• 专利附录
  • 专利说明
  • 权利要求
  • 类似技术
  • 同族专利
  • 引用文献
  • 法律状态
  • 优先权
    • 专利摘要:
    In the profragment region of prorenin, a method for screening a substance capable of inhibiting the activation of prorenin using the regulation ability of prorenin activation by the interaction between proteins, and the profragment region of prorenin Prorenin activation inhibitors having a donation that inhibits the activation of prorenin based on protein-to-protein interactions, as well as suppressors, organ hypertrophy inhibitors, and arterial thickening inhibitors containing the prorenin activation inhibitors as active ingredients. to provide.
    公开号:KR20020097217A
    申请号:KR1020027013545
    申请日:2001-04-09
    公开日:2002-12-31
    发明作者:유이치 이쉬다；스즈키푸미아키；무라카미가즈오
    申请人:유이치 이쉬다；
    IPC主号:
    专利说明:

    Coercion {Hypotensors}
    [2] Renin, angiotensin, and aldosteron systems are known to be involved in the rise of blood pressure (Dong-Yae University Magazine 60 (4): 342-350,1990). That is, angiotensinogen secreted from the liver is converted to angiotensin I (hereinafter referred to as AI) by the action of renin-derived renin, which in turn is caused by the action of angiotensin converting enzyme (ACE). To angiotensin II (hereinafter referred to as AII). AII directly acts on blood vessels, causing vasoconstriction, and also acting on the adrenal cortex, promoting biosynthesis and secretion of aldosterone, and raising blood pressure by causing the storage of sodium and water.
    [3] For this reason, as a treatment for the recent increase in blood pressure, drugs acting on the Lenin-Angiotensin system have become the newest forms of suppressors, and ACE inhibitors have been put into practical use. At the same time, renin inhibitors have been studied as a coercive agent, but development has been stopped because of the side effects. ACE inhibitors are specific inhibitors of angiotensin converting enzymes and are generally known as captopril. However, since ACE inhibitors also have side effects such as flatulence, and cannot be clinically effective to the extent of enzymatic inhibitory effects in vitro, combination therapy with renin-angiotensin inhibitors having different mechanisms of action is attempted. It is becoming. Therefore, the necessity of the effective antagonist, the vascular thickening inhibitor, and the long-term hypertrophy inhibitor which has the effect other than the conventional one reported or used is increasing.
    [4] Lenin is biosynthesized from pre-pro-renin consisting of 406 amino acids, its precursor in the kidney, and 23 amino acids at the N-terminus are cleaved to produce prorenin. It is cleaved from the end to form renin consisting of 340 amino acids. Lenin is a protease that hydrolyzes angiotensinogen specifically to produce AI. However, since prorenin, a precursor of renin, does not usually exhibit its enzymatic activity, despite the fact that the amount of prorenin in the blood is about 10 times that of renin, in the renin-angiotensin system, agonists are hydrolyzed by renin or prorenin. It is believed to be the produced Lenin.
    [5] The inventors have previously recognized the 43 amino fragments of the N-terminus (hereinafter referred to as 'profragments' or 'pf'), where the inactive renin precursor prorenin is cleaved when renin is produced from prorenin. The experiment using anti-human prorenin pf antibody showed that when combined with in vitro antibody to form immunocomplex, the primary structure does not change under physiological conditions and it expresses protein function, that is, enzyme activity (renin activity) non-enzymatically. (Patent No. 10-279600, U.S. Patent No. 5945512). To date, as a means of activating prorenin, it is converted to lenin by proteases, and changes in primary structure in acidic and cold conditions. Activation, etc. ( Nature , 288, 702-705, 1980; J. Biol. Chem. , 262, 2472-2477, 1987; Clin. Chem. , 37, 1811-1819, 1991; J. Biol. Chem. , 267, 11753-11759, 1992), and small molecules A method of binding a renin inhibitor to an enzyme active site hidden deep in the three-dimensional structure of prorenin and making it open ( J. Biol. Chem. , 267, 22837-22842, 1992) and the like are known. However, the mechanism of prorenin activation in vivo as well as its action is not clearly identified.
    [1] The present invention relates to hypertensives, organ hypertrophy inhibitors and arterial thickening inhibitors and their screening methods.
    [6] In light of the above circumstances, the present inventors conducted a study to administer a pf antibody capable of activating prorenin or a partial peptide derived from prorenin profragment, which is an antigen of the antibody, to a model animal having a high blood pressure condition. It was found that the effect, long-term hypertrophy effect, and camellia thickening effect can be obtained. In other words, by discovering that activation of prorenin can be controlled in vivo by inhibiting interprotein interactions in the profragment region of prorenin, for the first time that prorenin can be activated in vivo. It proved. Based on the findings described above, the present inventors have confirmed that by using a substance that inhibits the activation of prorenin in vivo, it is possible to provide an antihypertensive agent, an organ hypertrophy inhibitor, an arterial thickener, and a method for selecting them. The invention has been completed.
    [7] That is, the aspect of this invention,
    [8] 1) Selecting a substance capable of controlling the prorenin activation by using the regulation of prorenin activation, which is not accompanied by a change in the primary structure caused by the interaction between proteins in the profragment region of the prorenin, as an indicator How to,
    [9] 2) a method for screening substances capable of controlling the activation of prorenin using the regulation of prorenin activation by antibodies to the profragment region of prorenin,
    [10] 3) The screening method according to the above 1) or 2), wherein the screening method comprises a substance designed based on at least three amino acid sequence information of the amino acid sequence of the profragment region of prorenin;
    [11] 4) The screening method of 1) or 2) above, wherein the amino acid sequence of the profragment region of prorenin is at least three amino acid sequences of the 1st to 19th amino acid sequence of the N-terminus or the 27th to 41st amino acid sequence Screening methods for the selection of substances designed on the basis of information,
    [12] 5) In the selection method of 1) or 2), a substance designed based on at least three amino acid sequence information among the fifth to nineteenth amino acid sequences of the amino acid sequence N-terminus of the profragment region of prorenin is selected. Screening method to target,
    [13] 6) In the screening method of any one of 3) to 5), the screening method wherein the substance to be screened is a peptide,
    [14] 7) The screening method of any one of 3) to 5) wherein the substance to be screened is a low molecular compound,
    [15] 8) The selection method of any one of 1) to 7), wherein the prorenin is human prorenin,
    [16] 9) The selection method of any one of 1) to 7), wherein the prorenin is rat prorenin,
    [17] 10) a prorenin activation control substance having a function of controlling prorenin activation that is not accompanied by changes in primary structure caused by interprotein interactions in the profragment region of prorenin,
    [18] 11) a function of controlling prorenin activation that is not accompanied by a change in primary structure caused by interprotein interaction in the profragment region of prorenin selected by any one of 1) to 9). Having prorenin activation regulators,
    [19] 12) a prorenin activation control substance having a function of inhibiting prorenin activation that is not accompanied by a change in primary structure caused by protein interactions in the profragment region of prorenin,
    [20] 13) a prorenin activation control substance that has a function of inhibiting prorenin activation that does not involve primary structure changes caused by protein interactions in the profragment region of prorenin, and at the same time does not have renin inhibitory activity,
    [21] 14) inhibits prorenin activation, which is selected by the method of any one of 1) to 9) and which does not involve changes in primary structure caused by interprotein interactions in the profragment region of prorenin. Having prorenin activation regulators,
    [22] 15) inhibits prorenin activation which is selected by any one of 1) to 9) and which is not accompanied by changes in primary structure caused by interprotein interactions in the profragment region of prorenin. Prorenin activation control substance, which does not have renin inhibitory activity,
    [23] 16) a partial sequence selected from the amino acid sequence of the profragment region of prorenin, which has a function of inhibiting prorenin activation which is not accompanied by a change in primary structure caused by the interaction between proteins in the profragment region of prorenin Prorenin activation control material consisting of a peptide having a peptide equivalent thereto or
    [24] 17) has a function of inhibiting prorenin activation that does not involve changes in primary structure caused by interprotein interactions in the profragment region of prorenin and is selected from the amino acid sequence of the profragment region of prorenin. Prorenin activation modulators consisting of peptides with 3 to 10 subarrays,
    [25] 18) has a function of inhibiting prorenin activation that is not accompanied by changes in primary structure caused by interprotein interactions in the profragment region of prorenin, and is selected from the amino acid sequence of the profragment region of prorenin. Prorenin activation modulators consisting of peptides with 3 to 8 subarrays,
    [26] 19) has a function of inhibiting prorenin activation which is not accompanied by changes in primary structure caused by interprotein interactions in the profragment region of prorenin, and is selected from the amino acid sequence of the profragment region of prorenin. Prorenin activation modulators consisting of peptides with 3 to 6 subarrays,
    [27] 20) from the peptides set forth in SEQ ID NOS: 7-14, having the function of inhibiting prorenin activation that does not involve changes in primary structure caused by interprotein interactions in the profragment region of prorenin; Prorenin activation modulators composed of selected peptides,
    [28] 21) has the function of inhibiting prorenin activation that does not involve changes in primary structure caused by interprotein interactions in the profragment region of prorenin; A prorenin activation modulator consisting of a peptide selected from the peptides set forth in Nos. 15 to 20,
    [29] 22) It is a low molecular compound designed based on the amino acid sequence information of the profragment region of prorenin, and inhibits prorenin activation which is not accompanied by the change of primary structure caused by the interaction between proteins in the profragment region of prorenin. Prorenin activation regulator, which has a function to
    [30] 23) In any of the prorenin activation control substance of 16) to 22), it inhibits prorenin activation which is not accompanied by primary structural changes caused by interprotein interaction in the profragment region of prorenin. Prorenin activation regulators that have a function and at the same time have no renin inhibitory activity,
    [31] 24) The prorenin activation according to any one of 12) to 23) above, wherein the effect of inhibiting prorenin activation is due to antagonism of interprotein interaction in the profragment region of prorenin. Control Material,
    [32] 25) a prorenin activation control substance, which inhibits prorenin activation in vivo by elimination of prorenin due to an antigenic antibody response to the profragment region of prorenin,
    [33] 26) prorenin activation can be inhibited in vivo by elimination of prorenin due to antigen antibody reaction to the profragment region of prorenin, and a prorenin activation control substance consisting of an antibody to the profragment region of prorenin,
    [34] 27) an antihypertensive agent containing at least one of any of the prorenin activation inhibitors of 12) to 26) as an active ingredient,
    [35] 28) The pressure reducing agent according to 27), wherein the pressure reducing agent exhibits a lowering rate of about 10% or more within at least 12 hours after in vivo administration,
    [36] 29) The antagonist of 27), wherein the antagonistic effect is selectively exerted only on the body of a hypertensive patient,
    [37] 30) long-term hypertrophy inhibitor containing at least one kind of prorenin activation inhibitor in any one of 12) to 26) as an active ingredient,
    [38] 31) The organ hypertrophy inhibitor according to 30), wherein the organ hypertrophy inhibitor inhibits organ hypertrophy in the heart and / or kidney,
    [39] 32) arterial thickening inhibitor containing at least one kind of any of the prorenin activation inhibitors of 12) to 26) as an active ingredient,
    [40] 33) a method for treating hypertension using any of the coercive agents of 27) to 29),
    [41] 34) A method of treating heart failure or kidney failure using the long-term hypertrophy inhibitor of 30) or 31) above,
    [42] 35) may be a method for treating a disease associated with vascular thickening using the arterial thickening inhibitor described in 32).
    [43] Hereinafter, the present invention will be described in detail, and the technical and scientific terms used herein have the meanings that can be generally understood by those skilled in the art unless otherwise defined. will be. This specification refers to various methods known to those skilled in the art, and does not completely describe these methods by citing materials such as publications which disclose the known methods cited.
    [44] [Effect by Inhibiting Prorenin Activation in Vivo]
    [45] The present inventors have previously used anti-human pf antibodies to activate prorenin in an open structure exhibiting enzymatic activity without changing its primary structure in vitro by specific antibodies to amino acid fragments of the pf region of prorenin. (Japanese Patent Application Laid-Open No. 10-279600, US Patent No. 5945512). Based on these facts, an antibody (anti-rat) that specifically recognizes a rat pf peptide that is the same site in the sequence of amino acid sequence and peptide (human pf peptide) in the pf region of human prorenin in the primary structure of rat prorenin pf antibody) were prepared and administered to hypertensive spontaneous hypertensive rats (SHR). As a result, although the anti-rat pf antibody can activate rat prorenin in vitro without changing its primary structure, it showed a sustained suppressive effect in vivo as experimentally shown. In general, antibodies bind to antigens in the blood to form antigen antibody complexes, which are then removed from the blood by the immune system. In other words, it is thought that the coercive effect occurs because prorenin or activated prorenin is removed from blood by the binding of anti-pf antibodies. In addition, it is thought that the binding effect of prorenin or activated prorenin to its action site is inhibited by anti-pf antibody binding to prorenin or activated prorenin in blood.
    [46] In addition, the present inventors synthesized a partial peptide derived from rat prorenin profragment, which is an antigen of the anti-rat pf antibody capable of activating rat prorenin in vitro, and administered to SHR. These peptides can antagonize rat prorenin activation in vitro by anti rat pf antibodies, such as human pf peptides inhibit human prorenin activation in vitro by anti human pf antibodies. That is, these peptides do not involve changes in the primary structure of rat prorenin caused by protein-to-protein interactions, but may inhibit activation accompanying changes in higher structures. Administration of these peptides to the SHR was able to obtain sufficient coercive effect within at least 12 hours after administration as described below. However, blood pressure did not change even when these peptides were administered to normal rats. As shown in the experimental examples described below, the behavior of these peptides over time was different from that of the Lenin inhibitor H6137 (manufactured by Sigma) or the Lenin neutralizing antibody. Was not caused by renin inhibition. In addition, these peptides have been found to inhibit the hypertrophy of the heart and kidney, and especially to inhibit the thickening of arteries, that is, vascular remodeling, when administered to SHR for 14 consecutive days. These effects obtained by the administration of the peptide are presumed to be due to the phenomenon that the binding of rat prorenin to its action site is inhibited by the peptide in vivo.
    [47] From the above results, in SHR, prorenin interacts with prorenin-binding protein to express enzymatic activity, thereby activating the renin-angiotensin system, thereby causing hypertension. Prorenin's pf peptide and anti-pf peptide antibody Has been shown to exhibit a hypertensive action against hypertension by activating the Lenin-Angiotensin system. Prorenin activation in vivo by such involvement of prorenin in hypertension and protein interactions in the profragment region of prorenin was first identified here. In addition, prorenin activation control substances that can inhibit the interaction between the protein and the interaction between prorenin or activated prorenin and its action site are accompanied by structural changes caused by the interprotein interaction of prorenin. It was found to be useful as a hypertrophy inhibitor and an arterial thickener.
    [48] Rat prorenin pf peptides have the same amino acid residues as human prorenin pf peptides but different constituent amino acids. Despite this fact, the human pf peptide, which is an epitope peptide of an anti-human pf antibody capable of activating human prorenin in vitro as described above, and the rat pf peptide which is the same site in the amino acid sequence of the primary structure And the specific antibody to the pf peptide showed a coercive effect in the rat hypertension model. Therefore, it can be said that there is no species difference in the pharmacological action of prorenin involved in hypertension, the structural site involved in prorenin activation, and the mechanism of activation thereof. That is, the coercive effect of the peptide was recognized in rat hypertension disease model animals, but the same effect can be expected in human hypertension by selecting an appropriate peptide in consideration of the amino acid sequence of the pf peptide of human prorenin.
    [49] Moreover, when the inactive enzyme precursor interacts with the anti-pf peptide specific antibody in vitro and expresses the enzyme activity, that is, the protein function, as described above, the mechanism is conserved across species, so that the pf peptide, the pf peptide, Antibodies, condition model animals, and the like are useful as screening methods for screening therapeutic agents for conditions by protein expression.
    [50] [Selection method of prorenin activation control substance]
    [51] The method for screening a substance capable of controlling prorenin activation according to the present invention is characterized by using the regulation of prorenin activation by the interaction between proteins in the prorenin profragment region as an index. Herein, prorenin activation by interprotein interactions in the profragment region of prorenin is caused by interprotein interactions in the pf region and does not involve changes in the primary structure of prorenin, but changes in higher order structures. It refers to the opening of the enzymatic activity site involved. Therefore, prorenin activated by interprotein interaction has the same amino acid sequence as the primary structure of prorenin before activation, but can exhibit enzymatic activity as renin. Examples of proteins having protein-to-protein interactions in the pf region include antibodies that recognize and activate pf peptides, proteins that act on prorenin or activated prorenin in vivo, and blood proteins obtained by binding to and activating prorenin. For example.
    [52] The method of selecting the prorenin activation control substance can be constructed using a drug screening system known per se. For example, by measuring the binding of prorenin to the anti-pf antibody using an anti-pf antibody capable of activating prorenin and prorenin, a substance that inhibits prorenin activation or a substance that inhibits enzymatic activity of prorenin Can be screened. Of course, the sorting method is not limited to these. In addition, as described above, since the constituent amino acids of the amino acid sequence of the prorenin pf region are different from species to species, selection of a human prorenin activating control agent using an antibody against human prorenin and, for example, a human prorenin pf peptide is recommended. It can be said that it is preferable.
    [53] Specifically, for example, antibodies that can activate human prorenin by specifically recognizing human prorenin and human prorenin pf peptides prepared according to Japanese Patent Application Laid-Open No. Hei 10-279600 and U.S. Patent No. 5945512 are used. To build a screening system.
    [54] As a substance to be screened in the method for screening a prorenin activating control substance, a candidate substance to be screened for use in a drug screening system known in the art such as a low molecular weight compound, a natural compound, or a peptide can be widely used. .
    [55] In addition, peptides and small molecule compounds designed on the basis of amino acid sequence information of the pf region of prorenin, particularly human prorenin, may also be selected.
    [56] The amino acid sequence of the pf region of human prorenin is known and is equal to the 43 amino acid sequence (SEQ ID NO: 1 of SEQ ID NO) listed in Japanese Patent Laid-Open No. 8-285852. The amino acid sequence of the pf peptide of prorenin is useful information for the design of the substance which selects a prorenin activation control substance. In particular, at least 3 of the 1st to 19th (pf1 to pf19), especially the 5th to 19th (pf5 to pf19), and the 27th to 41st (pf27 to pf41) amino acids from the N-terminus of the prorenin pf region. Information of the dog, more preferably at least four consecutive amino acid sequences, is useful for the design of prorenin activation regulators. Examples of the amino acid sequence of human-derived pf1-19, human-derived pf5-19, human-derived pf27-41, rat-derived pf1-19, rat-derived pf5-19 and rat-derived pf27-41 are shown in SEQ ID NOs: 2, 3, 4, 6, 7, and 8 are shown. Hereinafter, the Mth to Nth peptides from the N-terminus of the pf region are described as pfM-N.
    [57] As described above, although the amino acid sequence of the prorenin pf region is different, the number of amino acid residues is the same, but the constituent amino acids are different. In the experimental example, the amino acid sequence (SEQ ID NO: 5) of the pf peptide of rat prorenin was exemplified as useful information for designing a target substance for selecting a prorenin activation regulator. In the experimental example, pf1-11 (SEQ ID NO: 9 of the Sequence Listing), pf5-19 (SEQ ID NO: 7 of the Sequence Listing), pf1-4 (SEQ ID NO: 10 of the Sequence Listing), p1, which are the amino acid sequence fragments of the rat prorenin pf region -7 (SEQ ID NO: 11), pf5-11 (SEQ ID NO: 12), pf12-19 (SEQ ID NO: 13), pf11-15 (SEQ ID NO: 14 of the Sequence Listing), and pf27-41 (SEQ ID NO: 8 of the Sequence Listing) was synthesized, and the coercive effect, long-term hypertrophy inhibitory effect, and arterial hypertrophy effect by inhibition of prorenin activation were confirmed, but are not necessarily limited to the arrangement. Those skilled in the art can synthesize at least three peptides by the means known per se, and design and synthesize the substances to be screened together with the synthetic peptides.
    [58] In addition, a specific antibody to the human pf peptide which is the epitope of the anti-human pf antibody capable of activating human prorenin in vitro and the rat pf peptide which is the same site in the amino acid sequence of the primary structure is a rat hypertension condition as described above. Since the model exhibits a coercive effect, it can be easily estimated that a human prorenin pf peptide having the same amino acid sequence in the primary structure as the rat pf peptide has the same effect in humans. As one of the preferable aspects, the information for designing the substance which selects the pf peptide of human prorenin is used for selection.
    [59] The substance to be screened can be provided for selection if the peptide is appropriately designed based on the amino acid sequence information of the prorenin pf region as described above. The amino acids constituting the peptide need not necessarily be identical to the amino acid sequence of the prorenin pf region, and if they have the same function as the peptide, a mutation, such as deletion, substitution, addition, insertion or the like, appropriately induced in the amino acid sequence of the peptide is induced. A peptide may be sufficient (henceforth an equivalent peptide).
    [60] Further, the substance to be selected may be a low molecular compound obtained by drug design based on the structural complementarity of information based on the secondary structure and / or tertiary structure of the amino acid sequence of the pf region.
    [61] Through the above method, a substance capable of inhibiting activation of human prorenin by an anti-human pf peptide antibody can be obtained. In addition, it accumulates information based on peptides having subarrays selected from the amino acid sequence of the pf region, known coercive agents, and the like, and suppresses the coercive effect, long-term hypertrophy effect, and arterial thickening effect among substances obtained by experiments using animal models. The substance having can be selected.
    [62] [Prorenin Activation Control Substances]
    [63] Thus, the present invention also provides a prorenin activation control substance capable of controlling prorenin activation based on interprotein interactions in the pf region of prorenin.
    [64] One aspect of the invention is a prorenin activation modulator capable of controlling human prorenin activation.
    [65] Prorenin activation control agent according to the present invention is a substance that competitively inhibits the interaction between proteins that activate prorenin, a substance that inhibits the binding of prorenin or activated prorenin antagonistically, prorenin Or a substance having an antigen-antibody reaction against activated prorenin, or a substance that acts on the site of action of prorenin or activated prorenin and inhibits the action of prorenin or activated prorenin.
    [66] It is a preferred aspect of the present invention that the prorenin activation regulator is a prorenin activation inhibitor that has a function of controlling prorenin activation based on interprotein interactions in the pf region of prorenin and at the same time does not have renin inhibitory activity. Is one of. That is, the prorenin activation control agent is a mechanism of action, and does not show a direct inhibitory action on renin. Therefore, it is possible to inhibit only the activity of prorenin activating the disease state without inhibiting the renin-angiotensin system, which plays an important role in the life support mechanism of the living body, such as a renin inhibitor. This is an important feature of the prorenin activation modulator according to the present invention.
    [67] Specific examples include a peptide having a subarray selected from the amino acid sequence of the prorenin pf region, an equivalent peptide of the peptide, a low molecular compound designed from information on the amino acid sequence of the prorenin pf region, an antibody to the prorenin pf region, and the like. It is not limited to these.
    [68] In one preferred embodiment, the peptide is a peptide having a subarray selected from the amino acid sequence of the human prorenin pf region or an equivalent peptide of the peptide. For example, the following peptide can be mentioned as a peptide derived from the amino acid sequence of a human prorenin pf region.
    [69] hp1: human prorenin pf1-11 (SEQ ID NO: 15 of the Sequence Listing)
    [70] hp2: human prorenin pf5-11 (SEQ ID NO: 16 in Sequence Listing)
    [71] hp3: human prorenin pf5-19 (SEQ ID NO: 3 in Sequence Listing)
    [72] hp4: human prorenin pf1-4 (SEQ ID NO: 17 in Sequence Listing)
    [73] hp5: human prorenin pf1-7 (SEQ ID NO: 18 in Sequence Listing)
    [74] hp6: human prorenin pf12-19 (SEQ ID NO: 19 in Sequence Listing)
    [75] hp7: human prorenin pf11-15 (SEQ ID NO: 20 of the Sequence Listing)
    [76] hp8: human prorenin pf27-41 (SEQ ID NO: 4 in Sequence Listing)
    [77] In the control of prorenin activation, one of these peptides may be used, or two or more types of peptides may be mixed and used.
    [78] In addition, as one preferred embodiment, the antibody is an antibody against a human prorenin pf region. For example, the anti-human pf antibody obtained using the following peptide derived from the amino acid sequence of human prorenin pf region as an antigen can be illustrated.
    [79] hp9: human prorenin pf1-15 (SEQ ID NO: 21 in Sequence Listing)
    [80] hp10: human prorenin pf18-30 (SEQ ID NO: 22 in Sequence Listing)
    [81] hp11: human prorenin pf30-41 (SEQ ID NO: 23 of the Sequence Listing)
    [82] In the control of prorenin activation, one type of antibody may be used, or two or more types of antibodies may be mixed and used. When the antibody is used in medicine as a prorenin activation control substance, the antibody is preferably a monoclonal antibody obtained by a known means, and moreover, the antibody is preferably prepared as a humanoid antibody. For the production of humanized antibodies, methods known per se can be used ( J. Immunol. Methods , 100, 5-40, 1987).
    [83] [Forced agent]
    [84] As one aspect of the present invention, there is provided an antihypertensive agent containing at least one kind of the prorenin activation control substance as an active ingredient. The coercive agent exhibits a sufficient coercive effect at least within 12 hours after administration in vivo and is extremely long in duration. For example, in experiments with hypertensive model rats, systolic blood pressure (SBP) values (%) showed a coercive rate of at least 10% within 12 hours after administration. Although this value is not absolutely high as a coercive agent, it can be seen by those skilled in the art that a high effect can be obtained in comparison with the usual case in consideration of the dosage, administration time, and / or administration method.
    [85] On the other hand, the in vivo effect of the renin inhibitor, a conventional antagonist, appears immediately after administration and thereafter is lost in minutes. In addition, the in vivo effect of ACE inhibitors lasts 6-8 hours after administration but then disappears. Therefore, considering the improvement of hypertension, the long-term sustained suppression effect of the suppressor according to the present invention can be said to be a very useful feature.
    [86] In addition, since the antihypertensive agent according to the present invention generates a hypertensive effect through a mechanism different from existing antihypertensive drugs, for example, ACE inhibitors or renin inhibitors, the antihypertensive agent may be used in combination with conventional antihypertensive therapy methods in the treatment of hypertension. .
    [87] Moreover, the coercive agent can only be exerted in hypertensive patients whose coercive effect is selectively. In comparative experiments between normal rats and hypertensive model rats, it was demonstrated that the antagonist only works in hypertensive model rats. That is, the hypertensive agent of the present invention is useful for the treatment of hypertension.
    [88] [Long-term antihypertensive]
    [89] The present invention also provides a novel long-term hypertrophy inhibitor containing at least one kind of the prorenin activation inhibitor as an active ingredient. Long-term hypertrophy inhibitor according to the present invention can suppress long-term hypertrophy in the heart and / or kidney. Therefore, the long-term hypertrophy inhibitor can be expected to have an effect on heart failure or kidney failure.
    [90] [Arterial thickening inhibitor]
    [91] The present invention provides a novel arterial thickening inhibitor containing at least one kind of the prorenin activation inhibitor as an active ingredient. Arterial thickening inhibitor according to the present invention can suppress the thickening of pulmonary arteries, femoral arteries, that is, vascular remodeling. Therefore, the arterial thickening inhibitor is effective for diseases involving vascular thickening, such as arteriosclerosis, that is, vascular remodeling.
    [92] [Shoemaking]
    [93] According to the physical properties of the peptide or the low molecular weight compound may be applied to the formulation of the hypertensive agent, organ hypertrophy inhibitor and arterial thickening inhibitor. For example, formulation methods such as tablets, capsules, aqueous solutions, ethanol solutions, ribosomes, fat emulsions, cyclodextrins and the like may be used.
    [94] Powders, pills, capsules and tablets include excipients such as lactose, glucose and mannitol, disintegrants such as starch, sodium alginate, magnesium stearate and talc ( lubricants such as talc), binders such as polyvinyl alcohol, hydroxy propyl cellulose and gelatin, surfactants such as fatty acid esters, and plasticizers such as glycerin Can be prepared. Individual pharmaceutical carriers are used to produce tablets or capsules.
    [95] Suspensions can be prepared using water, sugars such as sucrose, sorbitol, fractose, glycols such as polyethylene glycol (PEG: polyetghylene glycol), and oils.
    [96] Injectable solutions can be prepared using a carrier consisting of a saline solution, a glucose solution or a mixture of saline and glucose solution.
    [97] For example, ribosomalization is performed by adding a solution in which the substance is dissolved in a solvent (ethanol, etc.) to a solution in which a phospholipid is dissolved in an organic solvent (chloroform, etc.), followed by removing the solvent, and adding a phosphate buffer solution to the shake and sonication. And after centrifugation can be carried out by the process of filtration to recover the supernatant.
    [98] Fat emulsification is, for example, by mixing and heating a substance, an oil component (vegetable oil such as soybean oil, sesame oil, olive oil, medium chain triglycerid oil (MCT), etc.), an emulsifier (phospholipid, etc.) After that, the required amount of water can be added and emulsified and homogenized using an emulsifier (such as a homogenase, for example, a high pressure injection type or an ultrasonic type). It is also possible to lyophilize it. That is, when emulsifying, an emulsifying aid may be added, and examples of the emulsifying aid include glycerin and sugars (for example, glucose, sorbitol, and fructose).
    [99] Cyclodextrin inclusion can be performed, for example, by adding a solution in which cyclodextrin is dissolved in water or the like to a solution in which the substance is dissolved in a solvent (ethanol, etc.), and then filtering and sterilizing drying the precipitate that has been cooled and precipitated. In this case, the cyclodextrin to be used can be appropriately selected from cyclodextrins (α, β, and γ types) having different pore diameters depending on the size of the substance.
    [100] The dosage of the hypertensive agent, organ hypertrophy inhibitor or arterial thickener can be appropriately selected according to the condition, sex, age and weight of the patient. As the route of administration, either oral or parenteral can be used. One preferred embodiment is oral administration in the form of oral preparations. As the dosage, about 1-1,000 micrograms can be illustrated per day.
    [101] Hereinafter, the present invention will be described in more detail with reference to Examples, but the present invention is not limited thereto, and various applications can be made without departing from the technical spirit of the present invention.
    [102] Example 1 Preparation of Antibodies Recognizing Peptides from the Rat Prorenin pf Region
    [103] Pf peptides of SEQ ID NO: 24 (SEQ ID NO: 24), pf18-30 (SEQ ID NO: 25), and pf30-41 (SEQ ID NO: 26 of SEQ ID NO: 26), which are pf peptides of rat prorenin According to the method described in Japanese Patent Application Laid-Open No. 10-2769600 and U.S. Patent No. 5945512, the oligopeptides were synthesized according to the method. Antibodies Ab1, Ab2, and Ab3 were prepared. Specifically, the oligopeptide, which is an antigen, was immunized with rabbits of New Zealand White species together with an aschwant to obtain antiserum. The obtained antiserum was purified according to the method of the present publication.
    [104] Ab1: an antibody that specifically recognizes pf1-15
    [105] Ab2: an antibody that specifically recognizes pf18-30
    [106] Ab3: an antibody that specifically recognizes pf30-41
    [107] In addition, these antibodies were equally mixed and called Abmix. In vitro prorenin activation by these antibodies was examined by the method described in the publication, and it was confirmed that these antibodies can activate prorenin.
    [108] Example 2 Preparation of Peptides from the Rat Prorenin pf Region
    [109] The following peptide derived from the arrangement of the pf region of rat prorenin was synthesized according to a conventional method by the solid phase method.
    [110] p1: pf1-11 (SEQ ID NO: 9 in Sequence Listing)
    [111] p2: pf5-19 (SEQ ID NO: 7 in the Sequence Listing)
    [112] p3: pf27-41 (SEQ ID NO: 8 in Sequence Listing)
    [113] p1a: pf1-4 (SEQ ID NO: 10 of the Sequence Listing)
    [114] p1b: pf1-7 (SEQ ID NO: 11 in Sequence Listing)
    [115] p1c: pf5-11 (SEQ ID NO: 12 in Sequence Listing)
    [116] p2a: pf12-19 (SEQ ID NO: 13 in Sequence Listing)
    [117] p2b: pf11-15 (SEQ ID NO: 14 in Sequence Listing)
    [118] In addition, equal mixing of p1, p2, and p3 was called Pmix.
    [119] Experimental Example 1 Examination of the Coping Effect by Anti-pf Peptide Antibody and pf Peptide
    [120] Three groups of SHR male rats (12-14 weeks old, 258-310 g body weight), which are hypertensive model animals, were used. Abmix prepared in Example 1, Pmix dissolved in physiological saline prepared in Example 2 and anti-renin neutralizing antibodies and captopril as comparative test drugs were 0.44 mg / kg, 0.6 mg / kg, 0.12 mg / g, and 100 mg /, respectively. It was administered intravenously at a dose of kg. Blood pressure measurement was performed by connecting the canoes inserted in the total carotid and external carotid arteries to a pressure transducer (P23XL, Gould Electrics), to a pressure processor signal conditioner (Gould Electrics), and to 1 hour after administration of the test substance. Every minute thereafter, measurements were made at an hourly interval and recorded on a thermal array recorder (RS3400, Gould Electrics).
    [121] The anti-renin neutralizing antibody, a comparative test drug, showed no blood pressure drop until 6 hours after administration, and 11% and 14% of blood pressure lowering effects at 6 hours and 24 hours, respectively. Pmix and Abmix showed continuous blood pressure drop with up to 9% and up to 7% of blood pressure reduction after administration. The positive control captopril showed a maximum blood pressure drop (up to 31%) up to 6 hours after administration, and at the end of the experiment at 24 hours, no blood pressure drop action was confirmed.
    [122] As a result, it was found that Pmix and Abmix had a time-dependent stepping effect different from capenpril which is an anti-renin neutralizing antibody and an ACE inhibitor. That is, it was found that Pmix and Abmix have a sustained blood pressure lowering ability for a very long time immediately after administration. In addition, it could be estimated from this point that the suppressive effect of Pmix and Abmix was not due to inhibition of renin activity.
    [123] Experimental Example 2
    [124] On the basis of the data obtained in Experiment 1, the dose was increased for Pmix (2 mg / kg) and tested in the same manner as in Experiment 1 using 5 SHR. Saline was used as a control. As a result, 11% of transient hypotension was recognized immediately after the administration, and after 5 minutes of administration, it was recovered, but then the blood pressure was gradually lowered again and confirmed up to 9% of hypotension. Significant differences of 1 to 6 hours were identified. That is, it was confirmed that Pmix has a bilayer blood pressure lowering action.
    [125] Experimental Example 3
    [126] For p1 (pf1-11), p2 (pf5-19), and p3 (pf27-41) prepared in Experimental Example 2, experiments were carried out in the same manner as in Experimental Example 1 using two or four SHR. As a result, when P1 was administered 2 mg / kg, a transient blood pressure increase of about 6% was confirmed immediately after the administration, and the continuous effect was confirmed up to 24 hours. In addition, in the case of administration of 10 mg / kg, blood pressure drop of about 7% appeared after 6 hours of administration, and the continuous effect was confirmed up to 24 hours at up to 10%. When 2 mg / kg was administered, p2 showed a transient blood pressure drop of 35% immediately after administration, and recovered 5 minutes after administration, and continued to lower blood pressure up to 6% after 24 hours. . p3 showed a transient blood pressure increase of 14% immediately after administration of 2 mg / kg, and recovered after 5 minutes of administration, and no further action was observed. When 10 mg / kg was administered, 7% of the blood pressure was lowered after 2 hours, and 11% of the blood pressure was lowered up to 24 hours.
    [127] That is, from the N-terminal end of the rat prorenin pf region to the 27-41th amino acid sequence corresponding to the C-terminal side of the peptide and rat prorenin pf region consisting of the 1st to 11th or 5th to 19th amino acid sequences. The constructed peptides were found to have sustained hypotension.
    [128] In addition, the results indicate that the transient blood pressure drop immediately after administration by Pmix is the action of p2, and the subsequent blood pressure lowering action is due to the interaction of p1, p2, and p3.
    [129] Experimental Example 4
    [130] As a comparative experiment, the same experiment as in Experiment 2 was performed using 5 normal blood pressure rats (WKY rats) of the same age instead of SHR. No hypotension was found except for transient hypotension immediately after Pmix administration. Therefore, Pmix did not show a sustained hypotensive effect on normal rats.
    [131] Experimental Example 5
    [132] P2a for p1 (pf1-11), p1a (pf1-4), p1b (pf1-7), p1c (pf5-11), p2a (pf12-19) and p2b (pf11-15) prepared in Example 2 (pf12-19) and p2b (pf11-15) were carried out in the same manner as in Experiment 1 using two SHRs each at a dose of 1 mg / kg and other peptides at a dose of 10 mg / kg. It was.
    [133] As a result, p1 showed sustained blood pressure drop of up to 14% up to 24 hours after administration. p1a showed a transient blood pressure rise of 13% immediately after administration, but a sustained blood pressure drop of up to 13% until 24 hours after administration. p1b showed a transient blood pressure drop of 17% immediately after administration, but continued up to 13% of blood pressure up to 24 hours after administration. p1c showed a transient blood pressure rise of 21% immediately after administration, but continued to decrease blood pressure up to 16% until 24 hours after administration. p2a showed a transient blood pressure increase of 6% immediately after administration, but continued to have a continuous blood pressure drop of up to 14% until 24 hours after administration. In addition, the same effect was obtained even when using p2b.
    [134] As a result, the peptide derived from the 1st to 19th amino acid sequence from the N terminus of the amino acid sequence of the rat prorenin pf region was found to have a sustained blood pressure lowering action.
    [135] Experimental Example 6
    [136] P2a (pf12-19) prepared in Experimental Example 2 and H6137 (D-His-Pro-Phe-His-Leu-ψ- [CH2NH] -Leu-Val-Tyr, Sigma), which are renin inhibitors, were administered, respectively. After the dose of 1 mg / kg was administered to the external jugular vein using canure, changes in blood pressure over time were measured by the method described in Experimental Example 1.
    [137] As a result, p2a showed a sustained drop in blood pressure of up to 11% at 5-8 hours after administration, and the result lasted at least 12 hours after administration. On the other hand, H6137 showed a continuous blood pressure drop of up to 9% at 3 to 8 hours after administration, but the tendency of the result was confirmed after 6 hours after administration.
    [138] While p2a takes a long time to act, the action time is long, whereas H6137 exhibits a fasting effect but a short action time. From this difference in time-dependent action, it can be inferred that the force-reducing effect of the proenin activation control agent represented by p2a is not caused by renin inhibition.
    [139] Experimental Example 7
    [140] P1c (pf5-11) prepared in Example 2 was administered intravenously to four SHR 1 groups at 1.6 mg / kg once a day for 12 consecutive days. On day 14, the hemorrhage was performed to remove the heart and kidney and the weight thereof was measured. The results are shown in Table 1 in terms of organ weight per 100 g of body weight. As shown in Table 1, the weight of the heart and kidney of SHR administered p1c was lighter than that of the control group administered physiological saline, indicating that p1c suppressed cardiac hypertrophy and renal hypertrophy.
    [141] SubstanceOrgan weight (g / g body weight) HeartRight kidneyLeft kidney Saline solution120.430.470.380.390.400.44 Mean standard error0.450.020.390.000.420.02 p1c34560.370.400.390.400.340.350.350.330.340.350.350.35 Mean standard error0.390.010.340.000.350.00
    [142] Experimental Example 8
    [143] For the rats tested in Example 7, the pulmonary artery, superior mesenteric artery, and femoral artery were removed and their weights measured. The results were compared by converting the weight of each artery into the weight per area and converting the converted value into the weight per weight of each rat, and are shown in Table 2. As shown in Table 2, the weight of each artery of SHR to which p1c was administered was apparently light compared to the control group, indicating that p1c inhibited arterial thickening, ie, vascular remodeling.
    [144] SubstanceOrgan weight (g / g body weight) Pulmonary arterySuperior mesenteric arteryFemoral artery Saline solution120.000930.001190.000840.001070.001380.00135 Mean standard error0.001060.000130.000960.000120.001370.00002 p1c34560.000670.000870.000880.000820.000940.000950.000960.001000.000680.000940.001060.00087 Mean standard error0.000810.000050.000960.000010.000890.00008
    [145] Example 3
    [146] A screening system was constructed using antibodies capable of activating human prorenin by specifically recognizing human prorenin and human prorenin pf peptides prepared according to Japanese Patent Application Laid-Open No. Hei 10-279600 and US Pat. . By testing candidate compounds in the constructed screening system, the effect on human prorenin activation by anti-pf peptide antibody was confirmed. Through this method, a substance capable of inhibiting the activation of human prorenin by anti-human pf peptide antibody was obtained. In addition, substances having a coercive effect, long-term hypertrophy effect, and arterial hypertrophy effect, from substances obtained by experiments using animal models and the like which accumulate information based on peptides selected from human prorenin profragment regions, known coercive agents, and the like. Can be selected.
    [147] In accordance with the present invention, a method for screening a substance capable of inhibiting the activation of prorenin using the regulation ability of prorenin activation by the interaction between proteins in the profragment region of prorenin and the profragment region of prorenin Prorenin activation regulators are provided that have the function of controlling prorenin activation based on protein-protein interactions. In addition, by using the prorenin activation control substance, it is possible to provide a suppressor, a long-term hypertrophy inhibitor, and an arterial thickener having a novel mechanism. These drugs can be effectively used for the treatment of hypertension, heart failure, renal failure, and arteriosclerosis, ie diseases involving vascular remodeling, by being used alone or in combination with existing drugs.
    权利要求:
    Claims (35)
    [1" claim-type="Currently amended] A method for screening a substance capable of controlling the activation of prorenin, which is used as an indicator of the regulation of prorenin activation that does not involve a change in primary structure caused by protein-protein interaction in the profragment region of prorenin.
    [2" claim-type="Currently amended] A method for screening a substance capable of controlling the activation of prorenin using the regulation of the prorenin activation by the antibody to the profragment region of the prorenin.
    [3" claim-type="Currently amended] The method of claim 1 or 2, wherein the substance to be selected is a substance designed based on information of at least three amino acid sequences of the amino acid sequence of the profragment region of prorenin. Selection of Substances That Can Be.
    [4" claim-type="Currently amended] The substance to be selected is the 1st to 19th amino acid sequence of the N-terminus or 27th to 41st amino acid sequence of the amino acid sequence of the profragment region of prorenin, A method for screening a substance capable of controlling prorenin activation, characterized in that the substance is designed based on at least three amino acid sequence information.
    [5" claim-type="Currently amended] The material of claim 1 or 2, wherein the substance to be selected is based on at least three amino acid sequence information of the fifth to nineteenth amino acid sequence of the N-terminus of the amino acid sequence of the profragment region of prorenin. A method for screening a substance capable of controlling prorenin activation, characterized in that it is a designed substance.
    [6" claim-type="Currently amended] The method for selecting a substance capable of controlling prorenin activation according to any one of claims 3 to 5, wherein the substance to be selected is a peptide.
    [7" claim-type="Currently amended] The method for screening a substance capable of controlling prorenin activation according to any one of claims 3 to 5, wherein the substance to be screened is a low molecular weight compound.
    [8" claim-type="Currently amended] 8. The method of any one of claims 1 to 7, wherein the prorenin is human prorenin.
    [9" claim-type="Currently amended] 8. The method for screening a substance according to any one of claims 1 to 7, wherein the prorenin is rat prorenin.
    [10" claim-type="Currently amended] A prorenin activation control substance having a function of inhibiting prorenin activation that is not accompanied by a change in primary structure caused by protein-to-protein interaction in the profragment region of prorenin.
    [11" claim-type="Currently amended] Inhibiting prorenin activation, which is selected by the method of any one of claims 1 to 9 and which does not involve changes in primary structure, caused by interprotein interactions in the profragment region of prorenin Prorenin activation control substance having the function to.
    [12" claim-type="Currently amended] A prorenin activation control substance having a function of inhibiting prorenin activation that does not involve primary structural changes, which is caused by interprotein interactions in the profragment region of prorenin.
    [13" claim-type="Currently amended] A prorenin activation control substance having a function of inhibiting prorenin activation that does not involve primary structural changes caused by interprotein interactions in the profragment region of prorenin, and at the same time having no renin inhibitory activity.
    [14" claim-type="Currently amended] A method for inhibiting prorenin activation that is not accompanied by a primary structural change, which is selected by the method according to any one of claims 1 to 9 and is caused by interprotein interactions in the profragment region of prorenin. Prorenin activating control agent with function.
    [15" claim-type="Currently amended] A method for inhibiting prorenin activation that is not accompanied by a primary structural change, which is selected by the method according to any one of claims 1 to 9 and is caused by interprotein interactions in the profragment region of prorenin. Prorenin activation control substance having a function and at the same time does not have a renin inhibitory activity.
    [16" claim-type="Currently amended] Has a function of inhibiting prorenin activation that is not accompanied by primary structural changes, caused by interprotein interactions in the profragment region of prorenin, and has a partial sequence selected from the amino acid sequence of the profragment region of prorenin Prorenin activation control substance consisting of a peptide or an equivalent peptide thereof.
    [17" claim-type="Currently amended] A contiguous 3 selected from the amino acid sequence of the profragment region of prorenin, having the function of inhibiting prorenin activation not accompanied by a change in primary structure, caused by interprotein interactions in the profragment region of prorenin A prorenin activation control substance consisting of peptides having dog to ten subarrays.
    [18" claim-type="Currently amended] A contiguous 3 selected from the amino acid sequence of the profragment region of prorenin, having the function of inhibiting prorenin activation not accompanied by a change in primary structure, caused by interprotein interactions in the profragment region of prorenin A prorenin activation control substance consisting of peptides having dog to eight subarrays.
    [19" claim-type="Currently amended] A contiguous 3 selected from the amino acid sequence of the profragment region of prorenin, having the function of inhibiting prorenin activation not accompanied by a change in primary structure, caused by interprotein interactions in the profragment region of prorenin A prorenin activation control substance consisting of peptides having dog to six subarrays.
    [20" claim-type="Currently amended] Selected from the peptides set forth in SEQ ID NOs: 7-14 of the Sequence Listing, having the function of inhibiting prorenin activation that does not involve changes in primary structure caused by interprotein interactions in the profragment region of prorenin Prorenin activation regulator consisting of peptides.
    [21" claim-type="Currently amended] SEQ ID NOs: 3, 4, and 15-20 of the Sequence Listing, having the function of inhibiting prorenin activation that does not involve changes in primary structure, caused by interprotein interactions in the profragment region of prorenin A prorenin activation modulator consisting of a peptide selected from the described peptides.
    [22" claim-type="Currently amended] It is a low molecular compound designed from the amino acid sequence information of the profragment region of prorenin, and has a function of inhibiting prorenin activation which is not accompanied by the change of primary structure caused by the interaction between proteins in the profragment region of prorenin. Having prorenin activation regulator.
    [23" claim-type="Currently amended] 23. The method according to any one of claims 16 to 22, which has a function of inhibiting prorenin activation that does not involve changes in primary structure caused by interprotein interactions in the profragment region of prorenin. Prorenin activating control agent having no renin inhibitory activity.
    [24" claim-type="Currently amended] 24. The prorenin activation modulator according to any one of claims 12 to 23, wherein the effect of inhibiting prorenin activation occurs by antagonism of interprotein interactions in the profragment region of prorenin.
    [25" claim-type="Currently amended] A prorenin activation control substance, characterized in that it inhibits prorenin activation in vivo by elimination of prorenin based on an antigen antibody response to the profragment region of prorenin.
    [26" claim-type="Currently amended] A prorenin activation control substance which is capable of inhibiting prorenin activation in vivo by elimination of prorenin based on an antigenic antibody response to a profragment region of prorenin, and composed of an antibody to the profragment region of prorenin.
    [27" claim-type="Currently amended] A pressure reducing agent containing at least one kind of the activation control substance of prorenin according to any one of claims 12 to 26 as an active ingredient.
    [28" claim-type="Currently amended] 29. The antagonist of claim 27, wherein the antagonist exhibits a coercive rate of at least about 10% within 12 hours after in vivo administration.
    [29" claim-type="Currently amended] 29. The antihypertensive according to claim 27, wherein the hypertensive effect is selectively exerted only on hypertensive patients.
    [30" claim-type="Currently amended] A long-term hypertrophy inhibitor containing at least one of the prorenin activation control substances as described in any one of Claims 12-26 as an active ingredient.
    [31" claim-type="Currently amended] 31. The long-term hypertrophy inhibitor according to claim 30, wherein the long-term hypertrophy of the heart and / or the kidney can be suppressed.
    [32" claim-type="Currently amended] An arterial thickening agent containing at least one of the prorenin activation control substances according to any one of claims 12 to 26 as an active ingredient.
    [33" claim-type="Currently amended] The method of treating hypertension using the hypertensive agent of any one of Claims 27-29.
    [34" claim-type="Currently amended] 32. A method for treating heart failure or kidney failure using the long-term hypertrophy inhibitor according to claim 30 or 31.
    [35" claim-type="Currently amended] A method for treating a disease associated with vascular thickening using the arterial thickening agent according to claim 32.
    类似技术:
    公开号 | 公开日 | 专利标题
    US20170037150A1|2017-02-09|MASP-2, A Complement Fixing Enzyme, and Uses for It
    JP2018197234A|2018-12-13|Plasma kallikrein binding proteins
    US20180092961A1|2018-04-05|Extracellular histones as biomarkers for prognosis and molecular targets for therapy
    US20140302048A1|2014-10-09|Plasma kallikrein binding proteins
    Kirkby et al.2008|The endothelin system as a therapeutic target in cardiovascular disease: great expectations or bleak house?
    Ferrario2010|New physiological concepts of the renin-angiotensin system from the investigation of precursors and products of angiotensin I metabolism
    DE69637150T2|2008-02-28|For the beta-A4 | peptide specific monoclonal antibody
    Margolius1995|Kallikreins and kinins: some unanswered questions about system characteristics and roles in human disease
    CA2512545C|2015-06-30|Recombinant vhh single domain antibody from camelidae against von willebrand factor |
    Zuk et al.1998|Polarity, integrin, and extracellular matrix dynamics in the postischemic rat kidney
    JP4281852B2|2009-06-17|Blood coagulation inhibitory antibody and method of use thereof
    DE69628652T2|2004-04-29|Vegf related protein
    KR100881900B1|2009-02-04|Anti-osteopontin antibody and use thereof
    Gils et al.2004|The structural basis for the pathophysiological relevance of PAI-1 in cardiovascular diseases and the development of potential PAI-1 inhibitors
    AU2001247219B2|2007-01-04|Integrin antagonists
    RU2156468C2|2000-09-20|Method of assay of presence or absence of inhibitory activity of platelet aggregation |, purified and isolated inhibitor of platelet activity | and/or its truncated form, method of purification of pa1 from snake venom, pharmaceutical composition for prevention of thrombus formation, method of inhibition of thrombus formation
    US20160271234A1|2016-09-22|TISSUE INHIBITOR OF METALLOPROTEINASES | LINKED TO GLYCOSYLPHOSPHATIDYLINOSITOL | - ANCHORS FOR TREATMENT OF CANCER AND SKIN LESIONS
    JP4954426B2|2012-06-13|Angiogenesis regulating composition and use
    Kozma et al.1996|The GTPase-activating protein n-chimaerin cooperates with Rac1 and Cdc42Hs to induce the formation of lamellipodia and filopodia.
    AU2001265953B2|2005-09-29|Use of inhibitors of placental growth factor for the treatment of pathological angiogenesis, pathological arteriogenesis, inflammation, tumour formation and/or vascular leakage
    US6315995B1|2001-11-13|Methods for treating an ischemic disorder and improving stroke outcome
    ES2628867T3|2017-08-04|Methods to treat conditions associated with activation of the MASP-2-dependent complement
    Mercure et al.2008|Angiotensin | blunts hypertensive cardiac remodeling by a direct effect on the heart
    CN1826349B|2011-05-04|Wound care compositions
    AU601801B2|1990-09-20|Anticoagulant peptides
    同族专利:
    公开号 | 公开日
    CN1311238C|2007-04-18|
    AU2001246864B2|2006-07-27|
    EA005310B1|2004-12-30|
    US6900020B2|2005-05-31|
    MXPA02010028A|2004-08-19|
    EP1273914A4|2004-08-25|
    CA2404644A1|2001-10-18|
    AU4686401A|2001-10-23|
    EP1273914A1|2003-01-08|
    US20030165999A1|2003-09-04|
    US7361637B2|2008-04-22|
    WO2001077673A1|2001-10-18|
    KR100775980B1|2007-11-16|
    EA200201072A1|2003-06-26|
    CN1436305A|2003-08-13|
    US20050203021A1|2005-09-15|
    引用文献:
    公开号 | 申请日 | 公开日 | 申请人 | 专利标题
    法律状态:
    2000-04-10|Priority to JP2000108670
    2000-04-10|Priority to JPJP-P-2000-00108670
    2001-04-09|Application filed by 유이치 이쉬다
    2002-12-31|Publication of KR20020097217A
    2007-11-16|Application granted
    2007-11-16|Publication of KR100775980B1
    优先权:
    申请号 | 申请日 | 专利标题
    JP2000108670|2000-04-10|
    JPJP-P-2000-00108670|2000-04-10|
    [返回顶部]