CAS 811803-05-1); imatinib (GLEEVECO); ponatinib (AP24534, CAS 943319-70-8); tivozanib (AV951, CAS 475108-18-0); regorafenib (BAY73-4506, CAS 755037-03-7); vatalanib dihydrochloride (PTK787, CAS 212141-51-0); bribanib (BMS-540215, CAS 649735-46-6); vandetanib (CAPRELSAO or AZD6474); motesanib diphosphate (AMG706, CAS 857876-30-3, N- (2,3-dihydro-3,3-dimethyl-1H-indol-6-i1) -2 - [(4-pyridinylmethyl) amino] - 3-pyridinecarboxamide, described in PCT Publication No. WO 02/066470); lymphanib (ABT869, CAS 796967-16-3); cabozantinib (XL184, CAS 849217-68-1); lestaurtinib (CAS 111358-88-4); N- [5 - [[[5- (1,1-dimethylethyl) -2-oxazolyl] methyl | thio] -2-thiazolyl] -4-piperidinecarboxamide (BMS38703, CAS 345627-80-7); (3R, 4R) -4-amino-1 - ((4 - ((3-
[00650] Exemplary anti-VEGF antibodies that can be used in the combinations disclosed herein include, for example, a monoclonal antibody that binds to the same epitope as the A4.6.1 anti-VEGF monoclonal antibody produced by the ATCC hybridoma HB 10709; a recombinant humanized anti-VEGF monoclonal antibody generated according to Presta et al. (1997) Cancer Res. 57: 4593 to 4599. In one embodiment, the anti-VEGF antibody is Be- vacizumab (BV), also known as rhuMAb or AVASTINO VEGF. It comprises regions of mutated human IgG1 framework and complementary determinant regions of the anti-murine anti-hnvVEGF A.4.6.1 monoclonal antibody that blocks the binding of human VEGF to its receptors. Bevacizumab and other humanized anti-VEGF antibodies are further described in US Patent No. 6,884,879 issued February 26, 2005. Additional antibodies include antibodies from the G6 or B20 series (for example, G6-31, B20-4.1) , as described in PCT Publication No. WOZ2005 / 012359, PCT Publication No. WOZ2005 / 044853, the contents of these patent applications are expressly incorporated by reference in this document. For additional antibodies, see US Patent No. 7,060,269, 6,582,959, 6,703,020, 6,054,297, WOS98 / 45332, WO 96/30046, WOS94 / 10202, EP 0666868B1, Patent Application Publications No. US 2006/009360, 2005/0186208, 2003 / 0206899, 2003/0190317,
[00651] [00651] In certain embodiments, the anti-CD73 antibody molecule described herein is administered in combination with a c-MET inhibitor. In some embodiments, the combination is used to treat a cancer, for example, a cancer described in this document, for example, a solid tumor (for example, a non-small cell lung cancer, pancreatic cancer, cancer liver disease, thyroid cancer (for example, anaplastic thyroid carcinoma), brain tumor (for example, a glioblastoma), kidney cancer (for example, renal cell carcinoma) or head and neck cancer (for example, a squamous cell carcinoma of the head and neck)). In certain embodiments, the cancer is a liver cancer, for example, a hepatocellular carcinoma (HCC) (for example, an HCC that expresses c-MET).
[00652] [00652] In some embodiments, the c-MET inhibitor is Compound A17 or a compound described in U.S. Patent Nos. 7,767,675 and 8,420,645).
[00653] [00653] In some embodiments, the c-MET inhibitor is JNJ-
[00654] [00654] In some embodiments, the c-Met inhibitor is AMG 208. AMG 208 is a selective small molecule inhibitor of c-MET. AMG
[00655] [00655] In some embodiments, the c-Met inhibitor is AMG 337. AMG 337 is an oral bio-available c-Met inhibitor. AMG 337 binds selectively to c-MET, thereby disrupting the signal transduction paths of c-MET.
[00656] [00656] In some embodiments, the c-Met inhibitor is LY2801653. LY2801653 is a small molecule inhibitor, available orally from c-Met. LY2801653 selectively binds to c-MET, thereby inhibiting phosphorylation of c-MET and disrupting the signal transduction pathways of c-Met.
[00657] [00657] In some embodiments, the c-Met inhibitor is MSC2156119J. MSC2156119J is an oral bioavailable c-Met inhibitor. MSC2156119J selectively binds to c-MET, which inhibits phosphorylation of c-MET and disrupts c-Met-mediated signal transduction trajectories.
[00658] [00658] In some embodiments, the c-MET inhibitor is capmatine. Capmatinib is also known as INCB028060. Capminib is an oral bioavailable c-MET inhibitor. Capmatinib binds selectively to c-Met, thereby inhibiting phosphorylation of c-Met and disrupting the signal transduction pathways of c-Met.
[00659] [00659] In some embodiments, the c-MET inhibitor is crizotinib. Crizotinib is also known as PF-02341066. Crizotinib is an aminopyridine-based inhibitor available orally from anaplastic tyrosine kinase lymphoma kinase receptor (ALK) and the c-Met / hepatocyte growth factor receptor (HGFR). Crizotinib, in a competitive ATP manner, binds to and inhibits ALK kinase and ALK fusion proteins. In addition, crizotinib inhibits c-
[00660] [00660] In some embodiments, the c-MET inhibitor is golvatinib. Golvatinib is a c-MET and VEGFR-2 double kinase inhibitor orally bioavailable with potential antineoplastic activity. Golvatinib binds to and inhibits the activities of both c-MET and VEGFR-2, which can inhibit tumor cell growth and the survival of tumor cells that overexpress these receptor tyrosine kinases.
[00661] [00661] In some embodiments, the c-MET inhibitor is tivantinib. Tivantinib is also known as ARQ 197. Tivantinib is an orally bioavailable small molecule inhibitor of c-MET. Tivantinib binds to the c-MET protein and disrupts c-Met signal transduction pathways, which can induce cell death in tumor cells that overexpress the c-MET protein or that express c protein -Met constitutively activated. Exemplary IAP inhibitors
[00662] [00662] In certain embodiments, the anti-CD73 antibody molecule described herein is administered in combination with an Apoptosis Protein Inhibitor (IAP) inhibitor. In some embodiments, the combination is used to treat a cancer, for example, a cancer described in this document, for example, a solid tumor (for example, colorectal cancer (CRC), a lung cancer (for example, example, a non-small cell lung cancer (NSCLC)), a breast cancer (for example, a triple negative breast cancer (TNBC)), an ovarian cancer or a pancreatic cancer), for example, a haematological malignancy (for example, multiple myeloma).
[00663] [00663] In some embodiments, the IAP inhibitor is (S) -N - ((S) -1- cyclohexyl-2 - ((S) -2- (4- (4-fluorobenzoyl) thiazole-2- il) pyrrolidin-1-i1) -2-
[00664] [00664] In some embodiments, the combination described in this document includes an IAP inhibitor, (S) -N - ((S) -1-cyclohexyl-2- ((S) -2- (4- (4-fluorobenzoyl) thiazol-2-yl) pyrrolidin-1-11) -2-0x0ethyl) -2- (methylamino) propanamide (Compound A21), or a compound disclosed in US Patent No. 8,552,003 and an inhibitor of a molecule of immune checkpoint, for example, a CD73 inhibitor (for example, an anti-CD73 antibody molecule).
[00665] [00665] In certain embodiments, the anti-CD73 antibody molecule described herein is administered in combination with an Epidermal Growth Factor Receptor (EGFR) inhibitor. In some embodiments, the combination is used to treat a cancer, for example, a cancer described in this document, for example, a solid tumor (for example, a lung cancer (for example, a non-cell lung cancer) small), pancreatic cancer, breast cancer (for example, triple negative breast cancer (TNBC)) or colon cancer. In certain modalities, cancer is chosen from colorectal cancer (eg, microsatellite stable colorectal cancer (MSS CRC)), lung cancer (eg, non-small cell lung cancer) or a breast cancer (for example, a triple negative lung cancer (TNBC)).
[00666] [00666] In some embodiments, the EGFR inhibitor is (R, E) -N- (7-chloro-1- (1- (4- (dimethylamino) but-2-enoyl) azepan-3-yl) -1H - benzol [d] imidazol-2-yl) -2-methylisonicotinamide (Compound A40) or a compound disclosed in PCT Publication No. WO 2013/184757.
[00667] [00667] In some embodiments, the combination described in this document includes an EGFR inhibitor, (R, E) -N- (7-chloro-1- (1- (4-
[006688] [006688] In some embodiments, the EGFR inhibitor, (R, E) -N- (7-chloro-1- (1- (4- (dimethylamino) but-2-enoyl) azepan-3-yl) -1H -benzo [d] imidezol-2-i1) -2-methylisonicotinamide (Compound A40) or a compound disclosed in PCT Publication No. WO 2013/184757, is administered in combination with a CD73 inhibitor (for example, an anti-CD73 antibody molecule) to treat colorectal cancer (CRC) (for example, an MSS-CRC), lung cancer (for example, non-small cell lung cancer (NSCLC)) or cancer of the breast (for example, triple negative breast cancer (TNBC)).
[00669] [00669] In some embodiments, the EGFR inhibitor is chosen from one or more among erlotinib, gefitinib, cetuximab, pani- tumumab, - necitumumab, —PF-00299804, nimotuzumab or RO5083945.
[00670] [00670] In certain embodiments, the anti-CD73 antibody molecule described herein is administered in combination with a rapamycin target inhibitor (MTOR). In some embodiments, the combination is used to treat a cancer, for example, a cancer described in this document, for example, a solid tumor (for example, a prostate cancer, a breast cancer, a brain cancer, a bladder cancer, pancreatic cancer, kidney cancer or liver cancer, lung cancer (eg, small cell lung cancer or non-small cell lung cancer), respiratory / thoracic cancer , sarcoma, bone cancer, non-small cell lung cancer, endocrine cancer, astrocytoma, cervical cancer, neurological cancer, gastric cancer or melanoma), for example, hematological malignancy ( for example, leukemia (for example, lymphocytic leukemia), for example, lymphoma or, for example, multiple myeloma). In certain modalities, cancer is chosen from a colorectal cancer (for example, a microsatellite stable colorectal cancer (MSS CRC)), a lung cancer (for example, a non-small cell lung cancer) or breast cancer (for example, triple negative lung cancer (TNBC)).
[00671] [00671] In some embodiments, the mMTOR inhibitor is 8- (6-Methoxy-pyridin-3-yl) -3-methyl-1- (4-piperazin-1-yl-3-trifluoromethyl-phenyl) -1, 3-dihydro-imidazo [4,5-c] quinolin-2-o0na (Compound A41).
[00672] [00672] In some embodiments, the MTOR inhibitor is everolimus (also known as AFINITORG; Compound A36) or a compound disclosed in PCT Publication No. WO 2014/085318.
[00673] [00673] In some embodiments, the combination described in this document includes the mMTOR inhibitor, everolimus (Compound A36) or a compound disclosed in PCT Publication No. WO 2014/085318 and an inhibitor of an immune checkpoint molecule , for example, a CD73 inhibitor (for example, an anti-CD73 antibody molecule).
[00674] [00674] In some embodiments, the MTOR inhibitor, Everolimus (Compound A36) or a compound disclosed in PCT Publication No. WO 2014/085318, is administered in combination with the CD73 inhibitor (for example, the anti- CD73) to treat colorectal cancer, lung cancer (for example, non-small cell lung cancer (NSCLC)) or breast cancer (for example, triple negative breast cancer (NTBC)).
[00675] [00675] In some modalities, the MTOR inhibitor is chosen from one or more among rapamycin, temsirolimus (TORISELO),
[00676] [00676] Other exemplary MTOR inhibitors include, but are not limited to, temsirolimus; (1R, 2R, 48) -4 - [(2R) -2 [(1R, 98,128, / 15R, 16E, 18R, 19R, 21R, 23S, 24E, 26E, 282Z2,30S, 32S, 35R) ridaforolimus dimethylphosfinate -1,18-dihydroxy-19,30-dimethoxy-15,17,21,23,29,35-hexamethyl-2,3,10,14,20-pentaoxo-11,36-dioxa-4-azathricycle [30.3.1.04.9] hexatriaconta-16,24,26,28-tetraen-12-yl] propyl] -2-methoxycyclohexyl, also known as AP23573 and MK8669; everolim (RADOO01); rapamycin (AY22989); simapimod; (5- (2,4-bis [(3S) -3-methylmorpholin-4-yl] pyrido [2,3-apyrimidin-7-yl) -2-methoxyphenyl) methane! (AZD8055); 2-mino-8- [trans-4- (2-hydroxyethoxy) cyclohexyl] -6- (6-methoxy-3-pyridinyl) -4-methyl-pyrido [2,3-apirimidin-7 (8H) - ona (PFO4691502); and No.
[00677] [00677] In certain embodiments, the anti-CD73 antibody molecule described herein is administered in combination with a phosphatidylinositol-4,5-bisphosphate 3-kinase inhibitor (PI3SK), for example, phosphatidylinositol-4,5-bisphosphate Gamma and / or delta 3-kinase (PI3K-y, 5). In some modalities, the combination is used to treat a cancer, for example, a cancer described in this document, for example, a solid tumor (for example, a prostate cancer, a breast cancer, a brain cancer , bladder cancer, pancreatic cancer, kidney cancer, solid tumor, liver cancer, non-small cell lung cancer, endocrine cancer, ovarian cancer, melanoma, system cancer female reproductive system, digestive / gastrointestinal cancer, glioblastoma multiforme, head and neck cancer or colon cancer, eg hematological malignancy (eg leukemia (eg leukemia lymphocytic, for example, chronic lymphocytic leukemia (CLL) (for example, recurrent CLL)), for example, a lymphoma (for example, non-Hodgkin's lymphoma (for example, recurrent follicular B cell non-Hodgkin's lymphoma) or small recurrent lymphocytic lymphoma (SLL)) or, for example, multiple myeloma) .
[00678] [00678] In some embodiments, the PI3K inhibitor is an inhibitor of PI3K delta and gamma isoforms. Exemplary PI3SK inhibitors that can be used in combination are described, for example, in WO 2010/036380, WO 2010/006086, WO 09/114870, WO 05/113556, GSK 2126458, GDC-0980, GDC-0941, Sanofi XL147, XL756, XL147, PF-46915032, BKM 120, CAL-101, CAL 263, SF1126, PX-886 and a double PI3K inhibitor.
[00679] [00679] In some embodiments, the PI3K-y inhibitor, ô is idelalisable (CAS Registry Number: 870281-82-6). Idelalisib is also known as ZYDELIGGO, GS-1101, CAL-101 or 5-Fluoro-3-phenyl-2 - [(18) -1- (7H-purin-6-ylamino) propyl] -4 (3H) - quinazolinone. Idelalisib blocks P1105, the delta isoform of PISK. Idelalisib is disclosed, for example, in Wu et al. Journal of Hematology & Oncology (2013) 6:
[00680] [00680] In some embodiments, the PI3K-y, ô inhibitor is 8- (6- Methoxy-pyridin-3-yl) -3-methyl-1- (4-piperazin-1-yl-3-trifluoromethyl-phenyl ) -1,3-dihydro-imidazo [4,5-c] quinolin-2-0na (Compound A41).
[00681] [00681] In some embodiments, the PI3K-y inhibitor, ô is buparlisible (Compound A6) or a compound disclosed in PCT Publication No. WO 2007/084786.
[00682] [00682] Other exemplary PI3K-y inhibitors that can be used in the combination include, for example, pictilisib (GDC-0941), LY294002, pilaralisib (XL147), PI-3065, PI-103, VS-5584 (SB2343 ), CZC24832, duvelisib (IPI-145, INK1I197), TG100-115, CAY10505, GSK1059615, PF-O04691502, AS-605240, voxtalisib (SAR245409, XL765), IC-87114, omipalisibe (GSK2126458, TGK12126458, (PF-05212384, PKI-587), PKI-402, XL147 analogue, PIK-90, PIK-293, PIK-294, 3-Methyladenine (3-MA), AS-252424, AS-604850 or apitolisib (GDC -0980, RG7422).
[00683] [00683] In some embodiments, the PI3K inhibitor is Compound A8 or a compound described in PCT Publication No. WO 2010/029082.
[00684] [00684] In some embodiments, the PI3K inhibitor is a pan-PI3K inhibitor, (48S, 5R) -3- (2: -amino-2-morpholino-4 '- (trifluoromethyl) - [4,5'- bipyrimidin] -6-yl) -4- (hydroxymethyl) -5-methyloxazolidin-2-one (Compound A1I3) or a compound disclosed in PCT Publication No. WO 2013/124826.
[00685] [00685] Exemplary PI3K-y, -5 inhibitors include, but are not limited to, duvelisib and idelalisib. Idelalisib (also called GS-1101 or CAL-101; Gilead) is a small molecule that blocks the PI3K delta isoform. The structure of idelalisib (5-Fluoro-3-phenyl-2- [(1S) -1- (7H-purin-6-ylamino) propyl] -4 (3H) -quinazolinone) is shown below.
[00686] [00686] - Duvelisibe (also known as IPI-145; Infinity Pharmaceuticals and Abbvie) is a small molecule that blocks PI3K-ô, y. The structure of duvelisib (8-Chloro-2-phenyl-3 - [(1S) -1- (9H-purin-6-ylamino) ethyl] -1 (2H) -isoquinolinone) is shown below. c x O CO
[00687] [00687] In one embodiment, the inhibitor is a double phosphatidylinositol 3-kinase (PISK) and the MTOR inhibitor selected from 2- Amino-8- [trans-4- (2-hydroxyethoxy) cyclohexyl] -6 - (6-methoxy-3-pyridinyl) -4-methyl-pyrido [2,3-a | pyrimidin-7 (8H) -one (PF-04691502); N- [4 - [[4- (Dimethylamino) -1-piperidinyl] | carbonyl] phenyl] -N '- [4- (4,6-di-4-morpholinyl-1,3,5-tria- zin-2-yl) phenylJurea (PF-05212384, PKI-587); apitolisib (GDC-0980, RG7422); 2,4-Difluoro-N- (2- (methyloxy) -5- [4- (4-pyridazinyl) -6-quinolinyl] -3-pyridinyl) benzenesulfonamide (GSK2126458); 8- (6-methoxypyridin-
[00688] [00688] In certain embodiments, the anti-CD73 antibody molecule described herein is administered in combination with a Janus kinase inhibitor (JAK). In some embodiments, the combination is used to treat a cancer, for example, a cancer described in this document, for example, a solid tumor (for example, a colon cancer, a prostate cancer, a lung cancer, a cancer breast cancer or pancreatic cancer), for example, hematological malignancy (for example, leukemia (for example, myeloid leukemia or lymphocytic leukemia), for example, lymphoma (for example, non-Hodgkin's lymphoma) or myeloma multiple).
[00689] [00689] In some embodiments, the JAK inhibitor is 2-fluoro-N-methyl-4- (7- (quinolin-6-ylmethyl) imidazo [1,2-b] [1,2,4] triazin-2 -il) Dbenzamide (Compound A17) or a dihydrochloric salt thereof, or a compound disclosed in PCT Publication No. WO 2007/070514.
[00690] [00690] In some embodiment, the JAK inhibitor is ruxo-litinib phosphate (also known as JAKAFI; Compound A18) or a compound disclosed in PCT Publication No. WO 2007/070514.
[00691] [00691] Anti-CD73 antibody molecules can also be combined with a cell therapy, for example, a chimeric antigen receptor (CAR) therapy, a T cell therapy, a natural killer cell therapy ( NK) or dendritic cell therapy
[00692] [00692] The anti-CD73 antibody molecules described in this document can be administered in combination with a second therapeutic agent, for example, a cell comprising a chimeric antigen (CAR) receptor. The CAR can comprise i) an extracellular antigen-binding domain, ii) a trans-membrane domain and iii) an intracellular signaling domain (which can comprise one or both of a primary signaling domain and a co-stimulating domain). The CAR may additionally comprise a leader sequence and / or a successor sequence. In specific modes, the CAR construct comprises a scFv domain, where scFv can be preceded by an optional leader sequence, and followed by an optional successor sequence, a transmembrane region and an intracellular signaling domain, for example. example, where the domains are contiguous to and in the same reading frame to form a single fusion protein.
[00693] [00693] In some embodiments, the CAR molecule comprises a CD19 CAR molecule described herein, for example, a CD19 CAR molecule described in U.S. No. 2015/0283178, for example, CTLO19. In the modalities, the CD19 CAR comprises an amino acid, or has a sequence of nucleotides shown in document No. US 2015/0283178, incorporated in this document for reference in its entirety, or a sequence substantially identical to the same (for example , a sequence that has at least about 85%, 90% or 95% sequence identity with it).
[00694] [00694] In one embodiment, the CAR T cell that binds to CD19 has the designation USAN TISAGENLECLEUCEL-T. CTLO19 is produced by an insertion-mediated T-cell gene modification
[00695] [00695] In one embodiment, CAR CD19 comprises an amino acid sequence provided as SEQ ID NO: 12 in PCT publication WO 2012/079000. In the modality, the sequence of amino acids is:
[006968] [006968] -MALPVTALLLPLALLLHAARPdiqmtattsslsas | gdrvtis- crasqdiskylInwyqgqgkpdgtvklliyhtsrlhsgvpsrfsasasgtdysltisnlegediat- yfcaagntipytfgggtkleitagggsagggsgagggsevklgesgpglvap- sqaslsvtctvsgvs | pdygvswirgpprkglewlgviwasettyynsalksrItiik- dnsksqvflkmnslqatddtaiyycakhyyyggsyamdywgagtsvtvsstttpaprpptpap- tiasqplIslrpeacrpaaggavhtraldfacdiyiwaplagtcgvllls | vitlyckrgrk- Kllyifkapfmrpvattgeedgescripeeeeggcelrvkfsrsadapa- yKaganalynelnIgrreeydvldkrrgrdpemggkprrknpgeglynel- qgkdkmaeayseigmkgerrrgkghdglygglstatkdtydalhmgalppr (SEQ ID NO: 132) or a sequence substantially identical thereto (e.g., a sequence which has at least about 85%, 90% or 95% sequence identity with the same), with or without the signal peptide sequence indicated in capital letters.
[00697] [00697] In one embodiment, the amino acid sequence is:
[00698] [00698] - diqgmtattssisaslgdrvtiscrasqdiskylnwygagkpdgtvklli- yhtsrlhsgvpsrfsgsgsgtdysltisnlegediatyfcaggntlpytfgggtkleit- ggggsgggasggggsevklgesgpalvapsaslisvtctvsgvs | pdygvswirap- prkglewlgviwgsettyynsalksritikdnsksqvflkmnslqgtdd- taiyycakhyyyggsyamdywgagtsvtvsstttpaprpptpapti- asqplslrpeacrpaaggavhtraldfacdiyiwaplagtegvllislvitlyckrgrkklly- ifkgpfmrpvattgeedgcscríipeeeeggcelrvkfsrsadapaykqgggnaqlynelnigr-
[00699] [00699] The antigen binding domain can be any domain that binds to the antigen including, but not limited to, a monoclonal antibody, a polyclonal antibody, a recombinant antibody, a human antibody, a humanized antibody and a fragment functionality, including, but not limited to, a single domain antibody such as a heavy chain variable domain (VH), a light chain variable domain (VL) and a variable domain (VHH) of a nanocorp derived from camelid, and an alternative framework known in the art to function as the antigen binding domain, such as a recombinant fibronectin domain, a T cell receptor (TOR) or a fragment thereof, for example, single-chain TOR and the like. In some cases, it is beneficial that the antigen-binding domain is derived from the same species in which the CAR will ultimately be used. For example, for use in humans, it may be beneficial that the CAR antigen binding domain comprises human or humanized residues for the antigen binding domain of an antibody or antibody fragment.
[00700] [00700] In some embodiments, the CAR antigen binding domain is a scFv antibody fragment that is humanized compared to the murine scFv sequence from which it is derived.
[00701] [00701] In some embodiments, the antigen-binding domain binds to a tumor antigen described in this document. In the modalities, the tumor antigen is chosen from: CD19;
[00702] [00702] In one embodiment, the CAR molecule comprises a BCMA CAR molecule, for example, a BCMA CAR described in U.S. 2016/0046724 or WO 2016/014565, incorporated herein by reference. In the embodiments, the BCMA CAR comprises an amino acid, or has a nucleus sequence of a CAR molecule, or an antigen binding domain according to US 2016/0046724, or Table 1 or 16, SEQ ID NO: 271 or SEQ ID NO: 273 of WO 2016/014565, incorporated herein by reference, or a sequence substantially identical to any of the aforementioned sequences (for example, which is at least about 85%, 90% or 95% sequence identity with any of the above BCMA CAR sequences). The amino acid and nucleotide sequences encoding the BCMA CAR molecules and antigen binding domains (for example, including one, two, three VH CDRs; and one, two, three VL CDRs according to Kabat or Chothia) , are specified in WO 2016/014565.
[00703] [00703] Regarding the traunsmembrane domain, in several modalities, a CAR can be designed to understand a transmembrane domain that is linked to the extracellular domain of the CAR.
[00704] [00704] The transmembrane domain can be derived from a natural source or from a recombinant source. When the source is natural, the domain can be derived from any membrane-bound or transmembrane protein. In one aspect, the transmembrane domain is able to signal the intracellular domain (s) whenever the CAR has linked to a target. A transmembrane domain can include at least the transmembrane region (or regions) of, for example, the alpha, beta or zeta chain of the T cell receptor, CD28, CD27, CD3 epsilon, CD45, CD4, CD5, CD8 , CD9, CD16, CD22, CD33, CD37, CD6A4, CD80, CD86, CD134, CD137, CD154. In some embodiments, the transmembrane domain may include at least the transmembrane region (or regions), for example, from KIRDS2, OX40, CD2, CD27, LFA-1
[00705] [00705] In some cases, the transmembrane domain can be attached to the extracellular CAR region, for example, the CAR antigen binding domain, by means of a joint, for example, a joint of a human protein. For example, in one embodiment, the hinge can be a human Ig (immunoglobulin) hinge (for example, an IgG4 hinge, an IgD hinge), a GS linker (for example, a GS linker described here), a KIR2QDS2 hinge or a CD8a hinge. Domain of intracellular signaling of a chimeric antigen receptor (CAR)
[00706] [00706] The cytoplasmic domain or region of the CAR includes an intracellular signaling domain. An intracellular signaling domain is generally responsible for the activation of at least one of the normal effector functions of the immune cell where the CAR was introduced.
[00707] [00707] Examples of intracellular signaling domains for use in the CAR include the T cell receptor (TOR) cytoplasmic sequences and co-receptors that act in concert to initiate signal transduction after antigen receptor engagement, as well as - any derivative or variant of these sequences and any recombinant sequence that has the same functional capacity.
[00708] [00708] A primary signaling domain regulates the primary activation of the TCR complex in a stimulatory or inhibitory manner. Primary intracellular signaling domains that act in a stimulating manner may contain signaling motifs that are known as immunoreceptor tyrosine-based activation motifs or ITAMs.
[00709] [00709] Examples of ITAM containing primary intracellular signaling domains include those of CD3 zeta, FCR common gamma (FCER1G), Fc gamma Rlla, FcR beta (Fc Epsilon R1b), CD3 gamma, CD3 delta, CD3 epsilon, CD79a , CD79b, DAP10 and DAP12. In one embodiment, a CAR comprises an intracellular signaling domain, for example, a primary CD3-zeta signaling domain.
[00710] [00710] The intracellular signaling domain of the CAR can comprise the CD3-zeta signaling domain alone or can be combined with any (any) other intracellular signaling domain (s) useful (useful) in the context of a CAR of the invention. For example, the intracellular signaling domain of the CAR may comprise a portion of the CD3 zeta chain and a co-stimulatory signaling domain. The co-stimulatory signaling domain refers to a portion of the CAR comprising the intracellular domain of a co-stimulating molecule. A co-stimulatory molecule is a cell surface molecule that is not a receptor for antigens or their ligands that is necessary for an efficient lymphocyte response to an antigen. Examples of such molecules include CD27, CD28, 4-1BB (CD137), OX40, CD30, CD40, PD-1, ICOS, antigen associated with lymphocyte-1 (LFA-1), CD2, CD7, LIGHT function , NKG2C, B7-H3 and a linker that binds to CD83, and the like. For example, co-stimulation of CD27 has been shown to enhance the expansion, effector function and survival of human CART cells in vitro and increase human T cell persistence and in vivo antitumor activity
[00711] [00711] Immune effector cells such as T cells can be activated and expanded generally using methods as described, for example, in U.S. Patent No. 6,352,694; 6,534,055; 6,905,680; 6,692,964; 5,858,358; 6,887,466; 6,905,681; 7,144,575; 7,067,318; 7,172,869; 7,232,566; 7,175,843; 5,883,223; 6,905,874; 6,797,514; 6,867,041; and Patent Application Publication No. U.S. 2006/0121005, incorporated herein by reference.
[00712] [00712] Examples of immune effector cells include T cells, for example, alpha / beta T cells and gamma / delta T cells, B cells, natural killer cells (NK), natural killer T cells (NKT), mast cells and phagocytes derived from myeloid lineage.
[00713] [00713] Methods for producing cells expressing CAR are described, for example, in U.S. 2016/0185861, incorporated herein by reference. Exemplary Cancer Vaccines
[00714] [00714] The anti-CD73 antibody molecules can be combined
[00715] [00715] The CD73 block can be used in conjunction with a collection of recombinant proteins and / or peptides expressed in a tumor to generate an immune response to these proteins.
[00716] [00716] Other tumor vaccines may include virus proteins implicated in human cancers such as Human Papilloma Virus (HPV), Hepatitis Virus (HBV and HCV), Kaposi's Herpes Sarcoma Virus (KHSV ) and Epstein-Barr virus (EBV). Another form of tumor-specific antigen that can be used in conjunction with CD73 blockade is purified heat shock proteins (HSP) isolated from the tumor tissue itself. These heat shock proteins contain protein fragments from tumor cells and these HSPs are highly effective in delivering antigen presenting cells to elicit tumor immunity (Suot, R & Srivastava, P (1995) Science 269: 1585 to 1588; Tamura, Y. et al. (1997) Science 278: 117 to 120).
[00717] [00717] Dendritic cells (DC) are potent cells that present antigen that can be used to initiate specific antigen responses. DC's can be produced ex vivo and loaded with various protein and peptide antigens, as well as tumor cell extracts (Nestle, F. et a /. (1998) Nature Medicine 4: 328 to 332). DCs can also be transduced by genetic means to express these tumor antigens as well. DCs have also been fused directly to tumor cells for immunization purposes (Kugler, A. et al. (2000) Nature Medicine 6: 332-336). As a vaccination method, DC immunization can be effectively combined with blocking CD73 to activate more potent antitumor responses. Exemplary Oncolytic Viruses
[00718] [00718] The anti-CD73 antibody molecules can be administered in combination with the oncolytic virus. In modalities, oncolytic viruses are able to selectively replicate in, and trigger the death of, or retard the growth of, a cancer cell. In some cases, oncolytic viruses have no effect or have minimal effect on cancer cells. In some embodiments, the combination is used to treat cancer, for example, a cancer described in this document. In certain modalities, cancer is brain cancer, for example, a glioblastoma (GBM). An oncolytic virus includes, but is not limited to, an oncolytic adenovirus, Oncolytic Herpes Simplex Virus, oncolytic retrovirus, oncolytic parvovirus, oncolytic vaccine virus, oncolytic Sindbis virus, oncolytic influenza virus or oncolytic RNA virus (for example , oncolytic reovirus, Newcastle oncolytic disease virus (NDV), oncolytic measles virus or oncolytic vesicular somatitis virus (VSV)).
[00719] [00719] Exemplary oncolytic viruses include, but are not limited to, the following:
[00720] [00720] - Group B Oncolytic Adenovirus (ColoAd1) (PsiOxus Therapeutics Ltd.) (see, for example, Clinical Test Identifier: NCTO02053220);
[00721] [00721] - “ONCOS-102 (formerly called CGTG-102), which is an adenovirus that comprises granulocyte-macrophage colony stimulating factor (GM-CSF) (Oncos Therapeutics) (see, for example, Clinical Test: NCTO1598129);
[00722] [00722] VCN-01, which is a genetically modified human oncolytic adenovirus encoding human PH20 hyaluronidase (VCN Biosciences, S.L.) (see, for example, Clinical Test Identifiers: NCTO2045602 and NCT02045589);
[00723] [00723] —Conditionally Replicative Adenovirus ICOVIR-5, which is a virus derived from the wild type 5 human adenovirus (Had5) serotype that has been modified to selectively replicate in cancer cells with an unregulated retinoblastoma / E2F pathway (Institut Català d ' Oncology) (see, for example, Clinical Test Identifier: NCTO1864759);
[00724] [00724] Celyvir, which comprises bone marrow-derived autologous mesenchymal stem cells (MSCs) infected with ICOVIRS5, an oncolytic adenovirus (Hospital Infantil Universitario Niho Jesús, Madrid, Spain / Ramon Alemany) (see, for example, Identifi- Clinical Test indicator: NCTO1844661);
[00725] [00725] CGO0070, which is a conditionally replicated oncolytic serotype 5 (Ad5) adenovirus in which the human E2F-1 promoter drives the expression of the essential viral genes of Ela, thus restricting viral replication and cytotoxicity defective tumor cells in the Rb path (Cold Genesys, Inc.) (see, for example, Clinical Test Identifier: NCTO2143804); or
[00726] [00726] DNX-2401 (formerly called Delta-24-RGD), which is an adenovirus that has been genetically modified to selectively replicate in cells deficient in the retinoblastoma (Rb) pathway and to infect cells that express certain lipid integrins RGD research more effectively (Clinica Universidad de Navarra, Universidad de Navarra / DNAtrix, Inc.) (see, for example,
[00727] [00727] In some embodiments, an oncolytic virus described in this document is administered by injection, for example, subcutaneous, intra-arterial, intravenous, intramuscular, intrathecal or intraperitoneal injection. In modalities, an oncolytic virus described here is administered via intratumor, transdermal, transmucosal, oral, intranasal or pulmonary. Additional Exemplary Cancer Therapies
[00728] [00728] Exemplary combinations of anti-CD73 antibody molecules (alone or in combination with other stimulating agents) and cancer care standard, include at least the following. In certain embodiments, the anti-CD73 antibody molecule, for example, the anti-CD73 antibody molecule described herein, is used in combination with a standard chemotherapeutic cancer care agent including, but not limited to, trozol (Arimidexº), bicalutamide (Casodexº), bleomycin sulfate (Blenoxaneº), busulfan (Myleranº), busulfan injection (Busulfexº), capecitabine (Xeloda ), N4-pentoxicarbonyl-5-deoxy-5-fluorocytidine, carboplatin-paraplatin ), carmustine (BICNUº), chlorambucil (Leuke-ranº), cisplatin (Platinolº), cladribine (Leustatinº), cyclophosphamide (Cytoxanº or Neosarº), cytarabine, cytosine arabinoside (Cytosar-Uº), liposome injection dacarbazine (DTIC- Domeº), dactinomycin (Actinomycin D, Cosmegan), daunorubicin hydrochloride (Cerubidineº), dau- norubicin citrate liposome injection (DaunoXomeº), dexamethasone, docetaxel (Taxotereº), hydrochloride dexamide, hydrochloride dex Rubexº), etoposí deo (Vepeidº), fludarabine phosphate (Fludara ), 5-fluorouracil (Adrucilº, Efu- dexº), flutamide (Eulexinº), tezacitibine, Gemcitabine (difluorodoxycytidine), hydroxyurea (Hydreaº), Idarrubicin ifosfamide (IFEX9), irinotecan (Camptosarº), L-asparaginase (ELSPARSº), leucovorin calcium, melphalan (Alkeranº), 6-mercaptopurine (Purinetolº), methotrexate (Folexº), mitoxantrona (Novantroneº), milotar, phoenix (Yttrium90 / MX-DTPA), pentostatin, polyifeprosan 20 with carmustine implant (Gliadelº), tamoxifen citrate (Nolvadexº), teniposide (Vumonº), 6-thioguanine, thiotepa, tirapazamine (Tirazoneº), topecan hydrochloride (topecan hydrochloride) Hycamptinº), vinblastine (Velban banº), vincristine (Oncovinº), vinorelbine (Navelbineº), Ibrutinib, idelisysib and vedotinade brentuximab.
[00729] [00729] Exemplifying alkylating agents include, without limitation, nitrogen mustards, ethylenimine derivatives, alkylsulfonates, nitro-moles and triazenes: uracil mustard (Aminouracil MustardO, ChlorethaminacilO, DemethyldopanO, DesmethyldopantO, Uremanil , Urailcilmostaza € O, UramustinO, UramustineO), chlormetino (MustargenO), cyclophosphamide (CytoxanO, NeosarO, ClafenO, EndoxanO, ProcytoxO, RevimmuneTM), ifosfamida (MitoxanaO), melphalan (Alkeranil), (AmedelO, VercyteO), triethylene-melamine (Hemel &, HexalenO, HexastatO), triethylene thiophosphoramine, Temozolomide (TemodarGO), thiotepa (Thioplex6 & O), busulfan (Busilexex, MyleranO), carmine (estrogen) tozocine (ZanosarO) and Dacarbazine (DTIC-DomeO). Additional exemplary alkylating agents include, without limitation, Oxaliplatin (EloxatinO); Temozolomide (Temodarê & e TemodalO); Dactinomycin (also known as actinomycin-D, CosmegenO); Melfalan (also known as L-PAM, L-sarcolysin, and phenylaniline mustard, AlkeranO); Altretamine (also known as hexamethylmethylamine (HMM), HexalenO); Carmustine (BICNUGO); Bendamustina (Trepa € O); Busulfan (Busulfexô and Myleran &O); Carboplatin (Paraplatin &O); Lomustine (also known as CCNU, CeeNUG); Cisplatin (also known as CDDP, Platinol & and PlatinolO-AQ); Clo-
[00730] [00730] Exemplary anthracyclines include, for example, doxorubicin (AdriamycinO & O and RubexO); bleomycin (lenoxaneO); daunorubicin (dauorubicin hydrochloride, daunomycin and rubydomicin hydrochloride, Cerubidine &); liposomal daunorubicin (daunorubicin citrate liposome, DaunoXome6); mitoxantrone (DHAD, NovantroneO); epirubicin (Ellence "Y); idarubicin (Idamycin € O, Idamycin PFSO); mitomycin C (MutamycinO); geldanamycin; herbimycin; ravidomedicine; and deacetylravidomycin. Exemplary vinca alkaloids that can be used in combination with the molecules anti-CD73 antibodies, include, but are not limited to, vinorrelbine tartrate (NavelbineO), Vincristine (OncovinO) and Vindesine (EldisineO)); vinblastine (also known as vinblastine sulfate, vincaleuco-blastine and VLB , Alkaban-AQG and VelbanO), and vinorelbine (NavelbineO).
[00731] [00731] Exemplary proteasome inhibitors that can be used in combination with anti-CD73 antibody molecules include, but are not limited to, bortezomib (VelcadeG &); carfilzomib (PX-171-007, (S) -4-Methyl-N - ((S) -1 - (((S) -4-methyl-1 - ((R) -2-methyloxyran-2-yl) -1-oxopentan-2-yl) amino) -1-0x0-3-phenylpropan-2-yl) -2 - ((S) -2- (2-morpholinoacetamido) -4-phenylbutanamido) -pentanamide); marizomib (NPI-0052); ixazomib citrate (MLN-9708); delanzomibe (CEP-
[00732] [00732] In some embodiments, the anti-CD73 antibody molecule, for example, the anti-CD73 antibody molecule described herein, is used in combination with a tyrosine kinase inhibitor (for example, a tyrosine inhibitor recipient kinase (RTK)). The exemplary tyrosine kinase inhibitor includes, but is not limited to, an epidermal growth factor (EGF) pathway inhibitor (eg, an epidermal growth factor receptor (EGFR) inhibitor), a growth factor pathway inhibitor vascular endothelial (VEGF) inhibitor (for example, a vascular endothelial growth factor receptor (VEGFR) inhibitor (for example, a VEGFR-1 inhibitor, a VEGFR-2 inhibitor, a VEGFR-3 inhibitor)), a platelet-derived growth factor (PDGF) pathway inhibitor (for example, a platelet-derived growth factor receptor (PDGFR) inhibitor (for example, a PDGFR-B inhibitor)), an inhibitor of RAF-1, a KIT inhibitor and a RET inhibitor. In some modalities, the anticancer agent used in combination with the porcupine-type inhibitor is selected from the group consisting of: axitinib (AG013736), bosutinib (SKI-606), cedi ranib (RECENTIN'Y , AZD2171), dasatinib (SPRYCELO, BMS- 354825), erlotinib (TARCEVAG), gefitinib (IRESSAO), imatinib (Gleevec &, CGP57148B, STI-571), lapatinibe (TYKERBO, TYVERBO), lestaurtinibe (CEP) HKI-272), nilotinib (TASIGNAGO), semaxanib (semaxinib, SUS416), sunitinib (SUTENT, SU11248), toceranib (PALLADIAO), vandetanib (ZACTIMAG, ZD6474), vatala-nibe (PTK787, PTK / ZKuma, PTK / ZKuma, PTK / ZKuma ), bevacizumab (AVASTINO), rituximab (RITUXANO), cetuximab (ERBITUXO), pani- tumumab (VECTIBIXO), ranibizumab (Lucentis &), nilotinib (TA-
[00733] [00733] Radiation therapy can be administered through one of several methods, or a combination of methods, including, without limitation, external beam therapy, internal radiation therapy, implant radiation, stereotactic radiosurgery, systemic radiation therapy, radiotherapy and permanent or temporary interstitial brachytherapy. The term "brachytherapy" refers to radiation therapy delivered by a radioactive material confined spatially to the body in a tumor or close to the same or another site of proliferative tissue disease. The term is intended, without limitation, to include exposure to radioactive isotopes (for example, At-211, 1-131, 1-125, Y-90, Re-186, Re-188, Sm-153, Bi- 212, P-32 and the radioactive isotopes of Lu). Sources of radiation suitable for use as a cell conditioner of the present invention include both solids and liquids.
[00734] [00734] CD73 blockade can also be effectively combined with chemotherapeutic regimens. In such cases, it may be possible to reduce the dose of chemotherapeutic reagent administered.
[00735] [00735] Exemplary cytotoxic agents that can be administered in combination with an anti-CD73 antibody molecule include antimicrotubule agents, topoisomerase inhibitors, antimetabolites, mitotic inhibitors, alkylating agents, anthracyclines, vinca alkaloids, intercalating agents, capable agents to interfere with a signal transduction pathway, agents that promote apoptosis, proteasome inhibitors and radiation (for example, local or whole body irradiation).
[00736] [00736] In certain modalities, any of the combinations disclosed in this document, alternatively or in combination, additionally includes one or more of the agents described in Table 18. Table 18. Selected therapeutic agents that can be administered in combination with anti-CD73 antibody molecules, for example, as a single agent or in combination with other immunomodulators described herein. Each publication listed in this Table is incorporated into this document as a reference in its entirety, including all structural formulas therein.
[00737] [00737] In some embodiments, the additional therapeutic agent is chosen from one or more of: 1) a protein C kinase inhibitor (PKC); 2) a heat shock protein inhibitor 90 (HSP90); 3) a phosphoinositide 3-kinase inhibitor (PISK) and / or rapamycin target (mMTOR); 4) a cytochrome P450 inhibitor (for example, a CYP17 inhibitor or a 17 alpha-hydroxylase / C17-20 lyase inhibitor); 5) an iron chelating agent; 6) an aromatase inhibitor; 7) a p53 inhibitor, for example, an inhibitor of a p53 / Mdm 2 interaction; 8) an apoptosis inducer; 9) an angiogenesis inhibitor; 10) an aldosterone synthase inhibitor; 11) a smoothed receptor inhibitor (SMO); 12) a prolactin receptor inhibitor (PRLR); 13) a Wnt signaling inhibitor; 14) a CDKA4 / 6 inhibitor; 15) a fibroblast growth factor receptor (FGFR2) inhibitor / fibroblast growth factor receptor 4 (FGFRA4); 16) a macrophage colony stimulating factor inhibitor (M-CSF); 17) an inhibitor of one or more of c-KIT, histamine release, FIt3 (for example, FLK2 / STK1) or PKC; 18) an inhibitor of one or more of VEGFR-2 (for example, FLK-1 / KDR), PDGFRbeta, c-KIT or Raf kinase C; 19) a somatostatin agonist inhibitor and / or a growth hormone release; 20) an anaplastic lymphoma kinase inhibitor (ALK); 21) an insulin-like growth factor receptor inhibitor (IGF-1R); 22) a P-glycoprotein 1 inhibitor; 23) an inhibitor of vascular endothelial growth factor receptor (VEGFR); 24) a BCR-ABL kinase inhibitor; 25) an FGFR inhibitor; 26) a CYP11B2 inhibitor; 27) an HDM2 inhibitor, for example, an inhibitor of HDM2-p53 interaction; 28) a tyrosine kinase inhibitor; 29) a c-MET inhibitor; 30) a JAK inhibitor; 31) a DAC inhibitor; 32) a 118B-hydroxylase inhibitor; 33) an IAP inhibitor; 34) a PIM kinase inhibitor; 35) a Porcupine inhibitor; 36) a BRAF inhibitor, for example, BRAF V600E or wild-type BRAF; 37) an HER3 inhibitor; 38) a MEK inhibitor; or 39) a lipid kinase inhibitor, for example, as described in this document and Table 18.
[00738] [00738] The exemplary tyrosine kinase inhibitor includes, but is not limited to, an epidermal growth factor (EGF) pathway inhibitor (eg, an epidermal growth factor receptor (EGFR) inhibitor), a vascular endothelial growth factor (VEGF) pathway inhibitor (for example, a vascular endothelial growth factor (VEGFR) receptor inhibitor (for example, a VEGFR-1 inhibitor, a VEGFR-2 inhibitor , a VEGFR-3 inhibitor)), a platelet-derived growth factor (PDGF) pathway inhibitor (for example, a platelet-derived growth factor receptor (PDGFR) inhibitor (for example, an inhibitor PDGFR-RB)), an RAF-1 inhibitor, a KIT inhibitor and a RET inhibitor.
[00739] [00739] In one embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with a PKC inhibitor, Sotrastaurine (Compound A1), or a compound disclosed in PCT Publication No. WO 2005/039549, to treat a disorder, for example, a disorder described in the present document. In one embodiment, the PKC inhibitor is Sotrastaurine (Compound A1) or a compound disclosed in PCT Publication No.
[00740] [00740] In one embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with a BCR-ABL inhibitor, TASIGNA (Compound A2 ), or a compound disclosed in PCT Publication No. WO 2004/005281, to treat a disorder, for example, a disorder described in the present document. In one embodiment, the BCR-ABL inhibitor is TASIGNA or a compound disclosed in PCT Publication No. WO 2004/005281. In one embodiment, an anti-CD73 antibody molecule is used in combination with TASIGNA (Compound A2), or a compound as described in PCT Publication No. WO 2004/005281, to treat a disorder such as lymphocytic leukemia, Parkinson's disease, neurological cancer, melanoma, digestive / gastrointestinal cancer, colorectal cancer, myeloid leukemia, head and neck cancer or pulmonary hypertension.
[00741] [00741] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with an HSP90 inhibitor, to treat a disorder, for example , a disorder described in this document, for example, cancer, multiple myeloma, non-small cell lung cancer, lymphoma, gastric cancer, breast cancer, digestive / gastrointestinal cancer, pancreatic cancer , colorectal cancer, solid tumor or hematopoiesis disorder.
[00742] [00742] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, is used in combination with an inhibitor of PI3K and / or mMTOR, 8- (6-Methoxy -piridin-3-yl) -3-methyl-1- (4-piperazin-1-yl-3-trifluoromethyl-phenyl) -1,3-dihydro-imidazo [4,5-c] quinolin-2- ona (Compound A41), to treat a disorder, for example, a disorder described in this document. In one embodiment, the PI3K and / or mTOR inhibitor is 8- (6-Methoxy-pyridin-3-yl) -3-methyl-1- (4-piperazin-1-yl-3-trifluoromethyl-phenyl) -1 , 3-dihydro-imidazo [4,5-c] quinolin-2-ovna / (Compound A41). In one embodiment, an anti-CD73 antibody molecule is used in combination with 8- (6-Methoxy-pyridin-3-yl) -3-methyl-1- (4-piperazin-1-yl-3-trifluoromethyl-phenyl ) -1,3-dihydro-imidazo [4,5-clquinolin-2-o0na (Compound A41), to treat a disorder such as cancer, prostate cancer, leukemia (eg, lymphocytic leukemia), a breast cancer, brain cancer, bladder cancer, pancreatic cancer, kidney cancer, solid tumor or liver cancer.
[00743] [00743] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with a FGFR inhibitor, 3- (2,6 -dichloro-3,5-dimethoxyphenyl) -1- (6- ((4- (4-ethylpiperazin-1-yl) phenyl) amino) pyrimidin-4-yl) -1-methylurea (Compound A5) or a compound disclosed in US Patent 8,552,002, to treat a disorder, for example, a disorder described in the present document. In one embodiment, the FGFR inhibitor is 3- (2,6-dichloro-3,5-dimethoxyphenyl) -1- (6 - ((4- (4-ethylpiperazin-1-yl) phenyl) amino) pyrimidin-4 -il) - 1I-methylurea (Compound A5) or a compound disclosed in US Patent No. 8,552,002. In one embodiment, an anti-CD73 antibody molecule is used in combination with Compound A5, or a compound as described in U.S. No. 8,552,002, to treat a disorder
[00744] [00744] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with a PISK inhibitor, Buparlisib (Compound A6), or a compound disclosed in PCT Publication No. WO 2007/084786, to treat a disorder, for example, a disorder described in this document. In one embodiment, the PISK inhibitor is Buparlisib (Compound A6) or a compound disclosed in PCT Publication No. WO 2007/084786. In one embodiment, an anti-CD73 antibody molecule is used in combination with Buparlisib (Compound A6), or a compound disclosed in PCT Publication No. WO 2007/084786, to treat a disorder like, prostate cancer, cancer of non-small cell lung, endocrine cancer, leukemia, ovarian cancer, melanoma, bladder cancer, breast cancer, cancer of the female reproductive system, digestive / gastrointestinal cancer, cancer colorectal cancer, a glioblastoma multiforme, a solid tumor, a non-Hodgkin's lymphoma, a hematopoiesis disorder or a head and neck cancer. Compound A6 has the following
[00745] [00745] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with an FGFR inhibitor, 8- (2,6 -difluoro-3,5-dimethoxyphenyl) -N- (4- ((dimethylamino) methyl) -1H-imidazol-2-yl) quinoxaline-5-carboxamide (Compound A7) or a compound disclosed in PCT Publication No. WO 2009 / 141386 to treat a disorder, for example, a disorder described in this document. In one embodiment, the FGFR inhibitor is 8- (2,6-difluoro-3,5-dimethoxyphenyl) -N- (4 - ((dimethylamino) methyl) - 1H-imidazol-2-yl) quinoxaline-5-carboxamide (Compound A7) or a compound disclosed in PCT Publication No. WO 2009/141386. In one embodiment, the FGFR inhibitor is 8- (2,6-difluoro-3,5-dimethoxyphenyl) -N- (4 - ((dimethylamino) methyl) -1 H-imidazol-2-yl) | quinoxaline-5 - carboxamide (Compound A7). In one embodiment, an anti-CD73 antibody molecule is used in combination with 8- (2,6-difluoro-3,5-dimethoxyphenyl) -N- (4 - ((dimethylamino) methyl) -1H-imidazole | -2 -il) quinoxaline-5-carboxamide (Compound A7), or a compound disclosed in PCT Publication No. WO 2009/141386, to treat a disorder like a cancer characterized by angiogenesis.
[00746] [00746] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with a PI3K inhibitor, (S) -N1- ( 4-methyl-5- (2- (1,1,1-trifluoro-2-methylpropan-2-yl) pyridin-4-yl) thiazol-2-yl) pyrrolidine-1,2-dicarboxamide
[00747] [00747] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with a cytochrome P450 inhibitor (for example, the inhibitor of CYP17) or a compound disclosed in PCT Publication No. WO 2010/149755, to treat a disorder, for example, a disorder described herein. In one embodiment, the cytochrome P450 inhibitor (for example, the CYP17 inhibitor) is CFG920 or a compound disclosed in PCT Publication No. WO 2010/149755; U.S. No. 8,263,635 B2; or EP 2445903 B1. In one embodiment, an anti-CD73 antibody molecule is used in combination with a compound disclosed in PCT Publication No. WO 2010/149755, to treat prostate cancer.
[00748] [00748] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with an HDM2 inhibitor, (S) -1- (4-chlorophenyl) -7-isopropoxy-6-
[00749] [00749] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with an iron chelating agent, Deferasirox (also known as EXJADE; Compound A11), or a compound disclosed in PCT Publication No. WO 1997/049395 to treat a disorder, for example, a disorder described in this document. In one embodiment, the iron chelating agent is Deferasirox or a compound disclosed in PCT Publication No. WO 1997/049395. In one embodiment, the iron chelating agent is Deferasirox (Compound A11). In one instance, an anti-CD73 antibody molecule is used in combination with Deferasirox (Compound A11), or a compound disclosed in PCT Publication No. WO 1997/049395, to treat iron overload, hemochromatosis or myelodysplasia.
[00750] [00750] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with an aromatase inhibitor, Letrozole (also known as mo FEMARA; Compound A12), or a compound disclosed in US No. 4,978,672 to treat a disorder, for example, a disorder described in this document. In one embodiment, the aromatase inhibitor is Letrozole (Compound A12) or a compound disclosed in U.S. Patent No. 4,978,672. In one embodiment, an anti-CD73 antibody molecule is used in combination with Letrozole (Compound A12), or a compound disclosed in U.S. Patent No. 4,978,672, to treat a disorder such as cancer, leiomyosarcoma, endometrial cancer, a breast cancer, a cancer of the female reproductive system or a hormonal deficiency.
[00751] [00751] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with a PISK inhibitor, for example, an inhibitor of pan-PI3K, (48,5R) -3- (2'-amino-2-morpholino-4 '- (trifluoromethyl) - [4,5'-bipyrimidin] -6-yl) - 4- (hydroxymethyl) -5 -methyloxazolidin-2-0na (Compound A13) or a compound disclosed in PCT Publication No. WO2013 / 124826 to treat a disorder, for example, a disorder described in this document. In one embodiment, the PI3SK inhibitor is (48.5R) -3- (2'-amino-2-morpholino-4 '- (trifluoromethyl) - [4,5'-bipyrimidin] -6-yl) -4- (hydroxymethyl) -5-methyloxazolidin-2-one (Compound A13) or a compound disclosed in PCT Publication No. WO2013 / 124826. In one embodiment, an anti-CD73 antibody molecule is used in combination with (48.5R) -3- (2'-amino-2-morpholino-4 '- (trifluoromethyl) - [4,5'-bipyrimidin] - 6-yl) - 4- (hydroxymethyl) -5-methyloxazolidin-2-0na (Compound A13), or a compound disclosed in PCT Publication WO2013 / 124826, to treat a disorder such as cancer or tumor advanced solid.
[00752] [00752] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with a p53 inhibitor and / or a p53 interaction / Mdm2, (S) -5- (5-chloro-1-methyl-2-0x0-1,2-dihydropyridin-3-yl) -6- (4-chloropheni |) -2- (2.4 - dimethoxypyrimidin-5-yl) -1-isopropyl-5,6-dihydropyrrolo [3,4-d] imidazole- 4 (1H) -one (Compound A14), or a compound disclosed in PCT Publication No. WO2013 / 111105 to treat a disorder, for example, a disorder described in this document. In one embodiment, the inhibitor of p53 and / or an interaction of p53 / Mdm2 is (S) -5- (5-chloro-1-methyl-2-0x0-1,2-dihydropyridin-3- yl) -6- (4-chlorophenyl) -2- (2,4-dimethoxypyrimidin-5-yl) -1-isopropyl-5,6-dihydropyrrolo [3,4-d] imidazo | - 4 (1H) -one (Compound A14) or a compound disclosed in PCT Publication No. WOZ2013 / 111105. In one embodiment, an anti-CD73 antibody molecule is used in combination with (S) -5- (5-chloro-1-methyl-2-0x0-1,2-dihydropyridin-3-yl) -6- (4-chlorophenyl) -2- (2,4-dimethoxypyrimidin-5-yl) -1-isopropyl-5,6-dihydropyrrolo [3,4-d] imidazo | - 4 (1H) -one (Compound A14 ), or a compound disclosed in PCT Publication No. WO2013 / 111105, to treat a disorder such as cancer or soft tissue sarcoma.
[00753] [00753] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with a CSF-1R tyrosine kinase inhibitor, 4 - (((2 - (((1R, 2R) -2-hydroxycyclohexyl) amino) benzo [d] thiazol-6-yl) Oxy) -N-methylpicolinamide (Compound A15), or a compound disclosed in the PCT Publication in WO 2005/073224 to treat a disorder, for example, a disorder described in this document. In one embodiment, the CSF-IR tyrosine kinase inhibitor is 4 ((2 - (((1R, 2R) -2-hydroxycyclohexyl) amino) benzo [d] thiazol-6-yl) oxy) -N- methylpicolinamide (Compound
[00754] [00754] In certain embodiments, the CSF-1R tyrosine kinase inhibitor, - 4 ((2 - (((1R, 2R) -2-hydroxycyclohexyl) amino) benzo [d] thiazol-6- iN) oxide ) -N-methylpicolinamide (Compound A15), or a compound disclosed in PCT Publication No. WO 2005/073224, is administered in combination with the CD73 inhibitor (for example, the anti-CD73 antibody molecule) to treat a cancer, for example, a solid tumor (for example, an advanced solid tumor). Exemplary cancers that can be treated by the combination include, but are not limited to, brain cancer (e.g., glioblastoma multiforme (GBM), e.g., recurrent glioblastoma), breast cancer (e.g., triple negative breast cancer ( for example, NTBC)) or a pancreatic cancer (for example, advanced pancreatic cancer). Common features of these cancers include, for example, a tumor biology characterized by high levels of TAMs in the tumor microenvironment that can contribute to immunosuppression and immunosuppression. In some embodiments, blocking CSF-1R in conjunction with anti-CD73 therapy may, for example, promote reprogramming of TAMs and / or remove the immunosuppression of tumor-infiltrating lymphocytes (TIL).
[00755] [00755] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with an apoptosis inducer and / or an angiogenesis inhibitor , as Imatinib mesylate (also known as GLEEVECGO;
[00756] [00756] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with a JAK inhibitor, 2-fluoro-N- methyl-4- (7- (quinolin-6-ylmethyl) imidazo [1,2-b] [1,2,4] triazin-2-yl) benzamide (Compound A17), or a dihydrochloric salt thereof, or a compound disclosed in PCT Publication No. WO 2007/070514, to treat a disorder, for example, a disorder described herein. In a modality, the JAK inhibitor is 2-fluoro-N-methyl-4- (7- (quinolin-6-ylmethyl) imidazo [1,2-b] [1,2,4] triazin-2- il) benzamide (Compound A17), or a dihydrochloric salt thereof, or a compound disclosed in PCT Publication No. WO 2007/070514. In one embodiment, an anti-CD73 antibody molecule is used in combination with 2-fluoro-N-methyl-4- (7- (quinolin-6-ylmethyl) imidazo [1,2-b] [1,2 , 4] triazin-2-yl) benzamide (Compound A17), or a dihydrochloric salt thereof, or a compound disclosed in PCT Publication No. WO 2007/070514, to treat a disorder such as colorectal cancer, myeloid leukemia, cancer hematological, autoimmune disease, non-Hodgkin's lymphoma or thrombocythemia.
[00757] [00757] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with a JAK inhibitor, Ruxolitinib phosphate (also known - as JAKAFI; Compound A18) or a compound disclosed in PCT Publication No. WO 2007/070514 to treat a disorder, for example, a disorder described herein. In one instance, the JAK inhibitor is ruxolitinib phosphate (Compound A18) or a compound disclosed in PCT Publication No. WO 2007/070514. In one embodiment, an anti-CD73 antibody molecule is used in combination with ruxolitinib phosphate (Compound A18), or a compound disclosed in PCT Publication No. WO 2007/070514, to treat a disorder such as prostate cancer, leukemia lymphocytic, multiple myeloma, lymphoma, lung cancer, leukemia, cachexia, breast cancer, pancreatic cancer, rheumatoid arthritis, psoriasis, colorectal cancer, myeloid leukemia, hematological cancer, one autoimmune disease, non-Hundgkin lymphoma or thrombocythemia.
[00758] [00758] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with a deacetylase inhibitor (DAC), Panobinostate (Compound A19), or a compound disclosed in PCT Publication No. WO 2014/072493 to treat a disorder, for example, a disorder described herein. In one embodiment, the DAC inhibitor is Panobinostat (Compound A19) or a compound disclosed in PCT Publication No. WO 2014/072493. In one embodiment, a molecule
[00759] [00759] In some embodiments, the combination described in this document includes a deacetylase inhibitor (DAC), Panobinostate (Compound A19), or a compound disclosed in PCT Publication No. WO 2014/072493, and an inhibitor of an immune checkpoint molecule, for example, a CD73 inhibitor (for example, an anti-CD73 antibody molecule).
[00760] [00760] In one embodiment, the DAC inhibitor, Panobinostate (Compound A19), or a compound disclosed in PCT Publication No. WO 2014/072493, is administered in combination with the CD73 inhibitor (for example, the anti-antibody molecule -CD73) to treat colorectal cancer (for example, an MSS CRC), lung cancer
[00761] [00761] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with an inhibitor of one or more among cytochrome P450 (eg example, 11B2), aldosterone or angiogenesis, Osilodrostate (Compound A20), or a compound disclosed in PCT Publication No. WO 2007/024945 to treat a disorder, for example, a disorder described herein. In one embodiment, the inhibitor of one or more of cytochrome P450 (for example, 11B2), aldosterone or angiogenesis is Osilodrostate (Compound A20) or a compound disclosed in PCT Publication No. WO2007 / 024945. In one embodiment, an anti-CD73 antibody molecule is used in combination with Osilodrostate (Compound A20), or a compound disclosed in PCT Publication No. WO2007 / 024945, to treat a disorder such as Cushing's syndrome, hypertension or heart failure therapy.
[00762] [00762] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with an IAP inhibitor, (S) -N- ((S) -1-cyclohexyl-2 - ((S) -2- (4- (4-fluorobenzoyl) thiazol-2-yl) pyrrolidin-1-i1) -2-0x0ethyl) -2- (methylamino ) propanamide (Compound A21) or a compound disclosed in US No. 8,552,003 to treat a disorder, for example, a disorder described herein. In one embodiment, the IAP inhibitor is (S) -N - ((S) -1-cyclohexyl-2 - ((S) -2- (4- (4-fluorobenzoyl) thiazol-2-yl) pyrrolidin -1-i1) -2-0x0ethyl) -2- (methylamino) propanamide - (Compound A21) or a compound disclosed in US Patent No. 8,552,003. In one embodiment, an anti-CD73 antibody molecule is used in combination with (S) -N - ((S) -1-cyclohexyl-2 - ((S) -2- (4- (4-fluorobenzene) zoyl) thiazol-2-yl) pyrrolidin-1-11) -2-0x0ethyl) -2- (methylamino) propanamide (Compound A21), or a compound disclosed in U.S. Patent No.
[00763] [00763] In another embodiment, the combination, for example, a combination that comprises an anti-CD73 antibody molecule as described herein, includes or is used in combination with a Smoothened - (SMO) inhibitor, (R) -2- (5- (4- (6- benzyl-4,5-dimethylpyridazin-3-yl) -2-methylpiperazin-1-yl) pyrazin-2-yl) propan-2-0l (Compound A25 ), or a compound disclosed in PCT Publication No. WO 2010/007120 to treat a disorder, for example, a disorder described herein. In one embodiment, the SMO inhibitor is (R) -2- (5- (4- (6-benzyl-4,5-dimethylpyridazin-3-yl) -2-methylpiperazin-1-yl) pyrazin- 2-yl) propan-2-o0l (Compound A25) or a compound disclosed in PCT Publication No. WO 2010/007120. In one fashion, an anti-CD73 antibody molecule is used in combination with (R) -2- (5- (4- (6-benzyl-4,5-dimethylpyridazin-3-i1) -2-methylpiperazin- 1-yl) pyrazin-2-yl) propan-2-ol (Compound A25), or a compound disclosed in PCT Publication No. WO 2010/007120 to treat a disorder such as cancer, medulloblastoma, cell lung cancer prostate cancer, basal cell carcinoma, cancer
[00764] [00764] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with an AIlk inhibitor, ceritinib (also known as ZYKADIA ; Compound A23) to treat a disorder, for example, a disorder described herein. In one embodiment, the AIk inhibitor is ceritinib (Compound A23). In one embodiment, an anti-CD73 antibody molecule is used in combination with ceritinib (Compound A23), to treat a disorder such as non-small cell lung cancer or solid tumors.
[00765] [00765] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with a JAK and / or CDKA4 / 6 inhibitor, 7-cyclopentyl-N N-dimethyl-2- ((5- (piperazin-1-yl) pyridin-2-yl) amino) -7 H-pyrrolo [2,3-d] pyrimidine-6-carbo-xamide ( Compound A24), or a compound disclosed in U.S. Patent No.
[00766] [00766] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with a prolactin receptor inhibitor (PRLR), a human monoclonal antibody molecule (Compound A26) as disclosed in U.S. Patent No. 7,867,493), to treat a disorder, for example, a disorder described herein. In one embodiment, the PRLR inhibitor is a human monoclonal antibody (Compound A26) disclosed in U.S. No. 7,867,493. In one embodiment, an anti-CD73 antibody molecule is used in combination with the human monoclonal antibody molecule (Compound A26) described in U.S. Patent No. 7,867,493 to treat a disorder such as cancer, prostate cancer or breast cancer.
[00767] [00767] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with a PIM kinase inhibitor, N- (4- ((1R, 38.58) -3-amino-5-methylcyclohexyl) pyridin-3-i1) -6- (2,6-difluorophenyl) -5-fluoropicolinamide (Compound A27) or a compound disclosed in the Publication of PCT No. WO 2010/026124 to treat a disorder, for example, a disorder described in this document. In one embodiment, the PIM Kinase inhibitor is N- (4 - ((1R, 38.5S) -3-amino-5-methylcyclohexyl) pyridin-3-yl) - 6- (2,6-difluorophenyl) -S-fluoropicolinamide (Compound A27) or a compound disclosed in PCT Publication No. WO 2010/026124. In one embodiment, an anti-CD73 antibody molecule is used in combination with N- (4 - ((1R, 38.58) -3-amino-5-methylcyclohexyl) pyridin-3-i1) -6 - (2,6-difluorophenyl) -S-fluoropicolinamide (Compound A27), or a compound disclosed in PCT Publication No. WO 2010/026124, to treat a disorder such as multiple myeloma, myelodysplastic syndrome, myeloid leukemia or a non-Hodgkin lymphoma.
[00768] [00768] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with a Wnt signaling inhibitor, 2- (2 ', 3-dimethyl- [2,4'-bipyridin] -5-yl) - N- (5- (pyrazin-2-yl) pyridin-2-yl) acetamide (Compound A28) or a compound disclosed in the publication of PCT No. WO 2010/101849 to treat a disorder, for example, a disorder described in this document. In one embodiment, the Wnt signaling inhibitor is 2- (2 ', 3-dimethyl- [2,4'-bipyridin] -5-yl) -N- (5- (pyrazin-2-yl) pyridin-2 -yl) acetamide (Compound A28) or a compound disclosed in PCT publication No. WO 2010/101849. In one embodiment, the Wnt signaling inhibitor is 2- (2 ', 3-dimethyl- [2,4'-bipyridin] -5-yl) -N- (5- (pyrazin-2-yl) pyridin-2 - iNacetamide (Compound A28). In one embodiment, an anti-CD73 antibody molecule is used in combination with 2- (2 ', 3-dimethyl- [2.4 "- bipyridin] -5-yl) - N- ( 5- (pyrazin-2-yl) pyridin-2-yl) acetamide (Compound A2Z8), or a compound disclosed in PCT publication No. WO 2010/101849, to treat a disorder such as a solid tumor (for example, a cannula) head and neck cancer, squamous cell carcinoma, breast cancer, pancreatic cancer or colon cancer. In some modalities, cancer is chosen from skin cancer (for example, a melanoma), a solid microsatellite (MSl-high) solid tumor, pancreatic cancer or breast cancer (for example, triple negative breast cancer (TNBC)).
[00769] [00769] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with a BRAF inhibitor, to treat a disorder, for example For example, a disorder described in this document, for example, non-small cell lung cancer, melanoma or colorectal cancer.
[00770] [00770] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with a CDKA4 / 6, 7-cyclopentyl inhibitor N, N-dimethyl-2 - ((5- ((1R, 6S) -9-methyl-4-0x0-3,9-diazabicyclo [4.2.1] nonan-3-yl) pyridin-2-yl) ami - no) -7H-pyrrolo [2,3-d] pyrimidine-6-carboxamide (Compound A30), or a compound disclosed in PCT publication No. WO 2011/101409 to treat a disorder, for example, a disorder described herein document. In one embodiment, the CDK4 / 6 inhibitor is 7-cyclopentyl-N, N-dimethyl-2 - ((5 - ((1R, 68S) -9-methyl-4-0x0-3,9-diazabicyclo [4.2. 1] nonan-3-yl) pyridin-2-yl) amino) -7 H-pyrrolo [2,3-d] pyrimidine-6-carboxamide (Compound A30) or a compound disclosed in PCT publication No. WO 2011 / 101409. In one embodiment, an anti-CD73 antibody molecule is used in combination with 7-cyclopentyl-N, N-dimethyl-2 - ((5- ((1R, 68S) -9-methyl-4-0x0-3,9 -diazabicyclo [4.2.1] nonan-3-yl) pyridin-2-iN)> amino) -7H-pyrrolo [2,3-d] pyrimidine-6-carboxamide (Compound A3O), or a compound disclosed in the publication of PCT No. WO 2011/101409, to treat a disorder such as cancer, mantle cell lymphoma, liposarcoma, non-small cell lung cancer, melanoma, squamous cell esophageal cancer or breast cancer .
[00771] [00771] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with an HER3 inhibitor, (Compound A31), or a compound disclosed in PCT Publication No. WO 2012/022814, to treat a disorder, for example, a disorder described herein. In one embodiment, the HER3 inhibitor is (Compound A31) or a compound disclosed in PCT Publication WO 2012/022814. In one embodiment, an anti-CD73 antibody molecule is used in combination with Compound A31, or a compound disclosed in the Publication.
[00772] [00772] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with an FGFR2 and / or FGFR4 inhibitor, Compound A32, or a compound disclosed in a publication PCT Publication No. WO 2014/160160 (for example, an antibody molecule drug conjugated against a FGFR2 and / or FGFRA, for example, mAb 12425), to treat a disorder, for example, a disorder described in this document. In one embodiment, the FGFR2 and / or FGFRA4 inhibitor is Compound A32 or a compound disclosed in PCT Publication publication WO 2014/160160. In one embodiment, an anti-CD73 antibody molecule is used in combination with Compound A32, or a compound as described in Table 18, to treat a disorder such as cancer, gastric cancer, breast cancer ma, rhabdomyosarcoma, liver cancer, adrenal cancer, lung cancer, esophageal cancer, colon cancer or endometrial cancer.
[00773] [00773] In some embodiments, compound A32 is an antibody molecule drug conjugate against a FGFR2 and / or FGFRA, for example, mAb 12425.
[00774] [00774] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with an M-CSF inhibitor, Compound A33, or a compound disclosed in PCT Publication No. WO 2004/045532 (for example,
[00775] [00775] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with a MEK inhibitor, to treat a disorder as a non-small cell lung cancer, a multisystem genetic disorder, melanoma, ovarian cancer, digestive / gastrointestinal cancer, rheumatoid arthritis or colorectal cancer.
[00776] [00776] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with an inhibitor of one or more of c-KIT, histamine release, FIt3 (eg, FLK2 / STK1) or PKC, Midostaurin (Compound A35) or a compound disclosed in PCT Publication No. WO 2003/037347 to treat a disorder, for example, a disorder described in this document. In one embodiment, the inhibitor is Midastaurin (Compound A35) or the compound disclosed in PCT Publication No. WO 2003/037347. In one embodiment, the inhibitor of one or more of c-KIT, histamine release, FIt3 (eg, FLK2 / STK1) or PKC is Midostaurine. In one embodiment, an anti-CD73 antibody molecule is used in combination with Midostaurine (Compound A35), or compound disclosed in PCT Publication No.
[00777] [00777] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with a TOR inhibitor (for example, MTOR inhibitor ), Evero-limus (also known as AFINITOR; Compound A36) or a Compound disclosed in PCT Publication No. WO 2014/085318 to treat a disorder, for example, a disorder described in this document). In one embodiment, the TOR inhibitor is Everolimus (Compound A36) or a Compound disclosed in PCT Publication No. WO 2014/085318. In one embodiment, an anti-CD73 antibody molecule is used in combination with Everolimus (Compound A36) to treat a disorder such as colorectal cancer, interstitial lung disease, small cell lung cancer, respiratory cancer / chest cancer, prostate cancer, multiple myeloma, sarcoma, age-related macular degeneration, bone cancer, tuberous sclerosis, non-small cell lung cancer, endocrine cancer, lymphoma, neurological disorders, an astrocytoma, cervical cancer, neurological cancer, leukemia, immune disorders, transplant rejection, gastric cancer, melanoma, epilepsy, breast cancer (eg triple negative breast cancer (TNBC ) or a bladder cancer. In some modalities, the cancer is chosen from colorectal cancer (for example, microsatellite stable colorectal cancer (MSS CRC)), lung cancer (for example, a non-small cell lung cancer ena) or breast cancer (for example, triple negative lung cancer (TNBC)).
[00778] [00778] In some embodiments, the combination described in this document includes the mMTOR inhibitor, everolimus (Compound A36) or a compound disclosed in PCT Publication No. WO 2014/085318 and an inhibitor of an immune checkpoint molecule , for example, a CD73 inhibitor (for example, an anti-CD73 antibody molecule).
[00779] [00779] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with an inhibitor of one or more of VEGFR-2, PDGFRbeta, KIT or Raf kinase C, 1-methyl-5 - ((2- (5- (trifluoromethyl) -1 H-imidazol-2- iN) pyridin-4-yl) oxy) -N- (4- (trifluoromethyl) |) phenyl) - 1H-benzo [d] imidazole-2-amine (Compound A37) or a compound disclosed in PCT Publication No. WO 2007/030377 to treat a disorder, for example, a disorder described herein. In one embodiment, the inhibitor of one or more of VEGFR-2, PDGFRbeta, KIT or Raf kinase C is 1- methyl-5 - ((2- (5- (trifluoromethyl) -1 H-imidazol-2-yl) pyridine -4-i1) oxy) -N- (4- (triluoromethyl) phenyl) -1 H-benzo [d] imidazole-2-amine (Compound A37) or a compound disclosed in PCT Publication No. WO 2007/030377. In one embodiment, an anti-CD73 antibody molecule is used in combination with 1-methyl-5 - ((2- (5- (trifluoromethyl) -1H-imidazo | -2-yl) pyridin-4-yl) oxide) -N- (4- (trifluoromethi |) phenyl) -1H-benzo [d] imidazole-2-amine (Compound A37), or a compound disclosed in PCT Publication No. WO 2007/030377, to treat a disorder like a cancer , a melanoma or a solid tumor.
[00780] [00780] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with a somatostatin agonist and / or hormone release inhibitory growth, Pasireotid diaspartate (also known as SIGNIFOR; Compound A38) or a compound disclosed in PCT Publication No. WOZ2002 / 010192 or US Patent 7,473,761 to treat a disorder, for example, a disorder described in this document. In one embodiment, the somatostatin agonist and / or the inhibitory growth hormone release is Pasteurotide diaspartate (Compound A38) or a compound disclosed in PCT Publication No. WO2002 / 010192 or U.S. Patent No. 7,473,761. In one instance, an anti-CD73 antibody molecule is used in combination with Pasireotide diaspartate (Compound A38), or a compound disclosed in PCT Publication No. WO2002 / 010192 or US Patent No. 7,473,761, to treat a disorder like prostate cancer, endocrine cancer, neurological cancer, neuroendocrine group (NET) (for example, an atypical lung carcinoid tumor), skin cancer (for example, melanoma or carcinoma of Merkel cell), pancreatic cancer, liver cancer, Cushing's syndrome, gastrointestinal disorder, acromegaly, liver disorder and biliary tract or liver cirrhosis.
[00781] [00781] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with a signal transduction modulator and / or inhibitor of angiogenesis, for example, to treat a disorder such as cancer, respiratory / chest cancer, multiple myeloma, prostate cancer, non-small cell lung cancer, endocrine cancer or genetic disorder neurological.
[00782] [00782] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with an EGFR inhibitor, (R, E) - N- (7-chloro-1- (1- (4- (dimethylamino) but-2-enoyl) azepan-3-yl) -1H-benzo [d] imidazo | -2-yl) -2-methylisonicotin-
[00783] [00783] In some embodiments, the EGFR inhibitor, (R, E) -N- (7-chloro-1- (1- (4- (dimethylamino) but-2-enoyl) azepan-3-yl) -1H -benzo [d] imidezol-2-i1) -2-methylisonicotinamide (Compound A40), or a compound disclosed in PCT Publication No. WO 2013/184757, is administered in combination with a CD73 inhibitor (for example , an anti-CD73 antibody molecule) to treat colorectal cancer (CRC), lung cancer (for example, non-small cell lung cancer (NSCLC)), or breast cancer (for example, cancer triple negative breast cancer (TNBC)).
[00784] [00784] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with an ALK inhibitor, No.- (2-isopropoxy -5-methyl-4- (1-methylpiperidin-4-yl) phenyl) -Nº- (2- (isopropylsulfonyl) phenyl) -1 H-pyrazolo [3,4-d] lpyrimidine-4,6-diamine (Compound A42) or a compound disclosed in PCT Publication No. WO 2008/073687 to treat a disorder, for example, a disorder described herein. In one embodiment, the ALK inhibitor is Nº- (2-isopropoxy-5-methyl-4- (1-me-
[00785] [00785] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with an IGF-1IR inhibitor, 3- (4 - (4 - ((5-chloro-4 - (((5-methyl-1H-pyrazol-3-yl) amino) pyrimidin-2-yl) amino) -5-fluoro-2-methylphenyl) piperidin-1-ethyl 1,1-dioxide (Compound A43), 5-chloro-N2- (2-fluoro-5-methyl-4- (1- (tetrahydro-2H-pyran-4-yl) piperidin-4-yl) phenyl ) - Ni- (5-methyl-1 H-pyrazol-3-yl) pyrimidine-2,4-diamine - (Compound A44), or 5-chloro-N2- (4- (1-ethylpiperidin-4-iI) -2-fluoro-5-methylphenyl) -Nº- (5-methyl-1 H-pyrazol-3-iN) pyrimidine-2,4-diamine (Compound A45) or a compound disclosed in PCT Publication No. WO 2010/002655 to treat a disorder, for example, a disorder described. In one embodiment, the IGF-1R inhibitor is 3- (4- (4 - ((5-chloro-4 - ((5-methyl-1H-pyrazole-3 -yl) amino) pyrimidin-2-yl)> amino) -5-fluoro-2-methylphenyl) piperidin-1-yl) tiethane - 1,1-dioxide (Compound A43), 5-chlorine-No.- ( 2-fluoro- 5-methyl-4- (1- (tetrahydro-2H-pyran-4-yl) piperidin-4-yl) phenyl) - Nº- (5-methyl-1 H-pyrazol-3-yl) pyrimidine-2 , 4-diamine (Compound A44), B-chloro-N2- (4- (1-ethylpiperidin-4-i1) -2-fluoro-5-methylphenyl) - Nº- (5-methyl-1 H-pyrazol-3 -yl) pyrimidine-2,4-diamine (Compound A45), or a compound disclosed in PCT Publication No. WO 2010/002655. In one embodiment, an anti-HIV antibody molecule
[00786] [00786] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with a P-glycoprotein 1 inhibitor, Valspodar (also known as AMDRAY; Compound A46) or a compound disclosed in EP 296122 to treat a disorder, for example, a disorder described herein. In one embodiment, the P-glycoprotein 1 inhibitor is Valspodar (Compound A46) or a compound disclosed in EP 296122. In one embodiment, an anti-CD73 antibody molecule is used in combination with Valspodar (Compound A46), or a compound disclosed in EP 296122, to treat a disorder such as a cancer or a drug resistant tumor.
[00787] [00787] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with one or more of a VEGFR inhibitor, Vananib (Compound A47) or a compound disclosed in EP 296122 to treat a disorder, for example, a disorder described herein. In one embodiment, the VEGFR inhibitor is Vatalanib succinate (Compound A47) or a compound disclosed in EP 296122. In one embodiment, an antisense molecule
[00788] [00788] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with an HDI inhibitor or a compound disclosed in WO 2014/141104 to treat a disorder, for example, a disorder described in this document. In one embodiment, the HDI inhibitor is a compound disclosed in PCT Publication No. WO 2014/141104. In one embodiment, an anti-CD73 antibody molecule is used in combination with a compound disclosed in WO2014 / 141104 to treat a disorder like cancer.
[00789] [00789] In another embodiment, the combination, for example, a combination comprising an anti-CD73 antibody molecule as described herein, includes or is used in combination with a BCL-ABL inhibitor or a compound disclosed in Publications = PCT No. WOZ2Z013 / 171639, WOZ2Z013 / 171640, WO 2013/171641 or WO 2013/171642 to treat a disorder, for example, a disorder described in this document. In one mode, the BCL-ABL inhibitor is a compound disclosed in PCT Publications No. WO2013 / 171639, WO2013 / 171640, WO2013 / 171641 or WO2013 / 171642. In one embodiment, an anti-CD73 antibody molecule is used in combination with a compound disclosed in PCT Publications No. WOZ2013 / 171639, WOZ2013 / 171640, WO 2013/171641 or WO2013 / 171642 to treat a disorder like a cancer.
[00790] [00790] In another embodiment, the combination, for example, a combination that comprises an anti-CD73 antibody molecule as described herein, includes or is used in combinations
[00791] [00791] Methods of administering the antibody molecules are known in the art and are described below. Adequate dosages of the molecules used will depend on the age and weight of the subject and the particular drug used. The dosages and therapeutic regimens of the anti-CD73 antibody molecule can be determined by a skilled person.
[00792] [00792] In certain embodiments, the anti-CD73 antibody molecule is administered by injection (for example, intravenously) at a dose (for example, a flat dose) of about 60 mg to 2400 mg, for example, about from 100 mg to 2400 mg, about 100 mg to 2200 mg, about 100 mg to 2000 mg, about 100 mg to 1800 mg, about 100 mg to 1600 mg, about 100 mg to 1400 mg, about 100 mg to 1200 mg, about 100 mg to 1000 mg, about 100 mg to 800 mg,
[00793] [00793] In some embodiments, an anti-CD73 antibody molecule disclosed in this document is administered by injection (eg, subcutaneously or intravenously) at a dose (eg, a flat dose) of about 5 mg at 100 mg, about 100 mg to 500 mg, about 500 mg to 1000 mg, about 1000 mg to 1500 mg, about 1500 mg to 2000 mg, about 2000 mg to 2500 mg, about 2500 mg at 3000 mg, about 3000 mg to 3500 mg or about 3500 mg to 4000 mg, for example, once a week (QW), once every two weeks (Q2W) or once every four weeks ( Q4W). In certain embodiments, the antibody molecule is administered, for example, intravenously, at a dose of about 6 mg, about 20 mg, about 60 mg, about 200 mg, about 600 mg, about 1200 mg, about 2400 mg, about 3000 mg or about 3600 mg, for example, QW, Q2W or Q4W. In certain embodiments, the antibody molecule is administered, for example, intravenously, at a dose of about 60 mg Q2W. In certain embodiments, the antibody molecule is administered, for example, intravenously, at a dose of about 200 mg Q2W. In certain embodiments, the antibody molecule is administered, for example, intravenously, at a dose of about 600 mg Q2W. In certain embodiments, the antibody molecule is administered, for example, intravenously, at a dose of about 1200 mg Q2W. In certain embodiments, the antibody molecule is administered, for example, intravenously, at a dose of about 2400 mg Q2W. In certain embodiments, the antibody molecule is administered, for example, intravenously, at a dose of about 3000 mg Q2W. In certain embodiments, the antibody molecule is administered, for example, intravenously, at a dose of about 3600 mg Q2W.
[00794] [00794] In certain embodiments, the anti-CD73 antibody molecule is administered by injection (for example, intravenously) at a dose of about 1 to 30 mg / kg, for example, about 5 to 25 mg / kg , about 10 to 20 mg / kg, about 1 to 5 mg / kg, or about 3 mg / kg. In some embodiments, the anti-CD73 antibody molecule is administered at a dose of about 1 mg / kg, about 3 mg / kg, or 10 mg / kg, about 20 mg / kg, about 30 mg / kg or about 40 mg / kg. In some embodiments, the anti-CD73 antibody molecule is administered at a dose of about 1 to 3 mg / kg, or about 3 to 10 mg / kg. In some embodiments, the anti-CD73 antibody molecule is administered at a dose of about 0.5 to 2.2 to 4.2 to 5.5 to 150 mg at 20 mg / kg. The dosing schedule can vary, for example, once a week to once every 2, 3 or 4 weeks. In one embodiment, the anti-CD73 antibody molecule is administered at a dose of about 10 to 20 mg / kg every two weeks.
[00795] [00795] The antibody molecules can be used in non-conjugated or conjugated forms to a second agent, for example, a cytotoxic drug, radioisotope or a protein, for example, a protein toxin or a viral protein. This method includes: administering the antibody molecule, alone or in conjunction with a cytotoxic drug, to an individual who requires such treatment. Antibody molecules can be used to deliver a variety of therapeutic agents, for example, a cytotoxic moiety, for example, a therapeutic drug, a radioisotope, molecules of plant, fungal or bacterial origin or biological proteins (for example, toxins from protein) or particles (for example, recombinant viral particles, for example, by means of a viral coating protein), or mixtures thereof. Diagnostic Uses
[00796] [00796] In one aspect, the present invention provides a diagnostic method for detecting the presence of a CD73 protein in vitro (e.g., in a biological sample, such as a tissue biopsy, e.g., cancerous tissue) or in vivo (for example, in vivo imaging in an individual). The method includes: (i) placing the sample in contact with an antibody molecule described in this document, or administering the antibody molecule to the individual; (optionally) (ii) placing a reference sample, for example, a control sample (for example, a biological control sample, such as plasma, tissue, biopsy) or a control individual)); and (iii) detecting the formation of a complex between the antibody molecule and the sample or the individual, or the control or individual sample, in which a change, for example, a statistically significant change, in the formation of the complex in the sample or individual in relation to the control sample or individual is indicative of the presence of CD73 in the sample. The antibody molecule can be directly or indirectly identified with a detectable substance to facilitate detection of bound or unbound antibody. Suitable detectable substances include various enzymes, prosthetic groups, fluorescent materials, luminescent materials and radioactive materials, as described above and described in greater detail below.
[00797] [00797] The term "sample", since it refers to samples used to detect polypeptides, includes, but is not limited to, cells, cell layers, proteins or cell membrane extracts, body fluids or tissue samples.
[00798] [00798] The complex formation between the antibody molecule and CD73 can be detected by measuring or visualizing the binding molecule bound to the CD73 antigen or unbound binding molecule. Conventional detection assays can be used, for example, enzyme-linked immunosorbent assays (ELISA), a radioimmunoassay (RIA) or tissue immunohistochemistry. As an alternative to the identification of the antibody molecule, the presence of CD73 can be assayed in a sample by a competition immunoassay using standards identified with a detectable substance and an unidentified antibody molecule. In this assay, the biological sample, the identified standards and the antibody molecule are combined and the amount of identified standard bound to the unidentified binding molecule is determined. The amount of CD73 in the sample is inversely proportional to the amount of identified standard bound to the antibody molecule. Nucleic acids
[00799] [00799] The invention also features nucleic acids comprising nucleotide sequences that encode variable regions of heavy and light chain and CDRs or hypervariable loops of anti-CD73 antibody molecules, as described herein. For example, the invention features a first and second variable regions of heavy and light chain encoding nucleic acid, respectively, of an anti-CD73 antibody molecule chosen from one or more of the disclosed antibody molecules in this document. The nucleic acid can comprise a nucleotide sequence as shown in Table 1, or a sequence substantially identical to the same (for example, a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity). frequency with the same, or that differs by no more than 3, 6, 15, 30 or 45 nucleotides from the sequences shown in Table 1).
[00800] [00800] In certain embodiments, the nucleic acid may comprise a nucleotide sequence that encodes at least one, two or three CDRs, or hypervariable loops, from a variable region of heavy chain that has an amino acid sequence as shown. - seated in Table 1, or a sequence substantially homologous to it (for example, a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity to the same and / or that has one or more substitutions, for example, preserved substitutions). In other embodiments, the nucleic acid may comprise a nucleotide sequence that encodes at least one, two or three CDRs, or hypervariable loops, from a light chain variable region that has an amino acid sequence as shown in Table 1, or a sequence substantially homologous to the same (for example, a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity to the same and / or that has one, or more substitutions , for example, preserved substitutions). In yet another embodiment, the nucleic acid may comprise a sequence of nucleotides that encodes at least one, two, three, four, five or six CDRs or hypervariable loops, from variable regions of heavy and light chains that have a sequence amino acid as shown in Table 1, or a sequence substantially homologous to it (for example, a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity to it and / or that has one or more substitutions, for example, conserved substitutions).
[00801] [00801] In certain embodiments, the nucleic acid may comprise
[00802] [00802] In another aspect, the application presents host cells and vectors containing the nucleic acids described in this document. Nucleic acids can be present in a single vector or separate vectors present in the same host cell or separate host cells, as described in greater detail in the present document below.
[00803] [00803] Additionally, vectors comprising nucleotide sequences encoding an antibody molecule described herein are provided herein. In one embodiment, the vectors comprise nucleotides that encode an antibody molecule described herein. In one embodiment, the vectors comprise the nucleotide sequences described in this document. The vectors include, but are not limited to, a virus, plasmid, cosmid, lambda phage or an artificial yeast chromosome (YAC).
[00804] [00804] Numerous vector systems can be used. For example, a class of vectors uses elements of DNA that are derived from animal viruses, such as bovine papilloma virus, papilloma virus, adenovirus, vaccine virus, baculovirus, retro virus ( Rous sarcoma, MMTV or MOMLV) or SV40 virus. Another class of vectors uses RNA elements derived from RNA viruses such as Semliki Forest virus, Eastern Equine Encephalitis virus and Flavivirus.
[00805] [00805] - In addition, cells that have stably integrated DNA into their chromosomes can be selected by introducing one or more markers that allow the selection of transfected host cells. The marker can provide, for example, prototrophy to an auxotrophic host, resistance to biocide (e.g., antibiotics) or resistance to heavy metals such as copper or the like. The selectable marker gene can be directly linked to the DNA sequences to be expressed, or introduced into the same cell by cotransformation. Additional elements may also be required for optimal MRNA synthesis. These elements can include splicing signals, as well as transcriptional promoters, intensifiers and termination signals.
[00806] [00806] Once the expression vector or DNA sequence containing the constructs has been prepared for expression, the expression vectors can be transfected or introduced into a suitable host cell. Various techniques can be employed to achieve this purpose, such as protoplast fusion, calcium phosphate precipitation, electroporation, retroviral transduction, viral, biological transfection, lipid-based transfection or other conventional techniques. In the case of protoplasty fusion, cells are cultured in media and examined for appropriate activity.
[00807] [00807] The invention also provides host cells that comprise a nucleic acid that encodes an antibody molecule as described herein.
[00808] [00808] In one embodiment, host cells are genetically modified to comprise nucleic acids that encode the antibody molecule.
[00809] [00809] In one embodiment, the host cells are genetically modified using an expression cassette. The phrase "expression cassette" refers to nucleotide sequences, which are capable of affecting the expression of a gene in hosts compatible with such sequences. Such cassettes may include a promoter, an open reading frame with or without introns and a termination signal. Additional factors necessary or useful in effecting the expression can also be used, such as, for example, a promotion
[00810] [00810] The invention also provides host cells that comprise the vectors described in this document.
[00811] [00811] The cell can be, but without limitations, a eukaryotic cell, a bacterial cell, an insect cell or a human cell. Suitable eukaryotic cells include, but are not limited to, Vero cells, HeLa cells, COS cells, CHO cells (e.g., CHO-C8TD cells), HEK293 cells, BHK cells and MDCKII cells. Suitable insect cells include, but are not limited to, Sf9 cells. EXAMPLES
[00812] [00812] The examples below are presented to assist in understanding the inventions, however, they are not intended for, and should not be interpreted in such a way as to limit their scope in any way. Example 1: Generation and characterization of anti-CD73 antibodies Selection and optimization of anti-CD73 antibodies from synthetic yeast antibody libraries
[00813] [00813] Anti-CD73 monoclonal antibodies representing five distinct epitope bins were selected from eight libraries of virgin human synthetic yeast using the methods described below. Materials and methods
[00814] [00814] The antigens were biotinylated with the use of Pierce's Sulfo-NHS EZ-Link Biotinylation Kit. Goat F (ab ') FITC from human anti-kappa (LC-FITC), ExtrAvidin-PE (EA-PE) and Streptavidin-AF633 (SA-633) were obtained from Southern Biotech, Sigma and Molecular Probes , respectively. The Streptavidin and MACS LC Micro-Beads separation columns were purchased from Miltenyi Biotec. Goat PE anti-human IgG (Human PE) was obtained from Southern Biotech. Primary Finding
[00815] [00815] Eight libraries of virgin human synthetic yeast, each of —10º in diversity, were propagated as previously described (see, for example, Y. Xu et al, Addressing polyspecificity of antibodies selected from an in vitro yeast presentation system : a FACS-based, high-throughput selection and analytical tool.
[00816] [00816] The light chain diversification protocol was used during the primary finding phase for additional finding and enhancement of antibodies.
[00817] [00817] Light chain batch diversification protocol: The heavy chain plasmids from a virgin selection result were extracted from the yeast by crushing and picking, propagating
[00818] [00818] Antibody optimization was performed by introducing diversities into the heavy chain and variable regions of the light chain as described below.
[00819] [00819] Selection of CDRH1 and CDRH2: The CDRH3 of a single antibody was recombined in a pre-produced library with variants of CDRH1 and CDRH2 of a diversity of 1 x 10º and the selections were carried out with a cycle of MACS and four cycles of FACS as described in the virgin finding. In the different FACS cycles, the libraries were observed for PSR binding, cross-reactivity of species and pressure of affinity by titration or pre-formation of parental Fab complex and the classification was performed to obtain a population with the desired characteristics. Antibody production and purification
[00820] [00820] The yeast clones were cultured to saturation and then induced for 48 h at 30 ºC with agitation. After induction, yeast cells were pelleted and supernatants were collected for purification. IgGs were purified using a Protein A column and eluted with acetic acid, pH 2.0. Fab fragments were generated by papain digestion and purified using KappaSelect (GE Healthcare LifeSciences). ForteBio Kp measurements
[00821] [00821] ForteBio's affinity measurements were performed on an RED384 Octet usually as previously described (see, for example, Estep et al, High throughput solution-based measure of antibody-antigen affinity and epitope binning. Mabs 5 (2 ), 270 to
[00822] [00822] The ligand / epitope ligation block was performed using a standard sandwich cross-block assay. The control anti-target IgG was loaded on the AHQ sensors and unoccupied Fc binding sites on the sensor were blocked with an irrelevant human IgG1 antibody. The sensors were then exposed to 100 nM of the target antigen followed by a second antibody or anti-target ligand. Additional binding by the second antibody or ligand after association with the antigen indicates an unoccupied (non-competing) epitope, whereas no binding indicates epitope blocking (ligand or competitor blocking). MSD-SET kinetic assay
[00823] [00823] Balance affinity measurements performed as descriptions
[00824] [00824] Approximately 100,000 cells that overexpress the antigen were washed with wash buffer and incubated with 100 nM IgG at 100 μl for 5 minutes at room temperature. The cells were then washed twice with wash buffer and incubated with 100 µl of 1: 100 Human PE for 15 minutes on ice. The cells were then washed twice with wash buffer and analyzed on a FACS Canto |! (BD Biosciences). Results
[00825] [00825] Yeast cells that express a library of human antibodies on their surface have been examined for binding to human CD73. Two antibodies from epitope bin 4, 918 and 930, bound well to CD73 and inhibited the enzymatic activity of CD73 (data not shown). These two antibodies were subjected to affinity maturation that produced two strains of related antibodies, called lineage 1 and lineage 3, respectively (Table 19). These anti-CD73 antibodies were expressed in three different formats: IgG1 antibodies (called .C constructs, for example, 350.C), IgG4 antibodies that comprise an S228P mutation in the Fc region (IgG4 S228P, called .A constructs, for example 350.A) or IgG4 antibodies comprising S228P and L235E mutations in the Fc region (IgG4 S228P / L235E, called B constructs, for example 350.B), numbered according to Eu numbering. The sequences of these antibodies are disclosed in Table 1. For the 350.A antibody, two batches of antibodies were produced, hereinafter referred to as 350.A1 and 350.A2. Table 19. Two strains of anti-CD73 antibodies Parental jo18 Daughter EEM Daughter 358 [Daughter [376 Daughter 377
[00826] [00826] All tested anti-CD73 antibodies bind to human CD73 and cinomolgo. Lineage 1 antibodies also bind to murine CD73. Table 20 provides Ka values for these antibodies measured using Octet as described above.
[00827] [00827] Then, using ligand blocking / epitope binding studies, it was shown that the parental antibody 918 competed for binding to CD73 with progeny antibodies 350, 356 and
[00828] [00828] FabsdemaAbs 350 and 373 were generated by genetically modifying an interruption between the two proline residues above the 350 and 373 heavy chain nucleus hinge region. Both were expressed in Expi293F cells (ThermoFisher) and purified using CaptureSelect IgG CH1 affinity resin (ThermoFisher).
[00829] [00829] Biacore was used to measure affinity between species for Fab materials of mAbs 350 and 373. The proteins used were as follows: recombinant human CD73 (R&D Systems 5795-EN); Recombinant cynomolgus monkey CD73 (Sino Biological 90912-CO08H); Recombinant mouse CD73 (R&D Systems
[00830] [00830] Results for Ka (M) affinity for binding between 350 and 373 Fab species are shown in Table 21. Table 21. Affinities of anti-CD73 Fabs
[00831] [00831] In a separate study, the affinity of 373.A full-length antibody or 373A Fab fragments to human CD73, cynomol monkey, mouse and rat was determined using a Biacore method of captures anti-histidine antibody (His) using surface plasmon resonance (SPR). The anti-His Ab was directly immobilized on a CM5 chip surface by amine coupling. Human CD73 labeled His / His, CD73 monkey cynomolgo / His, mouse CD73 / His or rat CD73 / His was eluted and captured in a desired resonance unit (RU) of an Rmax of 20. Concentrations of antibody analyte in serial dilutions of IgG or Fab were eluted at 60 µl / min. The sensors were analyzed using the manufacturer's software for a 1: 1 connection model. Binding to mouse CD73 / His protein and mouse CD73 / His protein was undetectable for 373.A and
[00832] [00832] Affinities have been established for human and monkey cynomolgus CD73 with both 373.A and 373.A. The hydrogen-deuterium mass spectrometry and size exclusion chromatography studies support a CD73-dimer conformational locking model by 373.A in the open-open (inactive-inactive) conformation, supporting a bond bidentate of 1: 1 of 1 of Ab: 1 of CD73 dimer (see Example 2). Therefore, considering that the 1: 1 bidentate bond will favor avidity, the affinities of whole Ab were used instead of Fab measurements. The full length antibody 373.A binds to CD73 recombinant human with a Ka of 0.991 + 0.267 nM and cross-react with recombinant cynomolgus monkey CD73 with a Ka of 0.068 + 0.009 nM as determined by Biotech kinetic binding studies. Anti-CD73 Antibody Target Coupling
[00833] [00833] Whole blood target engagement was assessed by flow cytometry using whole blood from healthy human donors. Briefly, biotinylated antibodies were incubated with whole blood for 30 minutes before lysis and red blood cell fixation. Fixed cells were axed to CD3 and CD8 to identify CD8 + T cells, and streptavidin-APC to detect biotin. After staining, the cells were washed and subjected to flow cytometry analysis.
[00834] [00834] Dose-dependent binding, as measured by APC signal mean fluorescence intensity (MFI), was observed for the tested anti-CD73 antibodies (Figure 1). A biotinylated isotype control antibody showed no binding to CD8 + T cells (Figure 1).
[00835] [00835] The conceptual demonstration of the target occupancy (TO) of CD73 in whole human blood samples was performed by treating donor blood ex vivo with 373.A unidentified. A titration of 373.A or DNP-lgG4sm isotype control of 10 µg / mL at 0.17 ng / mL was titrated. As shown in Figure 24A, samples from the two donors that were treated with unidentified 373.A in larger doses (10 µg / mL to -0.1 µg / mL) prevented biotinylated 373.A from binding to cells , reducing the geometric mean fluorescence intensity (gMFI) values for a plateau at the fluorescence background level (-550 gMFI for both donors). This is indicative of target occupancy of total CD73. In contrast, gMFI values for cells that were pretreated with less than 373.A unidentified amounts (0.17 ng / mL and 0.51 ng / mL) were similar to samples that were pretreated with DNP-IlgG4sm isotype (-1600 gMFI for donor 1 and -2200 gMFI for donor 2). Samples treated with isotype control simulated blood that had zero target occupancy. The resulting% TO values are shown in Figure 24B. Inhibition of enzymatic activity of soluble recombinant CD73
[00836] [00836] CD73 of 5 'ectonucleotidase is the rate limiting step in the conversion of AMP to adenosine. The ability of anti-CD73 antibodies to inhibit the enzymatic activity of CD73 was measured using a malachite green phosphate assay. Briefly, 25 ng / ml of recombinant human CD73 was incubated with a dose titration of substrate adenosine monophosphate (AMP) (O at 500 µM) with buffer alone, or in the presence of an isotype control antibody at 1 µg / ml or the anti-CD73 350.C antibody at 1, 0.3 or 0.1 puo / ml. The release of inorganic phosphate (Pi) was measured using a malachite green phosphate assay kit (Enzo Life Sciences, nº
[00837] [00837] “As shown in Figure 2A, the control antibody at the concentration tested had no effect on the Michaelis constant (Km) of recombinant human CD73. In contrast, the anti-CD73 antibody
[00838] [00838] Next, anti-CD73 antibodies 350, 356, 373 and 374, expressed in .A or .B format, were tested for their ability to inhibit the enzymatic activity of recombinant human CD73 and cynomolgus monkey with the use of a malachite green phosphate assay as described above. Briefly, anti-CD73 antibodies were incubated for 10 minutes with 25 ng / ml human CD73 recombinant or cinomolg in the presence of 25 µM AMP. The release of inorganic phosphate (Pi) was measured using a malachite green phosphate assay kit (Enzo Life Sciences, Catalog number: BML-AK111). The normalized percentage inhibition (% INH) was determined using the zero time control as 100% INH and no antibody control as 0% INH.
[00839] [00839] “As shown in Figures 3A to 3C, all anti-CD73 antibodies tested inhibited the enzymatic activity of recombinant human CD73 or soluble cinomolgo. Inhibition of enzymatic activity of soluble endogenous CD73
[00840] [00840] Furthermore, the enzyme inhibiting activity of anti-CD73 antibodies has been tested against several endogenous CD73, for example, CD73 spilled from the cell surface.
[00841] [00841] In a first study, anti-CD73 antibodies 350 and 373, expressed in .A or .B format, or isotype control antibodies were incubated for 240 minutes with MDA-MB-231 serum-free medium (a strain human breast cancer cell) conditioned in the presence of 100 µM AMP. The disappearance of AMP was measured by a modified Glo Cell Titration (CTG) assay (Promega, Cat no. G9242 / 3). AMP inhibits the luciferase signal in the CTG kit. The luciferase signal increases as the added AMP is enzymatically consumed by CD73. The normalized percentage inhibition (% INH) was determined using the zero time control as 100% INH and no antibody control as 0% INH.
[00842] [00842] “As shown in Figure 4, the dose-dependent anti-CD73 antibodies inhibited the enzymatic activity of CD73 shed from the breast cancer cell line MDA-MB-231.
[00843] [00843] In a second study, anti-CD73 antibodies 350, 356, 358, 373, 374, 377 and 379, all expressed in .B format, were incubated for 60 minutes with diluted serum (12.5% v: v in PBS) of a pancreatic cancer patient in the presence of 100 µM AMP. Similarly to the first study, the disappearance of AMP was measured by the modified Cell Titration (CTG) assay and the normalized inhibition percentage (% INH) was determined using zero time control as 100% of INH and without antibody control such as 0% INH.
[00844] [00844] The anti-CD73 antibodies also inhibited CD73 enzyme activity in the pancreatic cancer patient's serum in a dose-dependent manner (Figure 5). Inhibition of CD73 enzymatic activity expressed on the cell surface
[00845] [00845] First, a malachite green phosphate assay was used to examine the ability of anti-CD73 antibodies 350, 356, 358, 373, 374, 377 and 379 (all in .B format) to inhibit CD73 expressed in a strain of breast cancer cell MDA-MB-231. Shortly, the antibodies were incubated for 180 minutes with cells in the presence of 100 µM AMP. The release of inorganic phosphate from AMP was measured using a malachite green phosphate assay kit (Enzo Life Sciences, Catalog number: BML-AK111). The normalized percentage inhibition (% INH) was determined using the zero time control as 100% INH and no antibody control as 0% INH.
[00846] [00846] “As shown in Figure 6, all anti-CD73 antibodies tested inhibited the enzymatic activity of CD73 expressed on the surface of the breast cancer cell line MDA-MB-231.
[00847] [00847] Next, since the antibodies of Lineage 1 cross-reacted with mouse CD73, whereas the antibodies of Lineage 3 did not, the antibodies of both strains were tested against CD73 expressed on the surface of a human or murine breast cancer cell line. Anti-CD73 antibodies were incubated for 240 minutes with a human breast cancer cell line MDA-MB-231 or a 4T1 murine breast cancer cell line in the presence of 100 µM AMP. The disappearance of AMP was measured by the modified Cell Titration Glo (CTG) assay described above and the percentage of normalized inhibition (% INH) was determined using zero time control as 100% INH and in the control of antibody the 0% INH.
[00848] [00848] Consistent with their binding profiles, antibodies from Line 1 918, 350, 356 and 358 inhibited both human and murine CD73 (Figures 7A and 7B), while antibodies from Line 3 930, 373, 374, 376, 377 and 379 inhibit human, but not murine, CD73 (Figures 7C and 7D).
[00849] [00849] In addition, two modified Cell Titration Glo (CTG) assays were conducted to test the enzyme inhibiting activity of anti-CD73 antibodies against CD73 expressed in a | i-
[00850] [00850] In both studies, all anti-CD73 antibodies tested were able to inhibit surface CD73 expressed in the human breast cancer cell line MDA-MB-231 or the cancer cell line of the human ovary SKOV3 (Figures 8A, 8B, 9A and 9B).
[00851] [00851] Next, a similar Glo Cell Titration (CTG) assay was performed to examine the ability of anti-CD73 antibodies to inhibit human CD73 expressed in a HEK 293 cell line. Briefly, a HEK 293 cell line was genetically modified to stably express human CD73 and incubated with anti-CD73 antibodies for 150 minutes in the presence of 100 µM AMP. The disappearance of AMP was measured by the modified Cell Titration Glo (CTG) assay described above and the percentage of normalized inhibition (% INH) was determined using the zero time control as 100% INH and in the control of antibody at 0% INH.
[00852] [00852] - As shown in Figure 10, anti-CD73 antibodies 350, 356, 373 and 374, in .A or .B format, inhibit membrane-bound human CD73 in a dose-dependent manner.
[00853] [00853] In addition, the enzyme inhibiting activity of anti-CD73 antibodies has also been examined with the use of human PBMCs.
[00854] [00854] “As shown in Figures 11A and 11B, the tested anti-CD73 antibodies inhibited the enzymatic activity of CD73 expressed in primary human PBMCs from both donors. Restoration of CD4 + and CD8 + T cell proliferation in the presence of AMP
[00855] [00855] Next, anti-CD73 antibodies were tested for their ability to alleviate AMP-mediated inhibition of CD4 + T cells. Briefly, CD4 + T cells were isolated from Peripheral Blood Mononuclear cells (PBMC) grouped from a healthy human donor. Prior to stimulation with anti-CD3 / 28 microspheres in the presence of 800 µM AMP, CD4 + T cells were stained with CellTrace Violet (CTV) (Thermo Fisher Scientific, Cat # C34557) to track cell division. On day 4, proliferation was determined by diluting CTV using flow cytometry. CTV-stained cells lose approximately half of their fluorescence signal as measured in flow cytometry with each division. The proliferation index was calculated as a measure of the level of T cell division for each condition where 100 represents maximum proliferation and O represents absence of proliferation.
[00856] [00856] “As shown in Figures 12A and 12B, all tested anti-CD73 antibodies were able to restore CD4 + T cell proliferation in the presence of AMP.
[00857] [00857] In a separate study, both CD4 + and T cells from
[00858] [00858] Furthermore, anti-CD73 antibodies have been examined for their enzyme inhibiting activity in vivo. Female Athymic nude mice (6 to 8 weeks old) were implanted with a CD73 (ATCC HTB-26) high-expression MDA-MB231 breast cancer cell line at 10x10º cells / mouse / 200 ul. Five mice per group were randomized when the tumors were 200 mm * and treated intraperitoneally with 20 or 200 µg / mouse of polyclonal human control IgG or a panel of anti-CD73 mAbs. The tested antibodies are the anti-CD73 antibodies 350, 356, 373 and 374, expressed in .A or .B format.
[00859] [00859] Plasma was collected three two post-doses at a ratio of one portion of plasma to five portions of methanol by volume. Samples cooled in methanol were stored at - 80 "C prior to use, at which time the samples were centrifuged. Precipitations were discarded and supernatants were transferred to new Eppendorf tubes. Standard stock solutions (IS, adenosine identified with C-13 and Inosine identified with N-15, Cambridge Isotope Laboratories, MA) were added to the final concentration of 50 nM. The prepared samples were then analyzed using a API-6500 QTrap LC / MS system (AB Sciex, US) coupled to a Shimadzu LC pump (LC-20AD) and a CTC self-sampler with DLW wash. For each sample, 5 uL were injected and separated with the use of a SeQuant ZIC-pHILIC column (sum, 150 x 2.1 mm, Millipore, MA) maintained at 40 º C. A binary gradient was used for the elution, in which the mobile phase B is 100% acetonitrile without additives and the mobile phase A is 12 mM ammonium formate and 12 mM formic acid in 1: 1 (v / v) mixture of water and acetonitrile. Elution was programmed as (0, 85, 0.6), (0.5, 85, 0.4), (2, 10, 0.4), (4.5, 10, 0.4), ( 5, 85, 0.4), (5.5, 85, 0.6), where the values in parentheses are time in minutes, percentage of mobile phase B and flow rate in mL / min in order. Adenosine and C13-adenosine were monitored from 0.5 to 4.5 minutes in positive ESI mode and mass transitions 268-> 136 and 273-> 136, respectively. Inosine and N15-Inosine were monitored from 0.5 to 4.5 minutes in a negative ESI fashion and mass transitions 267-> 135 and 271-> 139 respectively. The results were reported as nM of adenosine or inosine.
[00860] [00860] All tested anti-CD73 antibodies effectively reduced the accumulation of adenosine and inosine in the serum of immunocompromised mice implanted with the CD73 high-expression MDA-MB231 breast cancer cell line (Figure 13) . Example 2: Determination of the epitope and mode of binding of anti-CD73 antibodies
[00861] [00861] In this example, the epitope and mode of binding of anti-CD73 antibodies 350.A2, 350.B, 373.A and 373.B were determined by hydrogenation / fragmentation amide deuterium (HDx) exchange and size exclusion chromatography (SEC). Methods Hydrogen-Deuterium Fragmentation Mass Spectrometry (HDx-MS)
[00862] [00862] The HDx-MS experiments were performed as described previously (Park IH, et al., J. Chem. Inf. Model .; 55 (9): 1914 to 1925 (2015); Chalmers MJ, et al ., Anal. Chem .; 78 (4): 1005-14 (2006)). Antibody-antigen complexes were prepared and used at a 1: 1 molar ratio by incubating overnight at 4 ° C. It is important to note that this means that there is a 2-fold excess of Fab binding sites present in complex mixtures.
[00863] [00863] The exchange experiments at room temperature were carried out by manually adding 50 µl of buffer in exchange (50 mM phosphate, 150 mM NaCl, pH 7.1, in D20) at 10 µl of 0.5 mg / ml of rhCD73 protein (R&D systems, Catalog No. 5795, Trp27-Lys547 of AAH65937 fused to a 6-His C-terminal tag (SEQ ID NO: 922)) or an amount of molar equivalent of CD73-mAb complex ( 50mM phosphate, 150mM NaCl, pH 7.1). The samples were cooled after 60 s of exchange at room temperature by adding 250 μl of cooling buffer (4M guanidinium hydrochloride, 0.5M Tris (2-carboxyethyl) phosphine hydrochloride (TCEP-HCI), 0.2M phosphate, pH 2.5, 0 ºC) and after 30 s additionally diluted with 300 HL of cold storage buffer (20% glycerol, 0.25% formic acid in water) before instant freezing with nitro - liquid genius and storage at - 80 “C until use or transfer to a drawer at - 70 ºC fixed to the rail of a liquid handle (PAL HTS, LEAP Technologies, Carrboro, NC) located inside a compartment at O ºC. The frozen samples were condensed for 120 s by flowing N> gas over the sample vial (facilitated by the liquid knob together with a condensing accessory) and loaded into the sample loop of the injection valve (Valco, Houston, TX). Loading samples (load buffer 0.05% Trifluoroacetic acid (TFA), 500uL / min) in the column of the chromatographic system and digesting pepsin online (pepsin immobilized in Poros AL20, 2.1x150 mm, maintained at 15 ºC) was carried out with a double boma system (2x, Accela 1250, Thermo Scientific, Waltham MA) that allowed mixing by adding a second flow of 550uL / min (0.05% TFA, 500uL / min) of loading buffer after digestion using a mixing valve. The combined flow was directed to the chromatographic system maintained at O ºC to desalinate and the separation of gradient LC at a flow rate of 15uL / min (Dionex Ulti- Mate 3000, Thermo Scientific, Waltham, MA) and spectrometric analysis concurrent mass (QExactive, Thermo Scientific, Waltham, MA) of the chromatographic effluent. The chromatographic system consisted of a valve (15kPSI Valco, Houston, TX), a 4uL EXP Halo C18 inverted phase capture cartridge (Optimize Technologies Inc., Oregon City, OR), and an analytical column (2.1x10mm ID, Prozap 1.5um C18, Grace). Gradient separation was 0% to 40% B over 20 min followed by 40% to 75% B over 5 min using buffer compositions A: 99.75: 0.25% v / v (H20: formic acid) and B: 99.75: 0.25% v / v (acetonitrile: formic acid).
[00864] [00864] MS scans were acquired at a resolution of
[00865] [00865] Peptide identification was performed by converting raw data into .mgf format using Proteome Discoverer 1,4, researched against the construct sequence using MAS-COT 2.4 (Matrix Science, London, UK Kingdom) and filtered using Scaffold 1,4. The filtered results were imported into HDE-xaminar (v1.3, Sierra Analytics, Modesto, CA) along with the raw data files for deuteration quantification. The deceleration values were exported to Microsoft Excel to calculate deuteration differences, and normalization of deuterium incorporation by the number of observable amides (number of residues minus 2, further subtract the number of prolines in the sequence excluding the end - tion N and the penultimate residues) in a peptide. For the compression of peptide difference data for the sequence, the normalized difference deuteration values were averaged for each primary sequence residue through the peptide observations in which the respective amide was observable.
[00866] [00866] Water, deuterium oxide, guanidine hydrochloride, sodium chloride, glycerol, formic acid, trifluoroacetic acid (TFA), trifluoroethanol (TFE), acetonitrile (ACN) were from Sigma Chemical Company (St. Louis, MO ). Tris- (2-carboxyethyl) phosphine) TCEP was from Gold Biotechnology Inc (St. Louis, MO). Size Exclusion Chromatography (SEC)
[00867] [00867] An equimolar amount of rhCD73 (3.2UL, 1.6mg / mL) was mixed with mAb (2.6UL, 5mg / mL) and could form a complex overnight at 4 ºC. The samples were run using an Agilentent Autosampler connected to an Agilent 1200 series pump with an Agilent UV detector. The system was controlled by Agilent Chemstation software. The column used was the Sho- dex Protein KW-803 column (8 x 300 mm ID). Mobile Phase: 90% 2x PBS, 10% isopropanol by volume. Flow rate: 500uL / min. injection volume: 8UL. Detection wavelength: 220nm. Cycle time: 30min.
[00868] [00868] Antibody-antigen complexes were prepared and used at a ratio of 1: 1 with the use of concentration values supplied by incubation overnight at 4 ºC. It is important to note that this means that there is a nominally 2-fold excess of Fab binding sites in complex mixtures. Structure data
[00869] [00869] The structural data used for interpretation were retrieved from the Protein Database (www.rcesb.org) and consist of entries 4H2F, 4H2! | and 4H1S. The alignments of the various models and visualization were performed using PyYMOL (PyMOL Molecular Graphical System, Version 1.8 Schródinger, LLC). Results Hydrogen exchange data
[00870] [00870] The HDx MS protection profiles of 373.A and 373.B as shown in Figure 14 overlap the total sequence of CD73. This demonstrates the equivalence of antibodies in terms of their epitope.
[00871] [00871] The HDx MS protection profiles of 350.A2 and 350.B as shown in Figure 15 are in excellent compliance with each other throughout the total sequence of CD73. This demonstrates the equivalence of antibodies in terms of their epitope.
[00872] [00872] The sequences in which the strongest protection is observed are noted in the Figures and mainly correspond to CD73 loop regions. Its relative spatial arrangement is represented in the structural models shown in Figures 16A and 16B. While the protection observed for loop sequences A, B and D is attributable to the interactions of antibody-CD73, the protective effects observed for the sequence annotated as loop C are likely to arise from conformational locking. Loop C is folded in the open / inactive conformation of CD73 and unfolded in the closed / active conformation (Figure 16A). Soluble as well as membrane-bound CD73, it forms a dimer across the C-terminal domains (Figure 16B). There is a balance between open and closed conformations, since the N-terminal domain rotates by about 90 degrees to convert from open to closed. Loop C reports this balance again. If, by binding the antibody to CD73, the balance is shifted more towards the open conformation, a larger fraction of the C loops will be folded, which is a protected state. On the other hand, if the equilibrium is shifted more towards the closed conformation in the antibody binding to CD73, then a deprotective effect is expected. The unusually strong protection observed for 373.A and 373.B in loop C (Figure 14) suggests that they are not the most efficient for keeping CD73 in inactive conformation and / or doing it by a different mechanism .
[00873] [00873] From the profiles, it is inferred that the maximally protected sequences are the residue range 158 to 172, YLPYKVL- PVGDEVVG (SEQ ID NO: 108) assigned with loop A, residue range 206 to 215 , KLKTLNVNKI (SEQ ID NO: 109) assigned as loop B and residue range 368 to 387, MINNNLRHADETFWNHVSMC (SEQ ID NO: 110) assigned as loop C in all Figures, numbered according to SEQ ID NO: 105.
[00874] [00874] For antibodies 373.A and 373.B, residues in loop regions A and B appear to be equally important for the epitope as indicated by comparable amounts of protection seen in Figure 14. In the case of antibodies 350. A2 and 350.B, mainly loop A residues seem important for the epitope, even some loop B protection is observed, however, it is not clear whether this should be interpreted as direct coupling of the antibody with residues in such a band or is only an indirect effect due to the proximity of loop A. Size Exclusion Data
[00875] [00875] The size exclusion data correlates well with the HDx data discussed above. Antibody equivalence
[00876] [00876] The resources close to 17 minutes correspond to the CD73 dimer and free antibody in accordance with the molecular weight scale based on the calibrators, which are shown for reference. The resource at about 14.8min or about 300kDa is consistent with a CD73 / mAb dimer complex. Close to 12.4min, a feature of approximately 600kDa consistent with a composition of two dimers of CD73 + 2 mAbs (dimer of 2xCD73 / 2xmAb) is observed. Additional resources above 600kDa are affected by the column's exclusion limit. However, they can be attributed to higher oligomeric states that mean complexes that consist of 3 or more CD73 dimers in addition to 3 or more mAbs (3 * xCD73 / 3 * xmAb dimer).
[00877] [00877] The dominant species in the profiles of CD73-373.A and CD73-
[00878] [00878] The SEC profiles for CD73-350.A2 and CD73-350.B show
[00879] [00879] The percentages of anti-CD73 antibodies that formed a 1xCD73 / 1xmAb dimer complex, a 2xCD73 / 2xmAb dimer complex, or a higher oligomer are estimated by integrating SEC profiles in relation to distinguishable species ( Figure 19). A zero baseline was used for all integrations with integration intervals from 10.5min to 11.65min for higher oligomers and 11.65min to 13.5min for the 2XxCD73 / 2xmAb dimer complex consistently used for all samples. The integration intervals for the 1xCD73 / 1xmAb dimer complex species varied slightly and were chosen as 13.5min to 16.2min for CD73-373.A, 13.5min to 16min for CD73-
[00880] [00880] The HDxe SEC data above provides convincing evidence for the equivalence of 373.A and 373.B, as well as 350.A2 and
[00881] [00881] The data from the HDx and SEC experiments are consistent and fully support the model shown in Figure 20. According to the model, the epitopes for all antibodies are in the N-terminal domains. Antibodies 373.A and 373.B interact with the CD73 diameter to form almost exclusively the homogeneous species shown on the left of Figure 20. In such a species, both active sites of the CD73 dimer are blocked in the inactive conformation. . Furthermore, in this conformation, both N-terminal domains of the CD73 dimer are directed to the opposite side to the cell surface in the case of membrane-bound CD73, allowing 373.A and 373.B to inhibit soluble and membrane-bound versions of the target with equal effectiveness.
[00882] [00882] Antibodies 350.A2 and 350.B can form the homogeneous bidentate complex on the left, however, they do not show preference for such a shape through the 2xCD73 / 2xmAb dimer species shown in the middle of Figure 20. The oligomeric species superior figures shown on the right of Figure 20 are also observed. Presumably, the species distribution will shift towards the homogeneous bidentate complex in the case of membrane-bound CD73 or in low concentrations of antibody and antigen, which is the probable situation in vivo. Antibodies 350.A2 and 350.B are expected to be able to effectively inhibit all forms of CD73 by the inactive site locking mechanism. It is understood that, less in active formation, the target could still be catalytically inactive due to antibody binding due to other allosteric inhibition mechanisms.
[00883] [00883] “Based on the model, it would be expected that an anti-CD73 antibody that preferably forms oligomeric species illustrated in the middle and to the right of Figure 20 would be less effective in inhibiting soluble and membrane-bound CD73, especially at low concentrations. - tractions. The reason for this is that a large fraction of CD73 will always be in the active conformation and, although the antibody may inhibit catalysis under such circumstances by an indirect mechanism, any separation of the antibody will immediately produce catalytically active CD73. Furthermore, as the total antigen and antibody concentration goes towards zero, two antibody molecules will be needed to inhibit a single CD73 dimer, whereas for the e 350 antibodies 373 and 350, a single molecule would still be sufficient . summary
[00884] [00884] The putative binding sites of antibodies 373.A, 373.B,
[00885] [00885] The expression of CD73 in cancer has been reported, not only in cancer cells, but also in stromal elements that make up the tumor microenvironment. In addition, several stromal factors, including TGFB, have been suggested to provide potential amplification mechanisms for the generation of adenosine in the microambi-
[00886] [00886] For this purpose, a humanized IgG2 monoclonal antibody that binds to human and murine TGFB1 and TGFB2 with high affinity and to TGFB3 with lower affinity was used. It has been shown that this antibody has neutralizing activity against the TGFRB isoforms in vitro and in vivo. Female mice, BALB / c, immunocompetent (6 to 8 weeks of age) were implanted in the right flank with 4T1 triple murine negative breast cancer cell line at 10x10º cells / mouse / 100 ul. Ten mice per group were treated intraperitoneally on intercalated days with a TGFB pan blocking antibody or isotype control (hlgG2), both at a final concentration of 10 mg / kg body weight. The tumors were able to grow for 14 days, after which animals were euthanized and the tumors collected and subjected to flow cytometric analysis. Briefly, the tumors were removed from mice and digested using a combination of physical and enzymatic dissociation (DNasel, CollagenaseP, Dispase). Single cell suspensions were Fc-blocked for 30 minutes to prevent nonspecific antibody binding, and stained with an antibody panel that binds to specific cell surface markers including CD45 (to exclude hematopoietic cells), CD31 ( endothelial cells, CD90 (pan mesenchymal marker), CD26 and podoplanin, to allow the identification of stromal cell populations, CD73 expression was evaluated using an eFluor450 conjugated anti-CD73 antibody (eBioscience n 48-0731- 82 ), compared to the stained samples of controlled isotype (I9G1 of Mouse conjugated to eFluor 450, eBioscience nº 48-4301-82). After staining for 40 minutes at 4ºC, the samples were washed
[00887] [00887] To assess whether the CD73-dependent adenosine generation block can specifically inhibit class switching recombination (CSR) without impacting the effectiveness of B cell response, the anti-CD73 350.B antibody was tested. Virgin B cells (CD19 + CD27-IgM + IgD +) were purified from peripheral blood from two healthy donors, based on CD73 expression, and stimulated with a TLR9 agonist, an anti-CD40 antibody and cyclic tocins (IL-2 and IL-21) in the presence of antibody 350.B. Three different concentrations of the antibody were tested - 1 ng / ml, 10 ng / ml and 100 ng / ml - along with an IgG4 isotype control. IgM secretion was measured on day 7 in culture supernatants, and no difference was observed compared to the isotype control (Figure 22A). Notably, the inhibition of CD73 enzyme activity with a 350.B treatment resulted in reduced differentiation of IgG secretion cells, as measured by an ELISPOT assay to detect IgG secretion plasma cells (Millipore MSIPSA4510 plates, coated with 10 pg / ml of goat anti-human IgG, Southern Biotechnologies), as a strong effect on the concentration of 100 ng / ml (Figure 22C). The number of viable cells was also assessed and no difference was observed (Figure 22B), except for the link between low survival rate and unsatisfactory CSR efficacy observed in CD73 + virgin B cells after treatment with 350.B. As a control, the virgin CD73 cells were similarly stimulated, and no difference was observed (Figures 22A to 22C). Finally, the CSR evaluation under isotype control conditions confirmed the increased number of IgG ISCs in the virgin CD73 + compartment (Figure 22C). Taken together, these results demonstrated that blocking the enzymatic activity of CD73 has an impact on class switching recombination without changing the IgM response.
[00888] [00888] To determine the role of the co-blocking of the CD73 and PD-1 trajectories in mice carrying the tumor, anti-PD-1 and anti-CD73 antibodies (350.B) were administered to the immunocompetent mice inoculated with a cell line of syngeneic colon carcinoma, CT-26. BALB / c mice 6 to 10 weeks old, each, were inoculated in the flank with 1 x 10º of CT-26 tumor cells. On day 2 after inoculation, mice (n = 10 / group) were treated intraperitoneally with anti-
[00889] [00889] Treatment with 350.B or low-dose anti-PD-1 alone (300 µg) showed partial tumor growth inhibition (30.96% and 52.04%, respectively) on day 25. In contrast, low-dose anti-PD-1 + 350.B in combination resulted in 88.41% tumor growth inhibition (Figure 23). Tumor control with a high dose of anti-PD-1 alone was sufficient, so that the additional benefit with 350.B has not been observed. No notable loss in body weights was observed in any of the groups, and there were no significant clinical observations for the animals in the study. These data demonstrate that the combination treatment of subideal doses of anti-CD73 and anti-PD-1 antibodies can lead to enhanced tumor growth inhibition in the CT-26 synegene tumor model. Example 6: The pharmacokinetics and toxicology study of anti-CD73 373.A antibody in cynomolgus monkeys
[00890] [00890] “As a single agent, anti-CD73 373.A antibody was well tolerated when administered intravenously once weekly over four weeks to cynomolgus monkeys in doses as high as 100 mg / kg. There were no effects on cardiac function as assessed by the electrocardiogram, nor were there any toxicologically significant effects on findings of clinical pathology, immunophenotyping or histopathology. Therefore, the level of unobserved adverse effect (NOAEL) for 373.A in cyano-moss monkeys is 100 mg / kg, while the highest non-severely toxic dose (HNSTD) is> 100 mg / kg.
[00891] [00891] In a single-dose non-LPG toxicology study, cynomolgus macaques were administered with single | V control doses, 3, 30 or 100 mg / kg of antibody 373.A. After IV injection, 373.A suffers a rapid decline in the 3 mg / kg group probably due to the target-mediated drug disposition. In the 30 mg / kg and 100 mg / kg groups, the decline was generally in a multi-exponential way. No gender difference in Cmax and AUCo values at 168hrs of 373.A was observed. The exposure, assessed by 373.A means that the Cmax and AUC values at 168hr, increased with increasing dose levels from 3 to 100 mg / kg. The increase in 373.A means that the Cmax and AUCo values at 168nr were generally proportional to the dose, with the exception of AUCo at 168; between such levels of 3 and 30 mg / kg. In the 100 mg / kg dose cohort, Cmax and AUCo at 168 nm averaged 2540 upg / mL and 173,000 ug.hr/mL, respectively. The incidence of anti-drug antibody (ADA) in this study did not affect PK.
[00892] [00892] In a LPG toxicology study, monkeys were administered four weekly IV doses of control, 10, 30 or 100 mg / kg of 373.A. The sex differences in 373.A Cmax and AUC0o-168hr were minimal (between 0.7 to 1.2 times). The exposure, as assessed by Cmax and AUCo at 168hr average, increased proportionally from 10 to 100 mg / kg. After the IV bolus injection, PK of 373.A suffered a decline in a biexoponential manner, with an average elimination phase t12 value of 266 hours after the recovery phase on Day 22 in the 100 mg / kg group. In the 100 mg / kg dose cohort, Cmax and AUCo at 1681 averages after the day 22 dose, 5030 ug / mL and 365,000 ug.hr/mL, respectively. The reduced exposure of ADA incidence in an animal in the 30 mg / kg group.
[00893] [00893] In a separate study, cytokine release was evaluated in a soluble in vitro assay format after a 24-hour stimulation of human whole blood with 373.A, at 50, 100, 500 and 1000 upg / mL . Blood samples were collected from ten healthy human donors. For each sample, negative (whole blood and whole blood with medium) and positive (anti-human CD3) controls were included. Cytokine levels were measured using a Luminex multiplex method for IL-16, IL-2, IL-6, IL-8, 11-10, IFNy and TNFa. The results obtained with the buffer control were similar to the results obtained with the negative control (blood only) and with no tendency for a decrease or an increase in the cytokine release, it was noted with the diluent (0.5% Dextrose). For all donors, cytokine release was induced with positive anti-CD3 controls for all cytokines, indicating that the stimulus was adequate and cytokine release could be induced in all samples. The cytokine levels obtained after the 373.A stimulus were similar to the levels measured in the negative controls. In conclusion, the in-vitro stimulation of human whole blood with 373.A did not induce the release of cytokine in concentrations as high as
[00894] [00894] “During the single decade, immunotherapies that target different checkpoints one (for example, PD-1, PD-L1 and CTLA-4) show effectiveness in a number of cancer indications. However, although some patients achieve the goal and long-lasting responses to checkpoint block, most patients show modest or no clinical benefit, indicating that tumors use alternating immunosuppressive mechanisms to achieve the immunoescape (Allard et al ., Clin Câncer Res. 2013; 19 (20): 5626- 35; Vesely et al., Annu Rev Immunol 2011; 29: 235 to 271). Thus, concomitant blockade of multiple immunosuppressive trajectories may be necessary to induce clinically significant responses in a greater number of patients.
[00895] [00895] Over the past few years, the generation of adenosine and signaling have emerged as potential therapeutic targets in the treatment of cancer. Adenosine creates an immunosuppressive tumor microenvironment by reducing the cytotoxic anti-tumor immunoresponse, intensifying the proliferation and polarization of immunosuppressive cells, and increasing neovascularization (Young et al., Cancer Discovery 2014; 4 (8): 879: 879 -88). Preclinical data demonstrate that blocking CD73 can significantly slow the growth of primary tumor and inhibit the development of lung metastases in an immunocompetent singene mouse model (Stagg et al 2010). Similar results were observed in a study in which the A2aR genetic deletion in the host resulted in rejection of immunogenic tumors established in A2aR deficient mice without rejection seen in wild-type control mice (Ohta et al., PNAS 2006; 103 (35) : 13132-37).
[00896] [00896] A phase I / Ib, open identification, multicentralized study was designed to assess the safety, tolerability, preliminary antitumor activity, pharmacokinetics (PK) and pharmacodynamics (PD) of the anti-CD73 373.A antibody as an agent unique and in combination with the A2aaR antagonist, PBF509 and / or the anti-PD-1 antibody, BAPO49-Clone-E, in patients with advanced malignancies. Primary objectives should characterize safety and tolerability, and determine the recommended dose (RD) for 373.A as a single agent and in combination with PBF509 and / or BAPO49-Clone-E. Secondary objectives should evaluate the preliminary antitumor activity and PK of 373.A as a single agent and in combination with PBF509 and / or BAPO49-Clone-E, evaluate the immunogenicity of 373.A and BAPO049- Clone-E and characterize changes in immunoinfiltrate in tumors after treatment, for example, alteration of the baseline in tumor infiltrating lymphocytes (TIlLs), tumor-associated macrophages (TAMs), expression of CD8 + T-cells and PDL-1.
[00897] [00897] “BAPO49-Clone-E is a programmed humanized anti-killing IgG4 antibody (PD-1) of high affinity ligand block that blocks the binding of PD-L1 and PD-L2 to PD-1. BAPO49-Clone-E is tested in a phase I / Il study on advanced malignancies. PBF509, a new non-xanthine-based compound, is a potent oral adenosine A2asR antagonist. The BAPO49-Clone-E sequence is disclosed in Table 5.
[00898] [00898] Two ongoing studies of Phase I / lb and Phase | evaluate PBF509 as a single agent and / or in combination with BAPO049-Clone-E in patients with advanced non-small cell lung cancer (NSCLC) and solid tumors and non-Hodgkin lymphoma, respectively.
[00899] [00899] This I / lb study will initially enroll adult patients with advanced malignancies who have progressed or are intolerant to standard therapy in indications where moderate to high CD73 expression has been associated with the most unsatisfactory result, indicating mediated immunoescape by adenosine (Wu et al., Journal of Social Oncology 2012, 106 (2): 130 to 137; Gaudreau et al., Oncoimmunology; 2016, 5 (5): e1127496; Inoue et al, Oncotarget .; 2017, 8 (5): 8738-8751). These indications include non-small cell lung cancer (NSCLC), triple negative breast cancer (TNBC),
[00900] [00900] The study consists of two parts: (1) one part of the dose scale for single agent 373.A, duplicate combinations
[00901] [00901] In the expansion part, the patients in each indication will be equally randomized to the combination treatment arms. Randomization will be performed by indication, and additionally stratified in certain indications by treatment of previous PD-1 / PD-L1 (virgin or resistant). Dose selection and regimen
[00902] [00902] The initial dose of 60 mg of 373.A flat dose, administered intravenously every 2 weeks (Q2W), was selected based on preclinical safety, tolerability and PK data observed in cynomolgus monkeys as well as the published case history of CD73-deficient patients.
[00903] [00903] The dose of 60 mg is considered a minimally active pharmacological dose (mMPAD), as it is expected to provide (1) approximately 20 h of> 90% T cell CD8 + CD73 occupancy, (2) approximately 22 h of> 90% inhibition of adenosine, (3) approximately 17 h of> 90% occupancy of overall imputed CD73.
[00904] [00904] “Based on the TK data modeling of the cynomolgus Toxicology studies, ex-vivo CD8 + CD73 occupancy data and in-vitro data on the inhibition of adenosine formation, a 21200 mg dose of Q2W is expected to achieve> 90% target occupancy in CD8 + T cells over the dosing interval and a 2600 mg Q2W dose is expected to achieve> 90% inhibition of adenosine production.
[00905] [00905] The dose of 373.A will be scaled in sequential cohorts, guided by a Bayesian Logistic Regression (BLRM) model coupled with overdose control criteria (EWOC), until a maximum tolerated dose (MTD), or dose recommended (RD) for expansion, be identified. Pre-clinical data and modeling suggest that high antigen collection may be required and that high doses (eg, 21200 mg Q2W) may be required to achieve continuous target occupancy over the dose range. Dose escalation will be performed primarily with a Q2W regimen. However, if this regimen shows an elimination of 373.A rapid and lack of target saturation within the dosing range, a more frequent QW regimen can be tested. If, on the other hand, a Q4W regimen is not expected to have rapid elimination within the dosing interval, the Q4W regimen can be explored, instead. 373.A / PBF509 combination
[00906] [00906] Maximum initial doses for the doublet combination of
[00907] [00907] 200 mg of Q2W 373.A is a low dose of 373.A which is expected to reach —2.3 days of> 90% target occupancy in CD8 + T cells. The 200 mg Q2W dose of 373.A is 16% of the 1200 mg Q2W dose which is expected to reach> 90% target occupancy of CD8 + T cell over the dosing interval.
[00908] [00908] “PBF509 was tested as a single agent up to 640 mg of BID without safety problems in advanced / metastatic NSCLC (only 1 DLT out of 5 patients evaluable at a dose of 640 mg of BID). In the same study, PBF509 showed single agent activity ranging from 80 mg BID to 480 mg BID, with 2 partial responses (PRs) and 6 with stable disease (DS), in a total of 18 dosed patients.
[00909] [00909] “A dose escalation approach of 373.A and PBF509 will be initiated to determine the appropriate dose of each drug in combination, guided by Bayesian Logistic Regression (BLRM) modeling coupled with overdose control principle criteria (EWOC). 373.A / BAPO49-Clone-E combination
[00910] [00910] The maximum starting dose for the doublet combination
[00911] [00911] The justification for 200 mg of Q2W of 373.A has been described above. The 373.A at 200 mg of Q2W will be combined with the RD for BAPO49-Clone-E which is 400 mg of Q4W, which has been shown to be safe and effective.
[00912] [00912] The dose level of 373.A will be scaled sequentially with a fixed dose of BAPO49-Clone-E, guided by Bayesian Logistic Regression (BLRM) modeling coupled with the overdose control principle criteria ( EWOC). 373.A / BAPO49-Clone-E / PBF509 combination
[00913] [00913] The maximum starting dose for the triple combination of
[00914] [00914] “PBF509 was safely administered up to 160 mg of BID in combination with 400 mg of Q4W of BAPO49-Clone-E (only 1 DLT out of 6 evaluable patients), with combining activity in effectiveness: 2 PRs and 6 SDs, among a total of 10 patients. The 160 mg dose of PBF509 BID and 400 mg Q4W of BAPO49-Clone-E are currently being used in the Ph2 study.
[00915] [00915] A dose escalation approach for
[00916] [00916] Antibody 373.A (100 mg powder for solution for infusion) will be administered intravenously as an infusion of 1 h (up to 2 hours if clinically indicated). BAPO49-Clone-E (100 mg powder for solution for infusion) will be administered intravenously as a 30-minute infusion (up to 2 hours, if clinically indicated). When supplied in combination, 373.A and BAPO49-Clone-E must be administered on the same day using separate infusion materials (bag, lines, filters) for each infusion. The same access site can be used for both infusions. 373.A will undergo the infusion first followed by a 30-minute pause before the BAPO49-Clone-E infusion. PBF509 (40 mg and / or 80 mg and / or 160 mg capsule for oral use) will be taken orally twice daily (BID) continuously. In visits where
[00917] [00917] Tables 22 to 25 describe the starting dose and dose levels that can be assessed during this test. Patients treated with a single agent of 373.A or 373.A in combination with BAPO049- Clone-E and / or PBF5089 will begin study treatment on Day 1 of Cycle 1. Each cycle will consist of 28 days. 373. Q2W will be administered on Days 1 and 15 of a cycle. 373. QW will be administered on Days 1, 8, 15 and 22 of a cycle. 373.The Q4W will be administered on Day 1 of a cycle. BAPO49-Clone-E Q4W will be administered on Day 1 of a cycle. PBF509 IDB will be taken every day of a cycle. Table 22. Provisional dose levels for 373.A Dose level Dosage regimen Increase from the proposed dose * previous 2 "6 mg Q2W -233% A” 20 mg Q2W -200% 1 60 mg Q2W (starting dose) 2 200 mg Q2W 233% 3 600 mg Q2W 200% 4 1200 mg Q2W 100% 2400 mg Q2W 100% 6 3600 mg Q2W 50% * Additional and / or intermediate dose levels may be added during the course of the study. can be added at any dose level below BAT to better understand safety, PK or PD. ** Dose level -1 and -2 represent treatment doses for patients who need a dose reduction from the initial dose level. No dose reduction below dose level -2 is allowed for this study.
[00918] [00918] The Fab fragment of 350.A2 was generated by genetically modifying a stop codon between the two proline residues above the core articulation region of the 350.A2 antibody heavy chain. 350.A2 Fab comprises the heavy chain comprising the amino acid sequence of SEQ ID NO: 331 and a light chain comprising the amino acid sequence of SEQ ID NO: 23. The CD73 ECD with his-tag (SEQ ID NO: 171) was cloned into the pRS5a vector. Expression and Purification
[00919] [00919] Expi293F cells were transfected at 1: 1 ratio of 350.A2 to CD73 ECD and five days post-transfection, cells were pelleted at 2000 rpm. During that time, the
[00920] [00920] The 350.A2 Fab-CD73 complex was crystallized using the suspension-drop method. Before configuring the crystallization trays, the purified complex was spun at 14,000 rpm for 10 minutes to remove protein and aggregate debris. Then, 1.0 μl of the 9 mg / ml complex was mixed with 1.0 μl protein precipitant consisting of 0.2M Li2SOs, 1.2M NaH2PO.a, 0.8M K2HPO. , 0.1M Glycine, pH 10.5 and then suspended over a 300 µl protein precipitant well. The crystals grew in approximately 3 to 4 days. The crystals were collected and then cryopreserved using a protein precipitant solution combined with 25% (v: v) glycerol. The X-ray diffraction data were collected in Advanced Light Source, beam line 5.0.1. Diffraction data was usable at approximately 2.85 Angstroms. Structure and Refinement Solution
[00921] [00921] The 350.A2 structure was modeled using the AL-57 Fab fragment (RCSB accession code 3HI5) as a model using the MOE modeling package. The co-structure was dissolved using the PHASER program and the dimeric structure of human glycosylated CD73 (RCSB 4H18S) and the Fab homology model as research models. Briefly, the dimeric structure of human CD73 was used as the first research model, locating the position of two dimers of human CD73 in the asymmetric unit. This initial structure was refined using Phenix-Refine. Subsequently, the heavy and light chain homology model was used as a separate research model, thus locating the position of the two Fabs in relation to the two human CD73 dimers. The completed structure was refined to convergence through multiple cycles of refinement and reconstruction of real space. The final R-factor and free R factor were 22.78% and 27.12%, respectively. The electron density maps reveal that each CD73 monomer is glycosylated in asparagine-311. However, this glycosylation is heterogeneous and, therefore, it is not possible to determine the exact chemical structure of the glycosylation event. Because of this, glycosylation is not included in the structural model.
[00922] [00922] The cocrystal structure of 350.A2 Fab-CD73 reveals the interactions of the antibody with CD73. These interactions can be divided into direct enthalpic interactions, such as hydrogen bonding or electrostatic interactions, and hydrophobic and Van der Waal interactions that are conducted by format complementarity between the antibody and CD73.
[00923] [00923] The enthalpy and VDW / hydrophobic key interactions identified by the cocrystal structure are as follows. The following interactions occur between the Fab heavy chain and CD73. The Fab heavy chain residues are numbered according to the Kabat numbering and highlighted in Figures 26C and 26D (bold, italic and underlined with a line) . CD73 residues are numbered according to SEQ ID NO: 105 and highlighted in Figures 26A and 26B (bold, italic and underlined with a line).
[00924] [00924] The following interactions occur between the Fab light chain and CD73. Fab light chain residues are numbered according to Kabat numbering and highlighted in Figures 26G and 26H (bold, italic and underlined with a line). CD73 residues are numbered according to SEQ ID NO: 105 and highlighted in Figures 26E and 26F (bold, italic and underlined with a line).
[00925] [00925] Antibody comparisons with the use of TMT isotope encoding reagents were primarily conducted as described in John D. et al., Analytical Chemistry 2015 87 (15), 7540 to 7544, incorporated herein by way of reference in its entirety. TMT identification methods
[00926] [00926] The antibodies in DPBS had the following concentrations and were used without further processing: 374 (1.37 mg / mL),
[00927] [00927] The samples were combined 15 min after cooling hydroxylamine in a rotating concentrator and the total volume was reduced to -30 uL. This was followed by deglycosylation with PNGase F and reducing / denaturing the separation of PAGE from SDS by standard methods.
[00928] [00928] The bands containing CD73 were excised from the gel for gel management. Gel processing followed standard procedures for removing staining, reduction, alkylation and dehydration. For conditioning before elastase digestion, the dehydrated gel pieces were rehydrated with 50 mM pH 8 Tris-buffer, while for pepsin digestion, 1% formic acid was used in the last rehydration step before completing vacuum dehydration.
[00929] [00929] The elastase digestion samples were rehydrated by 1 hem ice by adding 25 μl of 12.5 ng / ulL elastase in 50 mM Tris, pH 8. The pepsin digestion samples were rehydrated for 1 h in ice by adding 25 μl of 10 ng / μl of pepsin in 1% formic acid.
[00930] [00930] After rehydration with the enzymes, the excess enzyme solution was discharged and 50 ul of the respective digestion buffer added before digestion for 4 h at 37 ºC. After digestion, the peptide recovery again followed the standard gel digestion methodology. LCMSMS
[00931] [00931] LOCMSMS was performed using an Orbitrap Lumos mass spectrometer (Thermo Scientific, Waltham, MA) coupled to an Easy-nLC 1200 chromatography system (Thermo Scientific, Waltham, MA). The separation column was a 75 µm capillary with a 15 cm ReproSil-Pur 120 C18 AQ 3 µm tip pack (catalog number r13.aqg). The proteolytic fragments were eluted using a segmented gradient from O to 35% B in 120 min, 35 to 63% B in 30 min, 63 to 100% B in 5 min, 100% B lied for 5 min, 100 to 0% B in 2 min, 0% B maintained for 2 min at a flow rate of 500 nL / min using elution buffers of A = 0.1% formic acid, water and B = 0.1% formic acid in 80% MCN.
[00932] [00932] The data acquisition method used the manufacturer's predefined method parameters for a TMT experiment with MSMS detection. Data reduction and analysis
[00933] [00933] Raw data were searched against the CD73 protein sequence using MASCOT 2.5.1 (MatrixScience, United Kingdom) using MASCOT Daemon (MatrixScienc, United Kingdom) and ExtractMSn (Thermo Scientific, Waltham, MA ) for converting and merging elastase and raw pepsin files for peak .mgf lists. For quantification, the predefined method of TMT10plex embedded in MASCOT was used. Research results including TMT ion-reporter intensities were exported as .csv and further refined / cured to remove anomalies before averaging weighted intensity reporter intensities using peptide spectral matches (PSMs) for a given lysine residue from the primary sequence using Microsoft Excel. Only peptide PSMs containing a single lysine residue were used in the analysis to avoid ambiguity at the identification site. Results
[00934] [00934] Normalized CD73 ion-reporter intensity ratios of weighted intensity covering observable primary sequence lysine residues are plotted as the difference between the ratio of the CD73 / anti-CD73 mAb and CD73 complex (1 per standardization). Plotting the reason in this way allows direct assessment
[00935] [00935] Figures 27A and 27B show the difference in ratios for 29 of the 35 lysines in the primary sequence of CD73 as seen in the 30s identification. Protection on lysines K162, K206 and K214 (Figure 27A) correlates well with the previous HDX interpretation of the 373 epitope (Example 2). Specifically, the HDX study assigned residues 158 to 172 (YLPYKVLPVGDEVVG, SEQ ID NO: 108, loop A), which encompass K162; and residues 206 to 215 (KLKTLNVNKI, SEQ ID NO: 109, loop B), which comprise K206 and K214, as part of the epitome of 373. Protection on lysines K285 and K291 (Figure 27B) is consistent with the minimum protection observed in the previous HDX data in the area around residue 300 (Figure 14 and Example 2). K262 and K274 are placed on the opposite sides of the N-terminal domain in relation to the epitope (K162, K206 and K214) and the increased identification in these two residues (Figure 27A) may reflect "destabilization"
[00936] [00936] Figures 28A and 28B show the difference in ratios for 29 of the 35 lysines in the primary sequence of CD73 as seen in the 300s identification. Protection in K162 is no longer observable for mAbs 374, 377, 379 and 373, while protection is still strong for
[00937] [00937] Notably, for all these antibodies, the protection in K206 is comparable to the protection in K214 (Figures 27A and 28A). INCORPORATION BY REFERENCE
[00938] [00938] All publications, patents and Access numbers mentioned in this document are hereby incorporated by reference in their entirety as if each individual publication or patent were specifically and individually indicated as being incorporated by reference. EQUIVALENTS
[00939] [00939] “Although specific modalities of the invention in question have been discussed, the above specification is illustrative and not restrictive. Many variations of the invention will become apparent to those skilled in the art after reviewing this specification and the claims below. The total scope of the invention must be determined by reference to the claims, together with its total scope of equivalents, and the specification, together with such variations.

权利要求:
Claims (1)
[1]
1. Antibody molecule that binds to human CD73, characterized by the fact that it comprises: (i). a heavy chain variable region (VH) comprising an amino acid sequence of heavy chain complementarity determining region 1 (VHCDR1) of SEQ ID NO: 88, an amino acid sequence of VÓHCDR 2 of SEQ ID NO: 89 and a VHCDR3 amino acid sequence of SEQ ID NO: 37; and (ii); a light chain variable region (VL) comprising an amino acid sequence of light chain complementarity determining region 1 (VLCDR1) of SEQ ID NO: 48, an amino acid sequence of VLCDR 2 of SEQ ID NO: 49 and a VLCDR3 amino acid sequence of SEQ ID NO: 50.
2. Antibody molecule according to claim 1, characterized in that it comprises: (i), a VH comprising a VHCDR1 amino acid sequence of SEQ ID NO: 38, a VHCDR2 amino acid sequence of SEQ ID NO: 36 and a VHCDR3 amino acid sequence of SEQ ID NO: 37; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 48, a VLCDR2 amino acid sequence of SEQ ID NO: 49 and a VLCDR3 amino acid sequence of SEQ ID NO: 50; (i))) a VH comprising a VÓHCDR1 amino acid sequence of SEQ ID NO: 72, a VHCDR2 amino acid sequence of SEQ ID NO: 71 and a VHCDR3 amino acid sequence of SEQ ID NO: 37; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 48, a VLCDR 2 amino acid sequence of SEQ ID NO: 49 and a VLCDR3 amino acid sequence of SEQ ID NO: 50; (iii) a VH comprising an amino acid sequence
VÓHCDR1 acids of SEQ ID NO: 38, a VHCDR2 amino acid sequence of SEQ ID NO: 71 and a VHCDR3 amino acid sequence of SEQ ID NO: 37; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 48, a VLCDR2 amino acid sequence of SEQ ID NO: 49 and a VLCDR3 amino acid sequence of SEQ ID NO: 50; (iv) a VH comprising a VHCDR1 amino acid sequence of SEQ ID NO: 137, a VHCDR2 amino acid sequence of SEQ ID NO: 136 and a VHCDR3 amino acid sequence of SEQ ID NO: 37; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 48, a VLCDR2 amino acid sequence of SEQ ID NO: 49 and a VLCDR3 amino acid sequence of SEQ ID NO: 50; (v) a VH comprising a VHCDR1 amino acid sequence of SEQ ID NO: 137, a VHCDR 2 amino acid sequence of SEQ ID NO: 146 and a VHCDR3 amino acid sequence of SEQ ID NO: 37; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 48, a VLCDR2 amino acid sequence of SEQ ID NO: 49 and a VLCDR3 amino acid sequence of SEQ ID NO: 50; or (vi) a VH comprising a VHCDR1 amino acid sequence of SEQ ID NO: 137, a VHCDR2 amino acid sequence of SEQ ID NO: 154 and a VÓHCDR3 amino acid sequence of SEQ ID NO: : 37; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 48, a VLCDR2 amino acid sequence of SEQ ID NO: 49 and a VLCDR3 amino acid sequence of SEQ ID NO: 50.
Antibody molecule according to claim 1, characterized in that it comprises a VH comprising a VHCDR1 amino acid sequence of SEQ ID NO: 38, a VÓHCDR2 amino acid sequence of SEQ ID NO: 36 and a VHCDR3 amino acid sequence of SEQ ID NO: 37; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 48, a VLCDR2 amino acid sequence of SEQ ID NO: 49 and a VLCDR3 amino acid sequence of SEQ ID NO:
50.
Antibody molecule according to any one of claims 1 to 3, characterized in that it comprises a variable region of heavy chain comprising the amino acid sequence of SEQ ID NO: 44, 77, 84, 142, 151 or 159, or an amino acid sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 44, 77, 84, 142, 151, or 159.
Antibody molecule according to any one of claims 1 to 4, characterized in that it comprises a light chain variable region comprising the amino acid sequence of SEQ ID NO: 55, or an amino acid sequence which it has at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 55.
Antibody molecule according to any one of claims 1 to 5, characterized in that it comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 46, 79, 86, 114, 116 or 117, or an amino acid sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 46, 79, 86, 114, 116, or 117.
Antibody molecule according to any one of claims 1, 2, 3, 4, 5 or 6, characterized by the fact that it comprises a light chain comprising the amino acid sequence of SEQ ID NO: 57 , or an amino acid sequence that has at least about 85%, 90%, 95%, or 99% sequence identity
with SEQ ID NO: 57.
Antibody molecule according to any one of claims 1 to 7, characterized in that it comprises: (i). a heavy chain variable region that comprises the amino acid sequence of SEQ ID NO: 44 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity thereto) and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 55 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity thereto); (li) a heavy chain variable region that comprises the amino acid sequence of SEQ ID NO: 77 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 55 (or a sequence having at least about 85%, 90%, 95%, or 99% sequence identity thereto) ; (iii) a heavy chain variable region that comprises the amino acid sequence of SEQ ID NO: 84 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 55 (or a sequence having at least about 85%, 90%, 95%, or 99% sequence identity thereto) ; (iv) a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 142 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with the same) and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 55 (or a sequence having at least about 85%, 90%, 95%, or 99%
sequence identity); (v) a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 151 (or a sequence having at least about 85%, 90%, 95%, or 99% sequence identity with the same) and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 55 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity to the same); or (vi) a heavy chain variable region that comprises the amino acid sequence of SEQ ID NO: 159 (or a sequence that has at least about 85%, 90%, 95%, or 99% identity of sequence with the same) and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 55 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with the same).
Antibody molecule according to any one of claims 1 to 8 characterized by the fact that it comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 44 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 55.
Antibody molecule according to any one of claims 1 to 9, characterized in that it comprises: (i)) a heavy chain comprising the amino acid sequence of SEQ ID NO: 46 (or a sequence having at least less than 85%, 90%, 95%, or 99% sequence identity thereto) and a light chain comprising the amino acid sequence of SEQ ID NO: 57 (or a sequence having at least about 85%, 90%, 95%, or 99% sequence identity with the same); (li), a heavy chain comprising the amino acid sequence of SEQ ID NO: 114 (or a sequence having at least about 85%, 90%, 95%, or 99% sequence identity thereto) and a light chain comprising the amino acid sequence of SEQ ID NO: 57 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity therewith);
(iii) a heavy chain comprising the amino acid sequence of SEQ ID NO: 79 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity therewith) and a light chain comprising the amino acid sequence of SEQ ID NO: 57 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity to it) ;
(iv) a heavy chain comprising the amino acid sequence of SEQ ID NO: 116 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity thereto) and a chain light that comprises the amino acid sequence of SEQ ID NO: 57 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity therewith);
(v) a heavy chain comprising the amino acid sequence of SEQ ID NO: 86 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity thereto) and a light chain comprising the amino acid sequence of SEQ ID NO: 57 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity to it) ; or
(vi) a heavy chain comprising the amino acid sequence of SEQ ID NO: 117 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity thereto) and a chain lightweight which comprises the amino acid sequence of SEQ ID NO: 57 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity therewith).
Antibody molecule according to any one of claims 1 to 10, characterized in that it comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 46 and a light chain comprising the amino acid sequence of SEQ ID NO: 57.
12. Antibody molecule that binds to human CD73, characterized by the fact that it comprises: (i), a VH that comprises a VHCDR1 amino acid sequence of SEQ ID NO: 90, a VHCDR2 amino acid sequence SEQ ID NO: 91 and a VHCDR3 amino acid sequence of SEQ ID NO: 3; and (ii). a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 14, a VLCDR2 amino acid sequence of SEQ ID NO: 15 and a VLCDR3 amino acid sequence of SEQ ID NO: 16.
Antibody molecule according to claim 12, characterized in that it comprises: (i), a VH comprising a VHCDR1 amino acid sequence of SEQ ID NO: 61, a VHCDR2 amino acid sequence of SEQ ID NO: 60 and a VHCDR3 amino acid sequence of SEQ ID NO: 3; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 14, a VLCDR2 amino acid sequence of SEQ ID NO: 15 and a VLCDR3 amino acid sequence of SEQ ID NO: 16; (li) a VH comprising a VHCDR1 amino acid sequence of SEQ ID NO: 4, a VHCDR2 amino acid sequence of SEQ ID NO: 26 and a VHCDR3 amino acid sequence of SEQ ID NO: 3; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 14, a VLCDR2 amino acid sequence of SEQ ID NO: 15 and a VLCDR3 amino acid sequence of SEQ ID NO: 16; (iii) a VH comprising a VÓHCDR1 amino acid sequence of SEQ ID NO: 4, a VHCDR2 amino acid sequence of SEQ ID NO: 2 and a VHCDR3 amino acid sequence of SEQ ID NO: 3; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 14, a VLCDR2 amino acid sequence of SEQ ID NO: 15 and a VLCDR3 amino acid sequence of SEQ ID NO: 16; or (iv) a VH comprising a VÓHCDR1 amino acid sequence of SEQ ID NO: 163, a VÓHCDR2 amino acid sequence of SEQ ID NO: 162 and a VHCDR3 amino acid sequence of SEQ ID NO: 162 : 3; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 14, a VLCDR2 amino acid sequence of SEQ ID NO: 15 and a VLCDR3 amino acid sequence of SEQ ID NO: 16.
Antibody molecule according to claim 12 or 13, characterized in that it comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 66, 31, 10, or 168, or an amino acid sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 66, 31, 10, or 168.
Antibody molecule according to any one of claims 12 to 14, characterized in that it comprises a light chain variable region comprising the amino acid sequence of SEQ ID NO: 21, or an amino acid sequence that it has at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 21.
An antibody molecule according to any one of claims 12 to 15, characterized in that it comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 68, 33, 12, 115, 113, or 112, or an amino acid sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 68, 33, 12, 115, 113, or
112.
17. Antibody molecule according to any one of claims 12 to 16, characterized in that it comprises a light chain comprising the amino acid sequence of SEQ ID NO: 23, or an amino acid sequence that has at least - about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 23.
18. Antibody molecule according to any one of claims 12 to 17, characterized in that it comprises: (i). a heavy chain variable region that comprises the amino acid sequence of SEQ ID NO: 66 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity thereto) and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 21 (or a sequence having at least about 85%, 90%, 95%, or 99% sequence identity thereto); (li) a heavy chain variable region that comprises the amino acid sequence of SEQ ID NO: 31 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 21 (or a sequence having at least about 85%, 90%, 95%, or 99%
sequence identity); (iii) a heavy chain variable region that comprises the amino acid sequence of SEQ ID NO: 10 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 21 (or a sequence which has at least about 85%, 90%, 95%, or 99% sequence identity thereto) ; or (iv) a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 168 (or a sequence that has at least about 85%, 90%, 95%, or 99% identity of sequence with the same) and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 21 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with the same).
19. Antibody molecule according to any one of claims 12 to 18, characterized in that it comprises: (i). a heavy chain comprising the amino acid sequence of SEQ ID NO: 68 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity thereto) and a light chain which comprises the amino acid sequence of SEQ ID NO: 23 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity to it); (ii), a heavy chain comprising the amino acid sequence of SEQ ID NO: 115 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity thereto) and a light chain comprising the amino acid sequence of SEQ ID NO: 23 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity therewith); (iii) a heavy chain comprising the amino acid sequence of SEQ ID NO: 33 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity therewith) and a light chain comprising the amino acid sequence of SEQ ID NO: 23 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity to it) ; (iv) a heavy chain comprising the amino acid sequence of SEQ ID NO: 113 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity thereto) and a chain light comprising the amino acid sequence of SEQ ID NO: 23 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity therewith); (v) a heavy chain comprising the amino acid sequence of SEQ ID NO: 12 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity therewith) and a light chain comprising the amino acid sequence of SEQ ID NO: 23 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity to it) ; or (vi) a heavy chain comprising the amino acid sequence of SEQ ID NO: 112 (or a sequence which has at least about 85%, 90%, 95%, or 99% sequence identity thereto) and a light chain comprising the amino acid sequence of SEQ ID NO: 23 (or a sequence that has at least about 85%, 90%, 95%, or 99% sequence identity therewith).
20. Antibody molecule that binds to human CD73,
characterized by the fact that it comprises: (i), a VH comprising an amino acid sequence of VHCDR1 of SEQ ID NO: 189, an amino acid sequence of VHCDR 2 of SEQ ID NO: 89 and a sequence of amino acids VHCDR3 of SEQ ID NO: 43; and (ii) a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 48, a VLCDR2 amino acid sequence of SEQ ID NO: 49 and a VLCDR3 amino acid sequence of SEQ ID NO: 50.
21. Antibody molecule according to claim 20, characterized by the fact that it comprises: (i)) a VH that comprises a VHCDR1 amino acid sequence of SEQ ID NO: 190, an amino acid sequence of VÓMHCDR 2 of SEQ ID NO: 36 and an amino acid sequence VHCDR3 of SEQ ID NO: 43; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 48, a VLCDR2 amino acid sequence of SEQ ID NO: 49 and a VLCDR3 amino acid sequence of SEQ ID NO: 50; (i))) a VH comprising a VHCDR1 amino acid sequence of SEQ ID NO: 191, a VHCDR 2 amino acid sequence of SEQ ID NO: 71 and a VHCDR3 amino acid sequence of SEQ ID NO: 43; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 48, a VLCDR2 amino acid sequence of SEQ ID NO: 49 and a VLCDR3 amino acid sequence of SEQ ID NO: 50; (iii) a VH comprising a VHCDR1 amino acid sequence of SEQ ID NO: 192, a VOMCDR 2 amino acid sequence of SEQ ID NO: 71 and a VHCDR3 amino acid sequence of SEQ ID NO: 43; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 48, a sequence
amino acid sequence of VLCDR2 of SEQ ID NO: 49 and an amino acid sequence of VLCDR3 of SEQ ID NO: 50; (iv) a VH comprising a VHCDR1 amino acid sequence of SEQ ID NO: 193, a VHCDR 2 amino acid sequence of SEQ ID NO: 136 and a VHCDR3 amino acid sequence of SEQ ID NO: 43; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 48, a VLCDR2 amino acid sequence of SEQ ID NO: 49 and a VLCDR3 amino acid sequence of SEQ ID NO: 50; (v) a VH comprising a VÓHCDR1 amino acid sequence of SEQ ID NO: 194, a VHCDR 2 amino acid sequence of SEQ ID NO: 146 and a VHCDR3 amino acid sequence of SEQ ID NO: 43; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 48, a VLCDR 2 amino acid sequence of SEQ ID NO: 49 and a VLCDR3 amino acid sequence of SEQ ID NO: 50; or (vi) a VH comprising a VHCDR1 amino acid sequence of SEQ ID NO: 195, a VHCDR 2 amino acid sequence of SEQ ID NO: 154 and a VHCDR3 amino acid sequence of SEQ ID NO: 43 ; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 48, a VLCDR2 amino acid sequence of SEQ ID NO: 49 and a VLCDR3 amino acid sequence of SEQ ID NO: 50.
22. Antibody molecule that binds to human CD73, characterized by the fact that it comprises: (i) a VH comprising a VMHCDR1 amino acid sequence of SEQ ID NO: 196, a VHCDR2 amino acid sequence of SEQ ID NO: 91 and a VHCDR3 amino acid sequence of SEQ ID NO: 9; and (ii) a VL comprising an amino acid sequence
of VLCDR1 of SEQ ID NO: 14, an amino acid sequence of VLCDR2 of SEQ ID NO: 15 and an amino acid sequence of VLCDR3 of SEQ ID NO: 16.
23. Antibody molecule according to claim 22, characterized in that it comprises: (i) a VH comprising a VHCDR1 amino acid sequence of SEQ ID NO: 197, a VHCDR2 amino acid sequence of SEQ ID NO: 60 and a VHCDR3 amino acid sequence of SEQ ID NO: 9; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 14, a VLCDR2 amino acid sequence of SEQ ID NO: 15 and a VLCDR3 amino acid sequence of SEQ ID NO: 16; (ii) a VH comprising a VHCDR1 amino acid sequence of SEQ ID NO: 198, a VHCDR2 amino acid sequence of SEQ ID NO: 26 and a VHCDR3 amino acid sequence of SEQ ID NO: 9; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 14, a VLCDR2 amino acid sequence of SEQ ID NO: 15 and a VLCDR3 amino acid sequence of SEQ ID NO: 16; (iii) a VH comprising a VHCDR1 amino acid sequence of SEQ ID NO: 199, a VHCDR2 amino acid sequence of SEQ ID NO: 2 and a VHCDR3 amino acid sequence of SEQ ID NO: 9; and a VL comprising a VLCDR1 amino acid sequence of SEQ ID NO: 14, a VLCDR2 amino acid sequence of SEQ ID NO: 15 and a VLCDR3 amino acid sequence of SEQ ID NO: 16; or (iv) a VH comprising a VÓHCDR1 amino acid sequence of SEQ ID NO: 200, a VHCDR 2 amino acid sequence of SEQ ID NO: 162 and a VHCDR3 amino acid sequence of SEQ ID NO: 9 ; and a VL that comprises a sequence
VLCDR1 amino acid sequence of SEQ ID NO: 14, a VLCDR2 amino acid sequence of SEQ ID NO: 15 and a VLCDR3 amino acid sequence of SEQ ID NO: 16.
24. Antibody molecule according to any one of the claims | to 23, which is a human antibody, a full length antibody, a bispecific antibody, Fab, F (ab ') 2, Fv or a single chain Fv fragment (scFv).
25. Antibody molecule according to any one of claims 1 to 24, characterized in that it comprises a heavy chain constant region selected from I9G1, I9G2, I9G3 and I9gG4, and a light chain constant region chosen from kappa or lambda light chain constant regions.
26. Antibody molecule according to any one of claims 1 to 25, characterized in that it comprises a heavy chain constant region comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 92 to 103, 119 and 120, and / or a light chain constant region comprising the amino acid sequence of SEQ ID NO:
104.
27. The antibody molecule according to any one of claims 1 to 26, which has one or more (for example, 2, 3,4,5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, or more, for example, all) of the following properties: (i) selects human CD73 with a dissociation constant (Kp) of less than about 1 x 108 M, for example, when the molecule antibody is tested as a bivalent antibody molecule using Octet; (ii) binds to soluble human CD73 and / or human CD73 | membrane-bound; (iii) does not bind to murine CD73, for example, as
mined with the use of Octet;
(iv) inhibits or reduces the enzymatic activity of CD73 (for example, soluble human CD73 or membrane-bound human CD73), for example, human CD73-mediated conversion of adenosine monophosphate (AMP) into adenosine, for example , as measured by a malachite green phosphate (MG) assay or a modified Glo Cell Titration (CTG) assay;
(v) inhibits at least about 60%, 70%, 80%, or 90% of the membrane-bound human CD73 enzymatic activity, for example, when the antibody molecule is tested as a bivalent antibody molecule with use of a modified Cell Titration Glo (CTG) assay;
(vi) increases the proliferation of stimulated anti-CD3 / anti-CD28 T cells, for example, CD4 + T cells, in the presence of adenosine monophosphate (AMP), for example, as measured by a proliferation test CellTrace Violet (CTV) cell;
(vii) binds to the N-terminal domain of human CD73;
(viii) reduces hydrogen-deuterium exchange in one or more regions of a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence of SEQ ID NO : 171) when linked to it, in which one or more regions are selected from the group consisting of residues 158 to 172, residues 206 to 215, residues 368 to 387 and residues 87 to 104 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry;
(ix) when linked to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 (for example, a protein consisting of the amino acid sequence
those of SEQ ID NO: 171), induces a conformational exchange in residues 368 to 387 of SEQ ID NO: 105;
(x) contacts, for example, directly or indirectly, at least one, two, three or four residues within residues 158 to 172 of SEQ ID NO: 105;
(xi) contacts, for example, directly or indirectly, at least one, two, three, four or five residues within residues 206 to 215 of SEQ ID NO: 105;
(xii) contacts, for example, directly or indirectly, at least one, two, three, four or five residues within residues 368 to 387 of SEQ ID NO: 105 or 106;
(xiii) contacts, for example, directly or indirectly, at least one, two, three, four or five residues within residues 87 to 104 of SEQ ID NO: 105;
(xiv) binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second CD73 monomer, where, when the antibody molecule comprises a first binding domain antigen and a second antigen binding domain, where the first antigen binding domain binds to the first CD73 monomer and the second antigen binding domain binds to the second CD73 monomer, for example, when tested using size exclusion chromatography;
(xv) binds to a closed catalytically active conformation of human CD73 with lower affinity, for example, 50%, 60%, 70%, 80%, 90%, 95%, or 99% lower affinity, than when hand - antibody molecule binds to a catalytically inactive open conformation of human CD73;
(xvi) locks the human CD73 in an open conformation that is catalytically inactive; or
(xvii) prevents or reduces the conversion of human CD73 from a catalytically inactive open conformation to a closed catalytically active conformation, for example, reduces conversion by at least 1.5 times, 2 times, 5 times, 10 times, 20 times, 30 times, 40 times, 50 times, 60 times, 70 times, 80 times, 90 times or 100 times compared to conversion in the absence of the antibody molecule.
28. Antibody molecule according to any one of claims 1 to 27, characterized in that it comprises the antibody molecule binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second CD73 monomer, where, when the antibody molecule comprises a first antigen binding domain and a second antigen binding domain, the first antigen binding domain binds to the first CD73 monomer and the second antigen binding domain binds to the second CD73 monomer.
29. Composition, characterized by the fact that it comprises a plurality of antibody molecules, as defined in any one of claims 1 to 27, wherein the antibody molecules bind to a human CD73 dimer, in which the said dimer consisting of a first CD73 monomer and a second CD73 monomer, each monomer comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, where, when the antibody molecules in the plurality, each comprises the same first antigen binding domain and the same second antigen binding domain, at least 30%, 35% or 40% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, where each of the said complex consists of an antibody molecule and a dimer of
CD73.
30. Composition according to claim 29, characterized by the fact that at least 70%, 75% or 80% of the antibody molecules in said composition bind to the CD73 dimer to form said complex.
31. Composition, characterized by the fact that it comprises a plurality of antibody molecules, according to any one of claims 1 to 27, in which the antibody molecules bind to a human CD73 dimer, in which the said dimer consisting of a first CD73 monomer and a second CD73 monomer, wherein each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, wherein, when the antibody molecules in plurality, each comprises the same first antigen binding domain and the same second antigen binding domain, at most 60%, 65% or 70% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, wherein each complex comprises two or more antibody molecules and two or more CD73 dimers.
32. Composition according to claim 31, characterized by the fact that at most 20%, 25% or 30% of the antibody molecules in said composition bind to the CD73 dimer to form said complex.
33. Composition according to any one of claims 29 to 32, characterized by the fact that each CD73 monomer consists of the amino acid sequence of SEQ ID NO: 171.
34. Antibody molecule according to any one of claims 1 to 28, characterized in that the antibody molecule binds to a closed catalytically active conformation of human CD73 with lower affinity, for example, 50%, 60%, 70%,
80%, 90%, 95%, or 99% lower affinity, than when the antibody molecule binds to a catalytically inactive open conformation of human CD73.
35. Antibody molecule according to any of claims 1 to 28 or 34, characterized in that the antibody molecule prevents or reduces the conversion of human CD73 from a catalytically inactive open conformation to a catalytically active closed conformation , for example, reduces conversion by at least 1.5 times, 2 times, 5 times, 10 times, 20 times, 30 times, 40 times, 50 times, 60 times, 70 times, 80 times, 90 times or 100 times compared to conversion in the absence of the antibody molecule.
36. Antibody molecule according to any one of claims 1 to 28, 34 or 35, characterized in that the antibody molecule reduces hydrogen-deuterium exchange in one or more regions of a protein comprising the sequence of amino acids from residues 27 to 547 of SEQ ID NO: 105 when linked to it, in which one or more regions are selected from the group consisting of residues 158 to 172, residues 206 to 215, residues 368 to 387 and residues 87 to 104 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry.
37. The antibody molecule according to any one of claims 1a 28, or 34 to 36, characterized by the fact that the antibody molecule, when attached to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, reduces the average hydrogen-deuterium exchange in the residue (or residues) Xc selected from the core C region (residues 368 to 387 of SEQ ID NO: 105) to a greater extent than in the residue ( or residues) Xa selected from nucleus A region (residues 158 to 172 of SEQ ID NO: 105), residue (or residues) Xg selected from nucleus B region (residues 206 to 215 of SEQ ID NO: 105) or residue ( or residues) Xp selected from the D-region (residues 297 to 309 of SEQ ID NO: 105), for example, when the antibody molecule is tested as a bivalent antibody molecule using exchange mass spectrometry of hydrogen deuterium conducted for 1 minute in exchange at pH 7 to 8 (eg pH 7.5) and ambient temperature and, where: Xc is equal to 1,2, 3,4,5,6,7,8,9,10,15, or 20, Xa is equal to 1, 2, 3,4, 5, 6,7 , 8,9, 10,11, 12,13, or 14, Xg is equal to 1, 2,3,4,5,6,7,8,9,0uU 10, and Xp is equal to 1, 2, 3 , 4,5,6,7,8,9,10, 11.12, or 13.
38. Antibody molecule according to claim 37, characterized in that the antibody molecule, when linked to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, reduces the exchange of average hydrogen-deuterium in 20 residues selected from the nucleus C region to a greater extent than in: (i) 14 residues selected from the nucleus A region, (ii) 10 residues selected from the nucleus B region, or (iii ) 13 selected residues from the D-nucleus region, for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange for pH 7 to 8 (eg pH 7.5) and room temperature.
39. Antibody molecule according to claim 37, characterized by the fact that the antibody molecule, when attached to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, reduces the exchange of average hydrogen-deuterium in 15 selected residues from the nucleus C region to a greater extent than in: (1) 10 residues selected from the nucleus A region, (ii) 8 selected residues from the nucleus B region, or (iii ) 10 selected residues from the D-core region, for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange for pH 7 to 8 (eg pH 7.5) and room temperature.
40. Antibody molecule according to claim 37, characterized in that the antibody molecule, when attached to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, reduces the exchange of average hydrogen-deuterium in 10 selected residues from the nucleus C region to a greater extent than in: (i) 7 residues selected from the nucleus A region, (ii) 5 residues selected from the nucleus B region, or (iii ) 7 residues selected from the D-core region, for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange for pH 7 to 8 (eg pH 7.5) and room temperature.
41. Antibody molecule according to any of claims 37 to 40, characterized in that the antibody binding reduces the average hydrogen-deuterium exchange in the nucleus C region to a greater extent than in the nucleus region THE.
42. Antibody molecule according to any of claims 37 to 40, characterized in that the antibody binding reduces the average hydrogen-deuterium exchange in the nucleus C region to a greater extent than in the nucleus region B.
43. Antibody molecule according to any of claims 37 to 40, characterized in that the antibody binding reduces the average hydrogen-deuterium exchange in the nucleus C region to a greater extent than in the nucleus region D.
44. Antibody molecule according to any one of claims 37 to 40, characterized in that the antibody binding reduces the average hydrogen-deuterium exchange in the nucleus C region to a greater extent than in the nucleus region A and core region B.
45. Antibody molecule according to any of claims 37 to 40, characterized in that the antibody binding reduces the average hydrogen-deuterium exchange in the nucleus C region to a greater extent than in the nucleus region A and D core region.
46. Antibody molecule according to any one of claims 37 to 40, characterized in that the antibody binding reduces the average hydrogen-deuterium exchange in the nucleus C region to a greater extent than in the nucleus region B and D core region.
47. Antibody molecule according to any of claims 37 to 40, characterized in that the antibody binding reduces the average hydrogen-deuterium exchange in the nucleus C region to a greater extent than in the nucleus region A, region of nucleus B and region of nucleus D.
48. Antibody molecule according to any one of claims 1a 28, or 34 to 36, characterized by the fact that the antibody molecule, when linked to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, reduces the average hydrogen-deuterium exchange in the residue (or residues) Xa selected from the core A region (residues 158 to 172 of SEQ ID NO: 105) to a greater extent than in the residue (or residues) Xg selected from core B region (residues 206 to 215 of SEQ ID NO: 105), residue (or residues) Xc selected from core C region (residues 368 to 387 of SEQ ID NO: 105), or residue (or residues) ) Xp selected from core D region (residues 297 to 309 of SEQ ID NO: 105), for example, when the antibody molecule is tested as a bivalent antibody molecule using deuterium exchange mass spectrometry of hydrogen conducted for 1 minute in exchange for pH 7 to 8 (for example, pH 7.5) and room temperature, where: Xa is equal to 1, 2, 3.4, 5,6,7,8,9,10,11,12,13, or 14, Xg is equal to 1,2, 3,4,5,6,7 , 8,9, or 10, Xc is equal to 1, 2,3,4,5,6,7,8,9,10,15, or 20, and Xp is equal to 1, 2, 3,4,5 , 6,7,8, 9,10, 11,12, or 13.
49. Antibody molecule according to claim 48, characterized in that the antibody molecule, when linked to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, reduces the exchange of average hydrogen-deuterium in 14 selected residues from the region of nucleus À to a greater extent than in: (i) 10 residues selected from the region of nucleus B, (ii) 20 residues selected from the region of nucleus C, or (iii ) 13 selected residues from the D-nucleus region, for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange for pH 7 to 8 (eg pH 7.5) and room temperature
you.
50. Antibody molecule according to claim 48, characterized in that the antibody molecule, when linked to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, reduces the exchange of average hydrogen-deuterium in 10 residues selected from the nucleus A region to a greater extent than in: (i) 8 residues selected from the nucleus B region, (ii) 15 residues selected from the nucleus C region, or (ili ) 10 selected residues from the D-core region, for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange for pH 7 to 8 (eg pH 7.5) and room temperature.
51. Antibody molecule according to claim 48, characterized in that the antibody molecule, when linked to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, reduces the exchange of average hydrogen-deuterium in 7 residues selected from the region of nucleus À to a greater extent than in: (i) 5 residues selected from the region of nucleus B, (ii) 10 residues selected from the region of nucleus C, or (iii ) 7 residues selected from the D-core region, for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange for pH 7 to 8 (eg pH 7.5) and room temperature.
52. Antibody molecule according to any one of claims 48 to 51, characterized in that the antibody binding reduces the average hydrogen-deuterium exchange in the nucleus A region to a greater extent than in the nucleus B region .
53. Antibody molecule according to any one of claims 48 to 51, characterized by the fact that the antibody binding reduces the average hydrogen-deuterium exchange in the nucleus A region to a greater extent than in the nucleus C region .
54. Antibody molecule according to any one of claims 48 to 51, characterized in that the antibody binding reduces the average hydrogen-deuterium exchange in the nucleus A region to a greater extent than in the nucleus region D.
55. Antibody molecule according to any of claims 48 to 51, characterized in that the antibody binding reduces the average hydrogen-deuterium exchange in the nucleus A region to a greater extent than in the nucleus region B and C core region.
56. Antibody molecule according to any one of claims 48 to 51, characterized by the fact that the antibody binding reduces the average hydrogen-deuterium exchange in the nucleus A region to a greater extent than in the nucleus region B and D core region.
57. Antibody molecule according to any one of claims 48 to 51, characterized in that the antibody binding reduces the average hydrogen-deuterium exchange in the nucleus A region to a greater extent than in the nucleus region C and D core region.
58. Antibody molecule according to any one of claims 48 to 51, characterized in that the antibody binding reduces the average hydrogen-deuterium exchange in the nucleus A region to a greater extent than in the nucleus region B, region of num-
cleo C and D region.
59. Antibody molecule according to any one of claims 1 to 28, or 34 to 58, characterized in that the antibody molecule reduces the hydrogen-deuterium exchange in one or more regions of a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 when linked to it, where one or more regions are selected from the group consisting of residues 158 to 172, residues 206 to 215, residues 368 to 387 and residues 87 to 104 of SEQ ID NO: 105, in which the region that has the greatest reduction in the average hydrogen-deuterium exchange among one or more regions does not consist of residues 206 to 215 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (eg pH 7, 5) and room temperature.
60. Antibody molecule according to any one of claims 1 to 28, or 34 to 59, characterized in that the antibody molecule, when linked to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, leads to: (i). a reduction in the average hydrogen-deuterium exchange of more than, for example, 0.02, 0.03, 0.04, 0.05 or 0.06 Da per residue in residues 368 to 387 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5 ) and room temperature; or (li) a reduction in the average hydrogen-deuterium exchange of less than, for example, 0.05, 0.04, 0.03 or 0.02 Da per residue in residues 206 to 215 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a divalent antibody molecule using hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7 , 5) and room temperature.
61. Antibody molecule according to any one of claims 36 to 60, characterized in that the protein consists of the amino acid sequence of SEQ ID NO: 171.
62. Antibody molecule according to any one of claims 1 to 28, or 34 to 61, characterized in that the antibody molecule contacts, for example, directly or indirectly, at least one, two, three or four residues within residues 158 to 172 of SEQ ID NO: 105.
63. Antibody molecule according to any one of claims 1 to 28, or 34 to 62, characterized in that the antibody molecule contacts, for example, directly or indirectly, at least one, two, three, four or five residues within residues 206 to 215 of SEQ ID NO: 105.
64. Antibody molecule according to any one of claims 1 to 28, or 34 to 63, characterized in that the antibody molecule contacts, for example, directly or indirectly, at least one, two, three, four or five residues within residues 368 to 387 of SEQ ID NO: 105 or 106.
65. Antibody molecule according to any one of claims 1 to 28, or 34 to 64, characterized in that the antibody molecule contacts, for example, directly or indirectly, at least one, two, three, four or five residues within residues 87 to 104 of SEQ ID NO: 105.
66. Antibody molecule according to any one of claims 1 to 28, or 34 to 65, characterized in that the antibody molecule reduces tandem mass tag (TMT) identification in residue K136 (numbered according to SEQ ID NO: 105) of a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 when linked to the same, for example, when the antibody molecule is tested as an antibody molecule bivalent with the use of TMT isotope identification, for example, with the methods described in Example 9, for example, with the use of TMT isotope identification conducted by 30 s identification time, optionally where the antibody molecule further reduces the identification of TMT in one or more of residues K133, K162, K179, K206, K214, K285, K291 and K341 (numbered according to SEQ ID NO: 105) of the protein, for example example, in one or more of the residues K162, K206, K214, K285, K291 and K341 (number those according to SEQ ID NO: 105) of the protein.
67. Antibody molecule according to any one of claims 1 to 28, or 34 to 66, characterized in that the antibody molecule increases the identification of TMT in one or both residues K262 and K274 (numbered from according to SEQ ID NO: 105) of the protein when bound to them, for example, when the antibody molecule is tested as a bivalent antibody molecule with the use of TMT isotope identification, for example, with the use of methods described in Example 9, for example, using the TMT isotope identification conducted by the identification time of 30 s.
68. Antibody molecule according to any one of claims 1 to 28, or 34 to 67, characterized by the fact that the antibody molecule reduces tandem mass tag (TMT) identification in residues K206 and K214 (numbered from according to SEQ ID NO: 105) of a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 when linked to it, where the reduction in residue K214 is not less than 90, 80, 70, 60 or 50% reduction in the K206 residue, for example, when the antibody molecule is tested as a bivalent antibody molecule using the TMT isotope identification, for example, using the methods described in Example 9 , for example, where the reduction in residue K214 is not less than 90, 80, 70, 60 or 50% of the reduction in residue K206 with the use of TMT isotope identification conducted by the identification time of 30 s, for example , in which the reduction in K214 residue is not less than 90, 80, 70, 60, 50, 40 or 30% of the reduction in residue K206 using the TMT isotope identification conducted by the identification time of 300 s.
69. Antibody molecule according to any one of claims 1 to 28, or 34 to 68, characterized in that the antibody molecule reduces tandem mass tag (TMT) identification in residue K162 (numbered according to SEQ ID NO: 105) of a protein that comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 when bound to it, when the antibody molecule is tested using the identification of isotope of TMT conducted by the identification time of 30 s, in which: (i) the antibody molecule does not reduce the identification of TMT in the K162 residue when tested using the use of TMT isotope conducted by the identification time of 300 s , or (ii) the antibody molecule reduces the identification of TMT in the K162 residue when tested using the TMT isotope identification conducted by the identification time of 300 s, in which the reduction in the K162 residue under the time of 300 s identification is not more than 20, 30, 40 or 50% reduction in K162 residue under 30 s identification time,
for example, when the antibody molecule is tested as a bivalent antibody molecule, for example, using the methods described in Example 9.
70. Antibody molecule according to any one of claims 1 to 28, or 34 to 69, characterized in that the antibody molecule binds to one or more CD73 residues, for example, through an electrostatic interaction and / or a hydrogen bond, where the one or more residues are selected from the group consisting of residues Y110, K136, L132, L157, K162, S155 and T209, numbered according to SEQ ID NO: 105, by example, as measured using crystal structure analysis, for example, using methods described in Example 8.
71. The antibody molecule according to any one of claims 1 to 28, or 34 to 70, characterized in that the antibody molecule comprises a heavy chain variable region and a light chain variable region, wherein the antibody molecule has one or more (for example, 1, 2, 3, 4 or all) of the following properties: (i) the antibody molecule (for example, the heavy chain variable region, for example, R54 of the heavy chain variable region, numbered according to Kabat numbering) binds to Y 110 (for example, the main chain carbonyl of Y110) or K136 (for example, the main chain carbonyl of K136) of CD73 (numbered according to SEQ ID NO: 105), for example, through an electrostatic interaction, for example, as measured using the crystal structure analysis, for example, using the methods described in Example 8, (ii) the antibody molecule (for example, the heavy chain variable region, for example, R31 of the heavy chain variable, numbered according to Kabat numbering) is linked to
CD13 L132 (eg L132 main chain carbonyl) or CD15 L157 (eg L157 main chain carbonyl) (numbered according to SEQ ID NO: 105), for example, by means of a electrostatic interaction, for example, as measured using crystal structure analysis, for example, using the methods described in Example 8,
(iii) the antibody molecule (for example, the heavy chain variable region, for example, S99 (for example, the main chain carbonyl of S99), R31 (for example, the main chain carbonyl of R31) or E95 (for example, the E95 side chain) of the heavy chain variable region, numbered according to Kabat numbering) binds to CD73 K162 (for example, the K162 side chain) (numbered according to SEQ ID NO: 105), for example, through an electrostatic interaction, for example, as measured using crystal structure analysis, for example, using the methods described in Example 8,
(iv) the antibody molecule (for example, the heavy chain variable region, for example, E98 of the heavy chain variable region, numbered according to Kabat numbering) binds to 8155 (for example, the side chain of S155) of CD73 (numbered according to SEQ ID NO: 105), for example, by means of a hydrogen bond, for example, as measured with the use of crystal structure analysis, for example, with the use of methods described in Example 8, and
(v) the antibody molecule (for example, the light chain variable region, for example, W32 (for example, the W32 side chain) of the light chain variable region, numbered according to Kabat numbering) binds to T209 (for example, the T209 side chain) of CD73 (numbered according to SEQ ID NO: 105), for example, via a hydrogen bond, for example, as measured using the analysis of crystal structure, for example, using the methods described in Example 8.
72. Antibody molecule according to any one of claims 1 to 28, or 34 to 71, characterized in that the antibody molecule binds to one or more regions of CD73, for example, by means of format complementarity and / or a Van der Waal interaction, in which one or more regions are selected from the group consisting of residues 155 to 170, 136 to 138 and 209 to 210, numbered according to SEQ ID NO : 105, for example, as measured using crystal structure analysis, for example, using the methods described in Example 8.
73. The antibody molecule according to any one of claims 1 to 28, or 34 to 72, characterized in that the antibody molecule comprises a heavy chain variable region and a light chain variable region, wherein the antibody molecule has one or more (for example, 1, 2 or all) of the following properties: (i) the antibody molecule (for example, the heavy chain variable region, for example, residue 33, 50, 52, 56, 97, 98, 100 or 100a of the heavy chain variable region, numbered according to the Kabat numbering) binds residues 155 to 170 of CD73 (numbered according to SEQ ID NO: 105), for example, through format complementarity and / or a Van der Waal interaction, for example, as measured using crystal structure analysis, for example, using the methods described in Example 8, (ii) the antibody molecule (for example, the heavy chain variable region, for example, residue 30 or 31 of the variable chain region heavy ia, numbered according to Kabat numbering) binds to residues 136 to 138 of CD73 (numbered according to
SEQ ID NO: 105), for example, through format complementarity and / or a Van der Waal interaction, for example, as measured with the use of crystal structure analysis, for example, with the use of methods described in Example 8, and (ili) the antibody molecule (for example, the light chain variable region, for example, residue 30 or 32 of the light chain variable region, numbered according to Kabat numbering) if binds to residues 209 to 210 of CD73 (numbered according to SEQ ID NO: 105), for example, as measured with the use of crystal structure analysis, for example, with the methods described in the Example 8.
74. Antibody molecule that binds to a human CD73 dimer, characterized by the fact that said dimer that consists of a first CD73 monomer and a second CD73 monomer, where, when the antibody molecule comprises a first antigen binding domain and a second antigen binding domain, the first antigen binding domain binds to the first CD73 monomer and the second antigen binding domain binds to the second CD73 monomer .
75. Composition, characterized by the fact that it comprises a plurality of antibody molecules that binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second CD73 monomer, wherein each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, where, when the antibody molecules in plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at least 30%, 35% or 40% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, each of which consists of a antibody molecule and a CD73 dimer.
76. Composition according to claim 75, characterized by the fact that at least 70%, 75% or 80% of the antibody molecules in said composition bind to the CD73 dimer to form said complex.
77. Composition, characterized by the fact that it comprises a plurality of antibody molecules that binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second CD73 monomer, wherein each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, where, when the antibody molecules in plurality each comprise the same first antigen binding domain and the same second antigen binding domain, a maximum of 60%, 65% or 70% of the antibody molecules in said composition bind to the CD73 dimer to form a complex, each of which comprises two or more more antibody molecules and two or more dimers of CD73.
78. Composition according to claim 77, characterized by the fact that at most 20%, 25% or 30% of the antibody molecules in said composition bind to the CD73 dimer to form said complex.
79. Composition according to any one of claims 75 to 78, characterized by the fact that each CD73 monomer consists of the amino acid sequence of SEQ ID NO: 171.
80. Antibody molecule that binds to human CD73, characterized by the fact that the antibody molecule binds to a closed, catalytically active conformation of human CD73 with lower affinity, for example, 50%, 60%, 70 %, 80%, 90%, 95% or 99% lower affinity than when the antibody molecule binds to a catalytically inactive open conformation of human CD73.
81. Antibody molecule that binds to human CD73, characterized by the fact that the antibody molecule prevents or reduces the conversion of human CD73 from a catalytically inactive open conformation to a catalytically active closed conformation, for example, reduces conversion at least 1.5 times, 2 times, 5 times, times, 20 times, 30 times, 40 times, 50 times, 60 times, 70 times, 80 times, 90 times or 100 times compared to conversion in the absence of the antibody molecule.
82. Antibody molecule that binds to human CD73, characterized by the fact that the antibody molecule reduces the hydrogen-deuterium exchange in one or more regions of a protein that comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 when connected to it, where one or more regions are selected from the group consisting of residues 158 to 172, residues 206 to 215, residues 368 to 387 and residues 87 to 104 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule with the use of hydrogen deuterium exchange mass spectrometry.
83. Antibody molecule that binds to human CD73, characterized by the fact that the antibody molecule, when bound to a protein that comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, reduces the average hydrogen-deuterium exchange in the residue (or residues) Xc selected from the core C region (residues 368 to 387 of SEQ ID NO: 105) to a greater extent than in the residue (or residues) Xa selected from core A region (residues 158 to 172 of SEQ ID NO: 105), residue (or residues) Xg selected from core B region (residues 206 to 215 of SEQ ID NO: 105) or residue (or residues) Xp selected from nucleus D region (residues 297 to 309 of SEQ ID NO: 105), for example,
when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and temperature environment, where: Xc is equal to 1,2, 3,4,5,6,7,8,9,10,15, or 20, Xa is equal to 1, 2, 3,4, 5,6,7, 8,9,10,11,12,13, or 14, Xg is equal to 1,2,3,4,5,6,7,8,9,0u 10, and Xp is equal to 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, or 13.
84. Antibody molecule according to claim 83, characterized in that the antibody molecule, when linked to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, reduces the exchange of average hydrogen-deuterium in 20 residues selected from the nucleus C region to a greater extent than in: (i) 14 residues selected from the nucleus A region, (ii) 10 residues selected from the nucleus B region, or (iii ) 13 selected residues from the D-nucleus region, for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange for pH 7 to 8 (eg pH 7.5) and room temperature.
85. Antibody molecule according to claim 83, characterized in that the antibody molecule, when linked to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, reduces the exchange of average hydrogen-deuterium in 15 selected residues from the nucleus C region to a greater extent than in: (i) 10 residues selected from the nucleus A region, (ii) 8 residues selected from the nucleus B region, or
(iii) 10 selected residues from the D-nucleus region, for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry conducted for 1 minute in changes at pH 7 to 8 (eg pH 7.5) and room temperature.
86. Antibody molecule according to claim 83, characterized in that the antibody molecule, when linked to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, reduces the exchange of average hydrogen-deuterium in 10 selected residues from the nucleus C region to a greater extent than in: (i) 7 residues selected from the nucleus A region, (ii) 5 residues selected from the nucleus B region, or (iii ) 7 residues selected from the D-core region, for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange for pH 7 to 8 (eg pH 7.5) and room temperature.
87. Antibody molecule according to any of claims 83 to 86, characterized in that the antibody binding reduces the average hydrogen-deuterium exchange in the nucleus C region to a greater extent than in the nucleus region THE.
88. Antibody molecule according to any of claims 83 to 86, characterized in that the antibody binding reduces the average hydrogen-deuterium exchange in the nucleus C region to a greater extent than in the nucleus region B.
89. Antibody molecule according to any of claims 83 to 86, characterized in that the antibody binding reduces the average hydrogen-deuterium exchange in the nucleus C region to a greater extent than in the nucleus region D.
90. Antibody molecule according to any of claims 83 to 86, characterized in that the antibody binding reduces the average hydrogen-deuterium exchange in the nucleus C region to a greater extent than in the nucleus region A and core region B.
91. Antibody molecule according to any of claims 83 to 86, characterized in that the antibody binding reduces the average hydrogen-deuterium exchange in the nucleus C region to a greater extent than in the nucleus region A and D core region.
92. Antibody molecule according to any of claims 83 to 86, characterized in that the antibody binding reduces the average hydrogen-deuterium exchange in the nucleus C region to a greater extent than in the nucleus region B and D core region.
93. Antibody molecule according to any of claims 83 to 86, characterized in that the antibody binding reduces the average hydrogen-deuterium exchange in the nucleus C region to a greater extent than in the nucleus region A, region of nucleus B and region of nucleus D.
94. Antibody molecule that binds to human CD73, characterized by the fact that the antibody molecule, when bound to a protein that comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, reduces the average hydrogen-deuterium exchange in the residue (or residues) Xa selected from the region of nucleus A (residues 158 to 172 of SEQ ID NO: 105) to a greater extent than in the residue (or residues) Xg selected from the region of cleo B (residues 206 to 215 of SEQ ID NO: 105), residue (or residues)
Xc selected from core C region (residues 368 to 387 of SEQ ID NO: 105) or residue (or residues) Xp selected from core D region (residues 297 to 309 of SEQ ID NO: 105), for example, when antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (eg pH 7.5) and room temperature, where: Xa is equal to 1, 2, 3.4, 5,6,7,8,9,10,11,12,13, or 14, Xg is equal to 1, 2, 3,4,5, 6,7,8,9, ou10, Xc is equal to 1, 2,3,4,5,6,7,8,9,10,15, or 20, and Xp is equal to 1, 2, 3,4 , 5,6,7,8,9,10, 11,12, or 13.
95. Antibody molecule according to claim 94, characterized in that the antibody molecule, when linked to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, reduces the exchange of average hydrogen-deuterium in 14 selected residues from the region of nucleus À to a greater extent than in: (i) 10 residues selected from the region of nucleus B, (ii), 20 residues selected from the region of nucleus C, or ( iii) 13 selected residues from the D-core region, for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature.
96. Antibody molecule according to claim 94, characterized in that the antibody molecule, when linked to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, reduces the exchange of average hydrogen-deuterium in 10 residues selected from the region of nucleus À to a greater extent than in: (i) 8 residues selected from the region of nucleus B, (ii) 15 residues selected from the region of nucleus C, or (iii ) 10 selected residues from the D-core region, for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange for pH 7 to 8 (eg pH 7.5) and room temperature.
97. Antibody molecule according to claim 94, characterized in that the antibody molecule, when linked to a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, reduces the exchange of average hydrogen-deuterium in 7 residues selected from the region of nucleus À to a greater extent than in: (i) 5 residues selected from the region of nucleus B, (ii) 10 residues selected from the region of nucleus C, or (iii ) 7 residues selected from the D-core region, for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange for pH 7 to 8 (eg pH 7.5) and room temperature.
98. Antibody molecule according to any of claims 94 to 97, characterized in that the antibody binding reduces the average hydrogen-deuterium exchange in the nucleus A region to a greater extent than in the nucleus region B.
99. Antibody molecule according to any of claims 94 to 97, characterized by the fact that the antibody binding reduces the average hydrogen-deuterium exchange in the region of
cleo A to a greater extent than in the C region.
100. Antibody molecule according to any one of claims 94 to 97, characterized by the fact that the antibody binding reduces the average hydrogen-deuterium exchange in the nucleus A region to a greater extent than in the nucleus region D.
101. Antibody molecule according to any one of claims 94 to 97, characterized in that the antibody binding reduces the average hydrogen-deuterium exchange in the nucleus A region to a greater extent than in the nucleus region B and C core region.
102. Antibody molecule according to any one of claims 94 to 97, characterized by the fact that the antibody binding reduces the average hydrogen-deuterium exchange in the nucleus A region to a greater extent than in the nucleus region B and D core region.
103. Antibody molecule according to any of claims 94 to 97, characterized in that the antibody binding reduces the average hydrogen-deuterium exchange in the nucleus A region to a greater extent than in the nucleus region C and D core region.
104. Antibody molecule according to any of claims 94 to 97, characterized in that the antibody binding reduces the average hydrogen-deuterium exchange in the nucleus A region to a greater extent than in the nucleus region B, region of nucleus C and region of nucleus D.
105. Antibody molecule that binds to human CD73, characterized by the fact that the antibody molecule reduces hydrogen-deuterium exchange in one or more regions of a protein that comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 when connected to it, where one or more regions
tions are selected from the group consisting of residues 158 to 172, residues 206 to 215, residues 368 to 387 and residues 87 to 104 of SEQ ID NO: 105, in which the region that has the greatest reduction in hydrogen- mean deuterium among one or more regions does not consist of residues 206 to 215 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule using mass spectrometry of hydrogen deuterium exchange conducted for 1 minute in exchange for pH 7 to 8 (for example, pH 7.5) and room temperature.
106. Antibody molecule that binds to human CD73, characterized by the fact that the antibody molecule, when bound to a protein that comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, leads a: (i): a reduction in the average hydrogen-deuterium exchange of more than, for example, 0.02, 0.03, 0.04, 0.05 or 0.06 Da per residue in residues 368 to 387 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a bivalent antibody molecule using hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7.5) and room temperature; or (ii) a reduction in the average hydrogen-deuterium exchange of less than, for example, 0.05, 0.04, 0.03 or 0.02 Da per residue in residues 206 to 215 of SEQ ID NO: 105, for example, when the antibody molecule is tested as a divalent antibody molecule using hydrogen deuterium exchange mass spectrometry conducted for 1 minute in exchange at pH 7 to 8 (for example, pH 7 , 5) and room temperature.
107. Antibody molecule according to any one of claims 82 to 106, characterized in that the protein consists of the amino acid sequence of SEQ ID NO: 171.
108. Antibody molecule that binds to human CD73, characterized by the fact that the antibody molecule contacts, for example, directly or indirectly, at least one, two, three or four residues within residues 158 to 172 of SEQ ID NO: 105.
109. Antibody molecule that binds to human CD73, characterized by the fact that the antibody molecule contacts, for example, directly or indirectly, at least one, two, three, four or five residues within residues 206 to 215 SEQ ID NO: 105.
110. Antibody molecule that binds to human CD73, characterized by the fact that the antibody molecule contacts, for example, directly or indirectly, at least one, two, three, four or five residues within residues 368 to 387 SEQ ID NO: 105 or
106.
111. Antibody molecule that binds to human CD73, characterized by the fact that the antibody molecule contacts, for example, directly or indirectly, at least one, two, three, four or five residues within residues 87 to 104 SEQ ID NO: 105.
112. Antibody molecule that binds to human CD73, characterized by the fact that the antibody molecule reduces tandem mass tag (TMT) identification at residue K136 (numbered according to SEQ ID NO: 105) of a protein that comprises the amino acid sequence of residues 27 to 547 of SEQ | D NO: 105 when bound to it, for example, when the antibody molecule is tested as a bivalent antibody molecule using identification of the TMT isotope, for example, using the methods described in Example 9, for example, using the TMT isotope identification conducted by an identification time of 30 Ss, optionally in which the antibody molecule reduces additionally the identification of TMT in one or more among residues K133, K162, K179, K206, K214, K285, K291 and K341 (numbered according to
SEQ ID NO: 105) of the protein, for example, in one or more of residues K162, K206, K214, K285, K291 and K341 (numbered according to SEQ ID NO: 105) of the protein.
113. Antibody molecule according to claim 112, characterized by the fact that the antibody molecule increases the identification of TMT in one or both residues K262 and K274 (numbered according to SEQ ID NO: 105) protein when bound to them, for example, when the antibody molecule is tested as a bivalent antibody molecule using the TMT isotope identification, for example, using the methods described in Example 9, for example, with the use of TMT isotope identification conducted by the 30 s identification time.
114. Antibody molecule that binds to human CD73, characterized by the fact that the antibody molecule reduces tandem mass tag (TMT) identification in residues K206 and K214 (numbered according to SEQ ID NO: 105) of a protein comprising the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 when attached to it, where the reduction in residue K214 is not less than 90, 80, 70, 60 or 50% of the reduction at residue K206, for example, when the antibody molecule is tested as a bivalent antibody molecule using the TMT isotope identification, for example, using the methods described in Example 9, for example, in which the reduction in residue K214 is not less than 90, 80, 70, 60 or 50% of the reduction in residue K206 with the use of TMT isotope identification conducted by the identification time of 30 s, for example, in which the reduction in residue K214 is not less than 90, 80, 70, 60, 50, 40 or 30% of the reduction in residue K206 using the identifi cation of TMT isotope conducted by the identification time of 300 s.
115. Antibody molecule that binds to human CD73, causes
characterized by the fact that the antibody molecule reduces tandem mass tag (TMT) identification at residue K162 (numbered according to SEQ ID NO: 105) of a protein that comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105 when bound to it, when the antibody molecule is tested using the TMT isotope identification conducted by the 30 s identification time, where: (i) the antibody molecule does not reduce the TMT identification on the K162 residue when tested using the TMT isotope identification conducted by the 300 s identification time, or (ii) the antibody molecule reduces the TMT identification on the K162 residue when tested with the use of the TMT isotope identification conducted by the 300 s identification time, where the reduction in the K162 residue under the 300 s identification time is not more than 20, 30, 40 or 50% of the reduction in the K162 residue under the identification time of 30 s, for example, when the molecule antibody cell is tested as a divalent antibody molecule, for example, using methods described in Example 9.
116. Antibody molecule that binds to human CD73, characterized by the fact that the antibody molecule binds to one or more CD73 residues, for example, through an electrostatic interaction and / or a binding of hydrogen, in which the one or more residues are selected from the group consisting of residues Y110, K136, L132, L157, K162, S155 and T209, numbered according to SEQ ID NO: 105, for example, as measured with the use of crystal structure analysis, for example, with the use of the methods described in Example 8.
117. Antibody molecule according to claim 116, characterized in that the antibody molecule comprises
has a heavy chain variable region and a light chain variable region, where the antibody molecule has one or more (for example, 1, 2, 3, 4 or all) of the following properties:
(i). the antibody molecule (for example, the heavy chain variable region, for example, R54 of the heavy chain variable region, numbered according to Kabat numbering) binds to Y110 (for example, the main chain carbonyl) Y110) or K136 (for example, the main chain carbonyl of K136) of CD73 (numbered according to SEQ ID NO: 105), for example, through an electrostatic interaction, for example, as measured with use crystal structure analysis, for example, using the methods described in Example 8,
(ii) the antibody molecule (for example, the heavy chain variable region, for example, R31 of the heavy chain variable region, numbered according to Kabat numbering) binds to L132 (for example, carbonyl from main chain of L132) or L157 (for example, the carbonyl of the main chain of L157) of CD73 (numbered according to SEQ ID NO: 105), for example, through an electrostatic interaction, for example, as measured using crystal structure analysis, for example, using the methods described in Example 8,
(iii) the antibody molecule (for example, the heavy chain variable region, for example, S99 (for example, the main chain carbonyl of S99), R31 (for example, the main chain carbonyl of R31) or E95 (for example, the E95 side chain) of the heavy chain variable region, numbered according to Kabat numbering) binds to CD73 K162 (for example, the K162 side chain) (numbered according to SEQ ID NO: 105), for example, through an electrostatic interaction, for example, as measured with the use of crystal structure analysis, for example, with the use of
all described in Example 8, (iv) the antibody molecule (for example, the heavy chain variable region, for example, E98 of the heavy chain variable region, numbered according to Kabat numbering) binds to S155 ( (eg, the S155 side chain) of CD73 (numbered according to SEQ ID NO: 105), for example, by means of a hydrogen bond, for example, as measured using crystal structure analysis, for example example, using the methods described in Example 8, and (v) the antibody molecule (for example, the light chain variable region, for example, W32 (for example, the W32 side chain) of the variable region light chain, numbered according to Kabat numbering) binds to CD20 T209 (for example, T209 side chain) (numbered according to SEQ ID NO: 105), for example, via a hydrogen, for example, as measured using crystal structure analysis, for example, using methods described in Example 8.
118. Antibody molecule according to claim 116 or 117, characterized in that the antibody molecule binds to one or more regions of CD73, for example, by way of complementary format and / or an interaction Van der Waal, in which one or more regions are selected from the group consisting of residues 155 to 170, 136 to 138 and 209 to 210, numbered according to SEQ ID NO: 105, for example, as measured with the use of crystal structure analysis, for example, with the use of the methods described in Example 8.
119. Antibody molecule according to any one of claims 116 to 118, characterized in that the antibody molecule comprises a heavy chain variable region and a light chain variable region, wherein the antibody molecule has one or more (for example, 1, 2 or all) of the following properties: (i). the antibody molecule (for example, the heavy chain variable region, for example, residue 33, 50, 52, 56, 97, 98, 100, or 100a of the heavy chain variable region, numbered according to the numbering of Kabat) binds to residues 155 to 170 of CD73 (numbered according to SEQ ID NO: 105), for example, through format complementarity and / or a Van der Waal interaction, for example, as measured with use of crystal structure analysis, for example, using the methods described in Example 8, (ii) the antibody molecule (for example, the heavy chain variable region, for example, residue 30 or 31 of the variable region - heavy chain level, numbered according to Kabat numbering) binds to residues 136 to 138 of CD73 (numbered according to SEQ ID NO: 105), for example, through format complementarity and / or an interaction of Van der Waal, for example, as measured using crystal structure analysis, for example, using methods described in E example 8, and (iii) the antibody molecule (for example, the light chain variable region, for example, residue 30 or 32 of the light chain variable region, numbered according to Kabat numbering) binds to the residues 209 to 210 of CD73 (numbered according to SEQ ID NO: 105), for example, as measured with the use of crystal structure analysis, for example, with the methods described in Example 8.
120. Antibody molecule according to any one of claims 82 to 119, characterized in that the antibody molecule binds to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second
of the CD73 monomer, where, when the antibody molecule comprises a first antigen binding domain and a second antigen binding domain, the first antigen binding domain binds to the first CD73 monomer and the second domain binding antigen binds to the second CD73 monomer.
121. Composition, characterized by the fact that it comprises a plurality of antibody molecules, as defined in any one of claims 82 to 119, wherein the antibody molecules bind to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second CD73 monomer, where each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, where, when the antibody molecules in the plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at least 30%, 35% or 40% of the antibody molecules in said composition bind to the CD73 dimer to form a complex , wherein each of said complex consists of an antibody molecule and a CD73 dimer.
122. Composition according to claim 121, characterized by the fact that at least 70%, 75% or 80% of the antibody molecules in said composition bind to the CD73 dimer to form said complex.
123. Composition, characterized in that it comprises a plurality of antibody molecules, as defined in any one of claims 82 to 119, wherein the antibody molecules bind to a human CD73 dimer, wherein said dimer consisting of a first CD73 monomer and a second CD73 monomer, where each monomer comprises the amino acid sequence of residues 27 to 547 of SEQ ID NO: 105, where, when the antibody molecules in the plurality each comprise the same first antigen binding domain and the same second antigen binding domain, at most 60%, 65% or 70% of the antibody molecules in said composition bind to the CD73 dimer to form a complex , wherein each of said complex comprises two or more antibody molecules and two or more CD73 dimers.
124. Composition according to claim 123, characterized by the fact that a maximum of 20%, 25% or 30% of the antibody molecules in said composition bind to the CD73 dimer to form said complex.
125. Composition according to any one of claims 121 to 124, characterized by the fact that each CD73 monomer consists of the amino acid sequence of SEQ ID NO: 171.
126. Antibody molecule according to any of claims 82 to 119, characterized in that the antibody molecule binds to a closed catalytically active conformation of human CD73 with lower affinity, for example, 50%, 60%, 70%, 80%, 90%, 95%, or 99% lower affinity, than when the antibody molecule binds to a catalytically inactive open conformation of human CD73.
127. Antibody molecule according to any of claims 82 to 119, characterized in that the antibody molecule prevents or reduces the conversion of human CD73 from a catalytically inactive open conformation to a catalytically active closed conformation, for example For example, it reduces conversion by at least 1.5 times, 2 times, 5 times, 10 times, 20 times, 30 times, 40 times, 50 times, 60 times, 70 times, 80 times, 90 times or 100 times compared to conversion in the absence of the antibody molecule.
128. Antibody molecule, characterized by the fact that it competes for binding to human CD73 with the antibody molecule or composition, as defined in any one of claims 1 to 127.
129. Antibody molecule, characterized by the fact that it binds to the same epitope as, substantially the same epitope as, an epitope that overlaps with, or an epitope that substantially overlaps with the epitope of the antibody molecule or composition as defined in any one of claims 1 to 128.
130. Pharmaceutical composition, characterized by the fact that it comprises the antibody molecule or composition, as defined in any of claims 1 to 129 and a pharmaceutically acceptable stabilizer, excipient or carrier.
131. Nucleic acid, characterized by the fact that it encodes the variable heavy or light chain variable region of the antibody molecule, as defined in any one of claims 1 to 28, 34 to 74, 80 to 120, or 126 to 129.
132. Nucleic acid, characterized by the fact that it encodes a variable region of heavy chain, in which the nucleic acid comprises the nucleotide sequence of SEQ ID NO: 45, 78, 85, 143, 152, 160, 67, 32 , 11, or 169, or a nucleotide sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 45, 78, 85, 143, 152, 160 , 67, 32, 11, or
169.
133. Nucleic acid according to claim 132, characterized by the fact that the nucleic acid encodes a heavy chain, wherein the nucleic acid comprises the nucleotide sequence of SEQ ID NO: 47, 80, 87, 69, 34, or 13, or a nucleotide sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 47, 80, 87, 69, 34, or 13.
134. Nucleic acid, characterized by the fact that it encodes a variable region of light chain, in which the nucleic acid comprises the nucleotide sequence of SEQ ID NO: 56, 144, 22 or 170, or a nucleotide sequence that has at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 56, 144, 22 or 170.
135. Nucleic acid according to claim 134, characterized by the fact that the nucleic acid encodes a light chain, wherein the nucleic acid comprises the nucleotide sequence of SEQ ID NO: 58 or 24, or a sequence of nucleotides that have at least about 85%, 90%, 95%, or 99% sequence identity with SEQ ID NO: 58 or 24.
136. Expression vector, characterized by the fact that it comprises nucleic acid, as defined in any one of claims 131 to 135.
137. Host cell, characterized by the fact that it comprises the nucleic acid, as defined in any one of claims 131 to 135 or the expression vector, as defined in claim 136.
138. Method for producing an antibody molecule, characterized by the fact that the method comprises culturing the host cell, as defined in claim 137, under conditions suitable for gene expression.
139. Method for stimulating an immune response in an individual, characterized by the fact that the method comprises administering to the individual the antibody molecule, composition or pharmaceutical composition, as defined in any one of claims 1 to 130, in a effective amount to stimulate the immune response.
140. Method for treating cancer in an individual, characterized in that the method comprises administering to the individual the antibody molecule, composition or pharmaceutical composition, as defined in any one of claims 1 to 130, in an amount effective to treat cancer.
141. Method according to claim 140, characterized by the fact that cancer is chosen from lung cancer (eg, non-small cell lung cancer), pancreatic cancer (eg, pancreatic ductal adenocarcinoma), breast cancer (for example, triple negative breast cancer), melanoma, head and neck cancer (for example, scaly head and neck cancer), colorectal cancer (for example, colorectal cancer microsatellite stable (MSS)), ovarian cancer or kidney cancer (eg renal cell carcinoma).
142. Method according to any one of claims 139 to141, characterized in that the antibody molecule is administered at a dose of about 100 mg to 1,600 mg, about 100 mg to 1,400 mg, about 100 mg to 1,200 mg, about 100 mg to
1,000 mg, about 100 mg to 800 mg, about 100 mg to 600 mg, about 100 mg to 400 mg, about 100 mg to 200 mg or about 100 mg, about 180 mg or about 200 mg, once every two weeks (Q2W), optionally in which the antibody molecule is administered at a dose of at least about 180 mg Q2W.
143. Method according to any one of claims 139 to141, characterized in that the antibody molecule is administered, for example, intravenously, at a dose of about 5 mg to 100 mg, about 100 mg to 500 mg, about 500 mg to 1,000 mg, about 1,000 mg to 1,500 mg, about 1,500 mg to 2,000 mg, about 2,000 mg to 2,500 mg, about 2,500 mg to 3,000 mg, about 3,000 mg to 3,500 mg or about 3,500 mg to 4,000 mg, for example, at a dose of about 6 mg, about 20 mg, about 60 mg, about 200 mg, about 600 mg, about 1,200 mg, about
2,400 mg, about 3,000 mg, or about 3,600 mg, for example, once a week (QW), once every two weeks (Q2W) or once every four weeks (Q4W), for example, Q2W.
144. Method according to any one of claims 139 to 143, characterized in that the antibody molecule is administered intravenously.
145. Method according to any one of claims 139 to 144, characterized in that the antibody molecule is administered in combination with a second procedure or therapeutic agent.
146. Method according to claim 145, characterized by the fact that the second procedure or therapeutic agent is chosen from one or more among chemotherapy, a targeted anticancer therapy, an oncolytic drug, a cytotoxic agent, a therapy with an immunological basis, a cytokine, surgical procedure, a radiation procedure, an activator of a co-stimulating molecule, an inhibitor of an inhibitory molecule, a vaccine or a cell therapy.
147. Method according to claim 145, characterized by the fact that the second therapeutic agent is chosen from one or more of: 1) a protein C kinase inhibitor (PKC); 2) a heat shock protein inhibitor 90 (HSP90); 3) a phosphoinositide 3-kinase inhibitor (PISK) and / or rapamycin target (mMTOR); 4) a cytochrome P450 inhibitor (for example, a CYP17 inhibitor or a 17 alpha-hydroxylase / C17-20 | iase inhibitor); 5) an iron chelating agent; 6) an aromatase inhibitor; 7) a p53 inhibitor, for example, an inhibitor of a p53 / Mdm 2 interaction; 8) an apoptosis inducer; 9) an angiogenesis inhibitor; 10) an aldosterone synthase inhibitor; 11) a soft receptor inhibitor (SMO); 12) a prolactin receptor inhibitor (PRLR); 13)
a Wnt signaling inhibitor; 14) a CDK4 / 6 inhibitor; 15) a fibroblast growth factor 2 receptor (FGFR2) / fibroblast growth factor 4 (FGFR4) receptor; 16) a macrophage colony stimulating factor inhibitor (M-CSF); 17) an inhibitor of one or more of c-KIT, histamine release, FIt3 (for example, FLK2 / STK1) or PKC; 18) an inhibitor of one or more of VEGFR-2 (for example, FLK-1 / KDR), PDGFRbeta, c-KIT or Raf kinase C; 19) a somatostatin agonist inhibitor and / or a growth hormone release; 20) an anaplastic lymphoma kinase inhibitor (ALK); 21) an insulin-like growth factor receptor inhibitor (IGF-1R); 22) a P-glycoprotein 1 inhibitor; 23) an inhibitor of vascular endothelial growth factor receptor (VEGFR); 24) a BCR-ABL kinase inhibitor; 25) an FGFR inhibitor; 26) a CYP11B2 inhibitor; 27) an HDM2 inhibitor, for example, an inhibitor of HDM2-p53 interaction; 28) a tyrosine kinase inhibitor; 29) a c-MET inhibitor; 30) a JAK inhibitor; 31) a DAC inhibitor; 32) an 11B-hydroxylase inhibitor; 33) an IAP inhibitor; 34) a PIM kinase inhibitor; 35) a Porcupine inhibitor; 36) a BRAF inhibitor, for example, BRAF V600E or wild-type BRAF; 37) an HER3 inhibitor; 38) a MEK inhibitor; or 39) a lipid kinase inhibitor.
148. Method according to claim 145, characterized in that the antibody molecule is administered in combination with a PD-1 inhibitor, optionally in which the PD-1 inhibitor is selected from the group consisting of PDROO01, Nivolumab, Pembrolizumab, Pidilizumab, MEDIO680, REGN2810, TSR-042, PF-O06801591 and AMP-224, optionally where: the PD-1 inhibitor is an anti-PD-1 antibody molecule, in whereas the anti-PD-1 antibody molecule is administered, for example, intravenously, at a dose of about 250 mg to 350 mg,
about 350 mg to 450 mg or about 450 mg to 550 mg, for example, at a dose of about 300 mg or about 400 mg, for example, once every three weeks (Q3W) or once every four weeks (Q4W), for example, at a dose of about 300 mg Q3W or at a dose of about 400 mg Q4W, optionally where:
the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 5 mg to 100 mg, for example, 20 mg, Q2W, and the anti-PD-1 antibody molecule is administered, for example, intravenously, at a dose of about 350 mg to 450 mg, for example, 400 mg, Q4W,
the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 5 mg to 100 mg, for example, 60 mg, Q2W, and the anti-PD-1 antibody molecule is administered, for example, intravenously, at a dose of about 350 mg to 450 mg, for example, 400 mg, Q4W,
the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 100 mg to 500 mg, for example, 200 mg, Q2W, and the anti-PD-1 antibody molecule is administered, for example, intravenously, at a dose of about 350 mg to 450 mg, for example, 400 mg, Q4W,
the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 500 mg to 1000 mg, for example, 600 mg, Q2W, and the anti-PD-1 antibody molecule is administered, for example, intravenously, at a dose of about 350 mg to 450 mg, for example, 400 mg, Q4W,
the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 1000 mg to 1500 mg, for example, 1200 mg, Q2W, and the anti-PD-1 antibody molecule is administered, for example, intravenously, at a dose of about 350 mg to 450 mg, for example, 400 mg, Q4W,
the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 2000 mg to 2500 mg, for example, 2400 mg, Q2W, and the anti-PD-1 antibody molecule is administered, for example, intravenously, at a dose of about 350 mg to 450 mg, for example, 400 mg, Q4W, the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 3000 mg to 3500 mg, for example, 3000 mg, Q2W, and the anti-PD-1 antibody molecule is administered, for example, intravenously, at a dose of about 350 mg to 450 mg, for example, 400 mg, Q4W, or the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 3500 mg to 4000 mg, for example, 3600 mg, Q2W, and the anti-PD antibody molecule -1 is administered, for example, intravenously, at a dose of about 350 mg to 450 mg, for example, 400 mg, Q4W.
149. Method according to claim 145, characterized in that the antibody molecule is administered in combination with an adenosine A2AR antagonist, optionally in which: (i) the adenosine APAR antagonist is selected from the group consisting of PBF509, CPI444, AZD4635, Vipade- nante, GBV-2034 and AB928; or (ii) the adenosine A2AR antagonist is selected from the group consisting of 5-bromo-2,6-di- (1H-pyrazol-1-yl) pyrimidine-4-amine; (S) -7- (5-methylfuran-2-yl) -3 - ((6 - ((((tetrahydrofuran-3-iN) oxy) mMethyl) pyridin-2-yl)] methyl) - 3H- [ 1,2,3] triazolo [4,5-d] pyrimidin-5-amine; (R) -7- (5-methylfuran-2-yl) -3 - ((6 - ((((tetrahydrofuran-3-yl) oxy) Methyl) pyridin-2-iNmethyl) -3H- [1,2 , 3] triazolo [4,5-d] pyrimidin-5-amine or racemate thereof; 7- (5-methylfuran-2-yl) -3 - ((6 - ((((tetrahydrofuran-3-yl) oxy) Methyl) pyridin-2-yl) methyl) -3H- [1,2,3] triazolo [4,5-d] pyrimidin-5-amine; and 6- (2-chloro-
6-methylpyridin-4-yl) -5- (4-fluorophenyl) -1,2,4-triazin-3-amine, optionally where:
the adenosine A2AR antagonist is administered, for example, orally, at a dose of about 20 mg to 60 mg, about 60 mg to 100 mg, about 100 mg to 140 mg, about 140 mg to 180 mg, about 180 mg to 220 mg, about 220 mg to 260 mg, about 260 mg to 300 mg, about 300 mg to 340 mg, about 340 mg to 380 mg, about 380 mg to 480 mg, about 480 mg to 580 mg or about 580 mg to 680 mg, for example, at a dose of about 40 mg, about 80 mg, about 160 mg, about 320 mg, about 480 mg or about 620 mg, for example, once a day (QD), twice a day (BID) or three times a day (TID), for example, BID, optionally where:
the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 5 mg to 100 mg, for example, 20 mg, Q2W, and the adenosine A2AR antagonist is administered, for example, orally, at a dose of about mg to 60 mg, for example, 40 mg, BID,
the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 5 mg to 100 mg, for example, 60 mg, Q2W, and the adenosine A2AR antagonist is administered, for example, orally, at a dose of about 20 mg to 60 mg, for example, 40 mg, BID,
the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 100 mg to 500 mg, for example, 200 mg, Q2W, and the adenosine A2AR antagonist is administered by example, orally, at a dose of about 60 mg to 100 mg, for example, 80 mg, BID,
the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 100 mg to 500 mg, for example, 200 mg, Q2W, and the adenosine A2AR antagonist is administered by example, orally, at a dose of about 140 mg to 180 mg, for example, 160 mg, BID, the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 500 mg a
1,000 mg, for example, 600 mg, Q2W, and the adenosine APAR antagonist is administered, for example, orally, at a dose of about 140 mg to 180 mg, for example, 160 mg, BID, the molecule of anti-CD73 antibody is administered, for example, intravenously, at a dose of about 1000 mg to 1500 mg, for example, 1200 mg, Q2W, and the adenosine A2AR antagonist is administered, for example, in an oral, at a dose of about 140 mg to 180 mg, for example, 160 mg, BID, the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 2000 mg to 2500 mg , for example, 2400 mg, QO2W, and the adenosine A2AR antagonist is administered, for example, orally, at a dose of about 140 mg to 180 mg, for example, 160 mg, BID, the antibody molecule anti-CD73 is administered, for example, intravenously, at a dose of about 2000 mg to 2500 mg, for example, 2400 mg, Q2W, and the adenosine A2AR antagonist is administered, for example, orally, at a dose of about 300 mg to 340 mg, for example, 320 mg, BID, the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 2000 mg at 2500 mg, for example, 2400 mg, Q2W, and the adenosine A2AR antagonist is administered, for example, orally, at a dose of about 480 mg to 580 mg, for example, 480 mg, BID, the molecule of anti-CD73 antibody is administered, for example, intravenously, at a dose of about 2000 mg at
2500 mg, for example, 2400 mg, Q2W, and the adenosine A2AR antagonist is administered, for example, orally, at a dose of about 580 mg to 680 mg, for example, 620 mg, BID,
the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 3000 mg to 3500 mg, for example, 3000 mg, Q2W, and the adenosine A2AR antagonist is administered, for example, orally, at a dose of about 300 mg to 340 mg, for example, 320 mg, BID,
the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 3000 mg to 3500 mg, for example, 3000 mg, Q2W, and the adenosine A2AR antagonist is administered, for example, orally, at a dose of about 480 mg to 580 mg, for example, 480 mg, BID,
the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 3000 mg to 3500 mg, for example, 3000 mg, QO2W, and the adenosine A2AR antagonist is administered, for example, orally, at a dose of about 580 mg to 680 mg, for example, 620 mg, BID,
the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 3500 mg to 4000 mg, for example, 3600 mg, Q2W, and the adenosine A2AR antagonist is administered, for example, orally, at a dose of about 300 mg to 340 mg, for example, 320 mg, BID,
the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 3500 mg to 4000 mg, for example, 3600 mg, Q2W, and the adenosine A2AR antagonist is administered, for example, orally, at a dose of about 480 mg to 580 mg, for example, 480 mg, BID, or the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 3500 mg The
4000 mg, for example, 3600 mg, Q2W, and the adenosine A2AR antagonist is administered, for example, orally, at a dose of about 580 mg to 680 mg, for example, 620 mg, BID.
150. Method according to claim 145, characterized in that the antibody molecule is administered in combination with a PD-1 inhibitor and an adenosine A2AR antagonist, optionally in which: the PD- inhibitor 1 is an anti-PD-1 antibody molecule, where the anti-PD-1 antibody molecule is administered, for example, intravenously, at a dose of about 250 mg to 350 mg, about 350 mg to 450 mg or about 450 mg to 550 mg, for example, at a dose of about 300 mg or about 400 mg, for example, once every three weeks (Q3W) or once at every four weeks (Q4W), for example, at a dose of about 300 mg Q3W or at a dose of about 400 mg Q4W, optionally where: the adenosine A2AR antagonist is administered, for example, orally, at a dose of about 20 mg to 60 mg, about 60 mg to 100 mg, about 100 mg to 140 mg, about 140 mg to 180 mg, about 180 mg to 220 mg, about 220 mg to 260 mg, about 260 mg to 300 mg, about from 300 mg to 340 mg or about 340 mg to 380 mg, for example, at a dose of about 40 mg, about 80 mg, about 160 mg, or about 320 mg, for example, once a day (QD), twice a day (BID) or three times a day (TID), for example, BID, optionally where: the anti-CD73 antibody molecule is administered, for example, intravenously, to a dose of about 5 mg to 100 mg, for example, 20 mg, Q2W, the anti-PD-1 antibody molecule is administered, for example, intravenously, at a dose of about 350 mg to 450 mg, per example, 400 mag, Q4W, and the adenosine AZAR antagonist is administered, for example, orally, at a dose of about 20 mg to 60 mg, for example, 40 mg, BID,
the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 5 mg to 100 mg, for example, 60 mg, Q2W, the anti-PD-1 antibody molecule is administered, by example, intravenously, at a dose of about 350 mg to 450 mg, for example, 400 mg, Q4W, and the adenosine AZAR antagonist is administered, for example, orally, at a dose of about 20 mg to 60 mg, for example, 40 mg, BID,
the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 100 mg to 500 mg, for example, 200 mg, Q2W, the anti-PD-1 antibody molecule is administered, by example, intravenously, at a dose of about 350 mg to 450 mg, for example, 400 mg, Q4W, and the adenosine AZAR antagonist is administered, for example, orally, at a dose of about 60 mg to 100 mg, for example, 80 mg, BID,
the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 100 mg to 500 mg, for example, 200 mg, Q2W, the anti-PD-1 antibody molecule is administered, by example, intravenously, at a dose of about 350 mg to 450 mg, for example, 400 mg, Q4W, and the adenosine AZAR antagonist is administered, for example, orally, at a dose of about 140 mg to 180 mg, for example, 160 mg, BID,
the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 500 mg to 1000 mg, for example, 600 mg, Q2W, the anti-PD-1 antibody molecule is administered, by example, intravenously, at a dose of about 350 mg to 450 mg, for example, 400 mg, Q4W, and the A2AR adenosine antagonist is administered, for example, orally, at a dose of about from 140 mg to 180 mg, for example, 160 mg, BID,
the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 1000 mg to 1500 mg, for example, 1200 mg, Q2W, the anti-PD-1 antibody molecule is administered, by example, intravenously, at a dose of about 350 mg to 450 mg, for example, 400 mg, Q4W, and the A2AR adenosine antagonist is administered, for example, orally, at a dose of about from 140 mg to 180 mg, for example, 160 mg, BID,
the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 2000 mg to 2500 mg, for example, 2400 mg, Q2W, the anti-PD-1 antibody molecule is administered, by example, intravenously, at a dose of about 350 mg to 450 mg, for example, 400 mg, Q4W, and the A2AR adenosine antagonist is administered, for example, orally, at a dose of about from 140 mg to 180 mg, for example, 160 mg, BID,
the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 2000 mg to 2500 mg, for example, 2400 mg, Q2W, the anti-PD-1 antibody molecule is administered, by example, intravenously, at a dose of about 350 mg to 450 mg, for example, 400 mg, Q4W, and the A2AR adenosine antagonist is administered, for example, orally, at a dose of about from 300 mg to 340 mg, for example, 320 mg, BID,
the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 2000 mg to 2500 mg, for example, 2400 mg, Q2W, the anti-PD-1 antibody molecule is administered, by example, intravenously, at a dose of about 350 mg to 450 mg, for example, 400 mg, Q4W, and the A2AR adenosine antagonist is administered, for example, orally, at a dose of about from 480 mg to 580 mg, for example, 480 mg, BID,
the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 2000 mg to 2500 mg, for example, 2400 mg, Q2W, the anti-PD-1 antibody molecule is administered, by example, intravenously, at a dose of about 350 mg to 450 mg, for example, 400 mg, Q4W, and the A2AR adenosine antagonist is administered, for example, orally, at a dose of about from 580 mg to 680 mg, for example, 620 mg, BID,
the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 3000 mg to 3500 mg, for example, 3000 mg, Q2W, the anti-PD-1 antibody molecule is administered, by example, intravenously, at a dose of about 350 mg to 450 mg, for example, 400 mg, Q4W, and the A2AR adenosine antagonist is administered, for example, orally, at a dose of about from 300 mg to 340 mg, for example, 320 mg, BID,
the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 3000 mg to 3500 mg, for example, 3000 mg, Q2W, the anti-PD-1 antibody molecule is administered, by example, intravenously, at a dose of about 350 mg to 450 mg, for example, 400 mg, Q4W, and the A2AR adenosine antagonist is administered, for example, orally, at a dose of about from 480 mg to 580 mg, for example, 480 mg, BID,
the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 3000 mg to 3500 mg, for example, 3000 mg, Q2W, the anti-PD-1 antibody molecule is administered, by example, intravenously, at a dose of about 350 mg to 450 mg, for example, 400 mg, Q4W, and the A2AR adenosine antagonist is administered, for example, orally, at a dose of about from 580 mg to 680 mg, for example, 620 mg, BID, the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose of about 3500 mg to 4000 mg, for example, 3600 mg, Q2W, the anti-PD-1 antibody molecule is administered, for example, intravenously, at a dose of about 350 mg to 450 mg, for example, 400 mg, Q4W, and the adenosine A2AR antagonist is administered, for example, orally, at a dose of about 300 mg to 340 mg, for example, 320 mg, BID, the anti-CD73 antibody molecule is administered, for example, intravenously, at a dose d and about 3500 mg to 4000 mg, for example, 3600 mg, Q2W, the anti-PD-1 antibody molecule is administered, for example, intravenously, at a dose of about 350 mg to 450 mg, for example , 400 mg, Q4W, and the adenosine A2AR antagonist is administered, for example, orally, at a dose of about 480 mg to 580 mg, for example, 480 mg, BID, or the antibody molecule anti-CD73 is administered, for example, intravenously, at a dose of about 3500 mg to 4000 mg, for example, 3600 mg, Q2W, the anti-PD-1 antibody molecule is administered, for example, in an intravenously, at a dose of about 350 mg to 450 mg, for example 400 mg, Q4W, and the A2AR adenosine antagonist is administered, for example, orally, at a dose of about 580 mg at 680 mg, for example, 620 mg, BID.
151. Method according to claim 145, characterized in that the antibody molecule is administered in combination with a PD-L1 inhibitor, optionally in which the PD-L1 inhibitor is selected from the group consisting of FAZO53, Atezolizumab, Avelumab, Durvalumabe and BMS-936559.
152. Method according to claim 145, characterized in that the antibody molecule is administered in combination with a PD-L1 inhibitor and an adenosine A2AR antagonist.
153. Method according to claim 145, characterized in that the antibody molecule is administered in combination with: (i): a CTLA-A4 inhibitor, optionally in which the CTLA-4 inhibitor is Ipilimumab or Tremelimumab; (ii) a TIM-3 inhibitor, optionally wherein the TIM-3 inhibitor is selected from the group consisting of MGB453, TSR-022 and LY3321367; (iii) an LAG-3 inhibitor, optionally wherein the LAG-3 inhibitor is selected from the group consisting of LAG525, BMS-986016, TSR-033, MK-4280 and REGN3767; (iv) a GITR agonist, optionally in which the GITR agonist is selected from the group consisting of GWN323, BMS-986156, MK-4166, MK-1248, TRX518, INCAGN1876, AMG 228 and INBRX-110 ; (v) an anti-CD3 multispecific antibody molecule, optionally wherein the anti-CD3 multispecific antibody molecule is an anti-CD3 x anti-CD123 antibody molecule (for example, XENP14045), or a bispecific antibody molecule anti-CD3 x anti-CD20 (for example, XENP13676); (vi) a cytokine molecule, optionally wherein the cytokine molecule is IL-15 complexed with a soluble form of IL-15 alpha receptor (IL-15Ra);
(vii) a STING agonist; (viii) a macrophage colony stimulating factor (M-CSF) inhibitor, optionally wherein the M-CSF inhibitor is MCS110; (ix) a CSF-1R inhibitor, optionally wherein the CSF-1R inhibitor is BLZ945; (x) an indolamine 2,3-dioxigenase (IDO) and / or 2,3-dioxigenase tryptophan (TDO) inhibitor; (xi) the TGF-B inhibitor; (xii) an oncolytic vaccine; (xiii) a chimeric antigen receptor (CAR) T cell therapy, optionally wherein the CAR T cell therapy is CTLO19; or (xiv) a compound disclosed in Table 18.
154. Method for detecting CD73 in a biological sample or in an individual, characterized by the fact that it comprises (i) putting the sample or the individual (and, optionally, a sample or reference individual) in contact with the antibody molecule or composition, as defined with any one of claims 1 to 94 under conditions that allow interaction of the antibody or composition molecule and CD73 to occur, and (ii) detecting the formation of a complex between the antibody molecule or composition and the sample or the individual (and, optionally, the sample or reference individual).

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法律状态:
2021-11-03| B350| Update of information on the portal [chapter 15.35 patent gazette]|

优先权:

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申请号 | 申请日 | 专利标题

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US201762523481P| true| 2017-06-22|2017-06-22|

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US62/523,481|2017-06-22|

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US201862636510P| true| 2018-02-28|2018-02-28|

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US62/636,510|2018-02-28|

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PCT/US2018/038775|WO2018237157A1|2017-06-22|2018-06-21|Antibody molecules to cd73 and uses thereof|

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