MXPA97004879A - Announcing monoclonal antibody that has an inhibitory activity against types ii type ii phospholipase and protein that comprises a part of my - Google Patents

Abstract

A novel monoclonal antibody capable of inhibiting type II phospholipase A2 activity is obtained. Since said antibody is capable of inhibiting the activity of human type II phospholipase A2, as well as phospholipase A2 of monkey type II and / or mouse type II phospholipase A2, it is not only clinically useful, but can also be used as such in preclinical tests with the use of monkey or mouse. The antibody, whose completely novel property of releasing type II phospholipase A2 bound to the cell membrane, seems to strongly inhibit the activity of said enzyme through a novel mechanism. The monoclonal antibody or protein comprising a part thereof of the present invention can be used as a therapeutic agent for cardiac infarction, cerebral infarction, etc., where it is implied by phospholipase A2 of type

Description

ANNOUNCING MONOCLONAL ANTIBODY THAT HAS AN INHIBITOR ACTIVITY AGAINST TYPE II PHOSPHOLIPASE A2 AND PROTEIN THAT COMPRISES A PART OF THE SAME FIELD OF THE INVENTION The present invention relates to novel monoclonal antibodies having a potent inhibitory activity against phospholipase A2 of type I I, and proteins comprising a part thereof. More specifically, the present invention relates to novel monoclonal antibodies, which are superior in the specificity and affinity for type II phospholipase A2, highly inhibitors against type II phospholipase A2, and useful as therapeutic agents for the treatment of diseases related to type II phospholipase A2 and protein comprising a part thereof. In addition, the present invention relates to cells that produce said monoclonal antibodies or proteins, DNAs that encode such monoclonal bodies or proteins, and recombinant vectors comprising said DNAs. In addition, the present invention relates to medical compositions comprising monoclonal antibodies or proteins, or an inhibitor of said phospholipase A2 of type I I.

BACKGROUND OF THE INVENTION Phospholipase A2 is known as the enzyme to catalyze the hydrolysis of the ester link at the C2 position of the 1,2-acylphosphoglyceride, a biomembrane component. This enzyme is distributed in several organs and cells of mammals, and not only regulates the generation and metabolism of the biomembrane phospholipid, but also acts as the regime limiting enzyme in the arachidonic acid cascade, the metabolic products of which they include prostaglandin, leukotriene, thromboxane, PAF, etc. , they are known to have a variety of physiological activities. Various types of phospholipase A2 are known, such as type I, type I I, intracellular type, etc. (Atsumi I. and others, Nippon Rinsho, Extra edition 1994, 202-206). Among them, phospholipase A2 of type I I is induced in the exudate of the inflammatory site, and released in large quantities in the bloodstream during the inflammatory reaction. Several reports indicate that this enzyme exacerbates the symptom of several inflammatory diseases or is a part of its cause. For example, Kikuchi-Yanoshita et al., Reported the elevation of this enzyme activity in rats with ischemic heart disease as an animal model of cardiac infarction, when organ damage progresses (Kikuchi-Yanoshita R. et al., J. Biochemistry, 1 14, 33-38, 1993). Leong et al. Reported the presence of a larger amount of this enzyme in the bloodstream of patients with cardiac infarction than in normal subjects (Leong L. L. et al., Clinical Experimental Pharmacology and Physiology, 19: 1 13-18 1892). Lauritzen and others reported the indication that this enzyme plays an important role in the exacerbation of symptoms of ischemic reperfusion disorder in patients with cerebral infarction (Lauritzen I. and others, Brain Research, 651: 353-356, 1994). Bauer and others also reported that this enzyme plays an important role in the exacerbation of symptoms in acute renal failure (Bauer M., Klin. Wochenschr., 67: 196-202, 1989). Murakami and others suggested that this enzyme stimulates the release of histamine from mast cells, which play central roles in allergic diseases such as asthma, etc. , and also that the inhibition of this enzyme can lead to a possible development of therapeutic agents for the treatment of allergic diseases including asthma, etc. , (Murakami M. et al., Journal of Immunology, 151: 5675-5684, 1993). Smith and others and Pruzanski and others reported the presence of larger amounts of this enzyme in the bloodstream of patients with chronic rheumatoid arthritis than in normal subjects, suggesting the possibility that this enzyme causes or exacerbates the disease (Smith G. M. and others, British Journal of Rheumatology, 31: 175-178, 1992; Pruzanski W. and others, J. Rheumatol. , 15: 1351-1355, 1988). Pruzanski and others similarly reported the presence of larger amounts of this enzyme in exudates from patients with osteoarthritis than in normal subjects, suggesting the possibility that this enzyme is a cause of the disease or its exacerbation (Pruzanski W. and others, Life Sciences, 48: 2457-2462, 1991). Vadas et al. And Green et al. Reported the presence of larger amounts of this enzyme in the bloodstream of patients suffering from septic shock than in normal subjects, suggesting that this enzyme is a possible cause of the disease or its exacerbation (Vadas P. and others, Life Sciences, 50: 807-81 1, 1992; Green, J., Inflammation, 15: 355-367, 1991). Nevalainen and others reported the presence of large amounts of this enzyme in the bloodstream of patients with especially severe pancreatitis, suggesting the possibility that this enzyme is a cause of the disease or its exacerbation (Nevalainen T. J et al., Gut, 34, 1 133-1 1 36, 1993). Anderson and others reported the presence of said enzyme in large quantities on the skin of patients with psoriasis, suggesting a possibility that this enzyme is a cause of the disease or its exacerbation (Anderson S. et al., Inflammation, 18: 1 - 12, 1994). Koike and others reported the suggestion that this enzyme plays a major role as the cause of multiple organ failure (MOF) (Koike K. et al., Surgery, 1 12: 173-180, 1992). Koeniger et al., Edelson et al., And Romaschin et al. Reported the presence of large amounts of this enzyme in the bloodstream of patients with acute respiratory distress syndrome (ARDS) than in normal subjects, suggesting the possibility that this enzyme be a cause of the disease or its exacerbation (Koeniger R., et al., Klin. Wochenschr., 67: 212-216, 1989; Edelson J, D., et al., Am. Rev. Respir. Dis., 143: 1 102-1199, 1991; Romaschin A. et al., Clin. Biochem., 25: 55-60, 1992). Minami and others reported the presence of large amounts of this enzyme in the bloodstream of patients with Crohn's disease and ulcerative colitis, suggesting the possibility that this enzyme is the cause of the diseases or their exacerbation (Minaki T. and others, Gut, 33: 914-921, 1992). In addition, it has been indicated that this enzyme is involved with uveitis, respiratory distress syndrome of the newborn (RDS), bronchopulmonary dysplasia (BRD), etc. As the anti-human type II phospholipase A2 antibody, the following antibodies are reported. Stoner et al. Purified human type II phospholipase A2 from human placenta and synovial fluid, and, using the purified enzyme as antigen, obtained a monoclonal antibody developed against it in mice (Stoner CR et al., Journal of Immunological Methods, 145 : 127-136, 1991). In this method, they first sensitized mice of the BALB / cByJ strain, with the purified phospholipase A2 mixed with Freund's complete adjuvant (5 μg) followed by the same enzyme (3 μg) 25 days later. In addition, they sensitized the mouse with the same preparation (5 μg) 3 days before the fusion of the spleen cells (36 days after the initial sensitization). Therefore, they immunized the mouse with the immunogen, totaling 13 μg, divided into three portions over a period of 36 days. The isolated spleen cells were fused with mouse myeloma PAI-0 cells, and screened in a HAT medium. The supernatant of the medium was examined through the ELISA system using phospholipase A2 to obtain the monoclonal antibodies (PLA 184, PLA 185, PLA 186 and PLA187). These antibodies were certainly able to inhibit human phospholipase A2. However, the inhibitory activity of these antibodies against phospholipase A2 of other animal species was examined using only rat type II phospholipase A2. They reported that these antibodies showed little activity or no cross-reactive activity with rat type I phospholipase A2, and, consequently, could not be used in animal experiments with rats. Takayama and others also purified human type II phospholipase A2 from the synovium of patients with rheumatism, and, using this purified enzyme as an antigen, obtained a monoclonal antibody in mouse (Takayama K. et al., Biochemical and Biophysical Research Communications, 167 : 1309-1315, 1990). In this immunization method, they adsorbed the purified human type II phospholipase A2 (10 μg) on nitrocellulose membrane, the resulting homogenate was homogenized and injected into the spleen of the BALB / c mouse, followed by a similar immunization two weeks after . Therefore, they immunized the animal with the immunization antigen with a total of 20 μg, were divided into two equal portions for a period of 28 days, and then, 3 days later, and performed the fusion of the spleen cells with mouse myeloma cells (X63-Ag8.6.5.3). They examined the supernatant of the medium through the ELISA system, using phospholipase A2 to obtain the reactive monoclonal antibodies (HP-1, HP-2, HP-3 and HP-4). Of these, HP-1 showed the highest inhibitory activity to human type II phospholipase A2. However, although HP-1 showed only weak inhibitory activity, inhibiting approximately 80% even by adding an excess of 200 molar. McCord et al. Purified a recombinant human type II phospholipase A2 and obtained a monoclonal antibody in the mouse, using this purified enzyme as the antigen (McCord M. et al., The Journal of Investigative Dermatology, 102: 980-986, 1994 ). For the immunization of animals, they immunized a mouse of the CAF1 race with phospholipase A2 mixed with complete Freund's assistant in amounts of 100, 50 and 25 μg, respectively, every four weeks, the final immunization was carried out three days before the fusion of Spleen cells In this manner, approximately 200 μg total of the immunogen divided into four portions, were immunized for a period of 56 days. The spleen cells were fused with SP2 / 0-AG14 mouse myeloma cells, and were classified for positive cells in the HAT medium. The supernatant of the medium for the inhibitory activity was examined, using phospholipase A2 to obtain a monoclonal antibody (3F10).

They examined the inhibitory activity of this antibody (1 μg) for human phospholipase A2, but without specifying the amount of the enzyme used, making it difficult to assess its inhibitory potency. In addition, in H4-506447 of Tokkai, Johnson L. K. described the purification of a recombinant human phospholipase A2, and the preparation of a polyclonal antibody in rabbits, using the purified enzyme as an immunogen. This antibody is not monoclonal, and, in addition, it used two types of peptide as antigens, which are parts of the human phospholipase A2 sequence, "GTKFLSYKFSNSGSRITC" (amino acids 67-85 of the N term) and "N KTTIN KKYQYYSN KHSRGSTPRC" ( amino acids 109-132 of the term N). He analyzed the inhibitory capacity of this antibody without specific amounts of the antibody and the human phospholipase A2 used in the specification, making it difficult to assess its inhibitory potency. Also in H7-109300 from Tokkai, an antibody was described which stimulates the binding of human phospholipase A2 to the sulfated polysaccharide, without confirming the inhibition of human type I phospholipase A2 by said antibody. In addition, there is no information with respect to the inhibitory activity of such known antibodies described above against type II phospholipase A2 of monkey, mouse, cat, dog, and rat, or their potencies to release bound human type I phospholipase A2. By immunizing the mouse with the human enzyme, the antibody is easily formed, which recognizes the different amino acid sequence between mouse and human enzymes. Thus, when the mouse is immunized with human type II phospholipase A2, it is theoretically assumed that almost all mouse antibodies, which bind to human type II phospholipase A2, do not bind to type II phospholipase A2. mouse . Also, the active center of the enzyme and its surroundings of phospholipase A2 of type I I, are generally conserved among several animal species, and the homology is completely high between the mouse and the human being. When the mouse is immunized with human type II phospholipase A2, it is difficult to obtain, due to immunological tolerance, antibodies that recognize the active center of the enzyme and its surroundings, resulting in the present situation, where it is impossible to acquire antibodies with a strong inhibitory activity for human type II phospholipase A2. No antibody has been known that not only strongly inhibits human type I I phospholipase A2, but also inhibits mouse type I I phospholipase A2 and / or monkey type I phospholipase A2. Before clinical testing of new drugs in humans, it is a prerequisite to clarify their pharmaceutical effects in animal experiments. Therefore, it is necessary to have a monoclonal antibody that shows pharmaceutical effectiveness in experimental animals, especially small ones such as mouse and monkey, that is, a monoclonal antibody that inhibits not only human type II phospholipase A2, but also the enzyme of these other animals. It is also known that phospholipase A2 must bind to cells when the enzyme exerts its hydrolytic action on the membranous phospholipid (Suga H., Et al., Eur. J. Biochem., 218: 807-813, 1993; Murakami M. et al. J. Biol. Chem., 268: 839-844, 1993). Therefore, if the antibodies are capable not only of inhibiting free phospholipase A2 in the bloodstream or exudates, but also of releasing said enzyme bound to the cells, their membranous phospholipids which will be hydrolyzed, these antibodies are expected to have potent effects. inhibitors in phospholipase A2 based on a novel mechanism of action. In addition, when the antibody binds to phospholipase A2, associated with the surface of the cell, the complement and / or effector cells can attack said cell, possibly exerting unfavorable side effects on it. Nevertheless, the antibodies that can release type I I phospholipase A2 bound to the cell, have not been obtained yet. Therefore, there has been a strong demand for antibodies against type II phospholipase A2 under the consideration of pharmaceuticals, wherein said antibodies strongly inhibit, not only human type II phospholipase A2, but also said enzyme derived from other species animals, and also have properties to be able to release type II phospholipase A2 bound to the cell.

DESCRIPTION OF THE INVENTION The present invention presents efforts to solve these problems, described above, and the following information is obtained to achieve the invention. That is, we immunized mice with a recombinant human type I phospholipase A2, mixed with Freund's complete adjuvant at a dose of 20 μg per mouse, 8 times at an interval of 2 to 3 weeks. Thus, we differ from other researchers in that it is intended to avoid immunological tolerance by increasing the frequency of immunization and extending the period of immunization. In addition, we tried to improve the system to analyze the inhibition of phospholipase A2 to classify antibodies. That is, we use a phospholipid derived from Escherichia coli as a highly sensitive substrate for the analysis of phospholipase A2 activity (Jacobson PB et al., Biochem. Pharm., 39: 1557, 1990), and we also improve the method for preparing said phospholipids of E. coli as the substrate. That is, we used Escherichia coli SN 17 deficient in phospholipase A2 (pldA, pla-2, trh-1, leuB6, thi) (Doi O. et al., J .. Biochem., 80: 1247-1258, 1976), and it incorporated tritium-labeled oleic acid into it to prepare its phospholipids. Using this microorganism, we were able to prepare the phospholipid of £. tritium-labeled coli with a high specific radioactivity efficiently without exposing it to degradation by E. coli phospholipase A2. Furthermore, by using a test system with a longer enzyme reaction time and a higher sensitivity than the conventional method (J. Biol. Chem., 261: 4239-4246, 1986), we were able to efficiently and accurately classify the hybridoma producing the antibody. In this way, we examined its inhibitory effects on phospholipase A2 activity derived from several animals. Also, in order to examine a novel function of the antibodies thus obtained, we used a test system to test the ability of these antibodies to release the phospholipase A2 bound to the cell. As a result, we succeeded in obtaining a characteristic monoclonal antibody by having the following properties. That is, in the monoclonal antibody obtained through the present inventors, it is reactive with phospholipase A2 of human type I I and highly inhibitor against it. Also, the antibody cross-reacts with type I I phospholipase A2 derived from monkey and mouse platelets. Therefore, using the antibody of the present invention, animal experiments with monkeys and mice can be performed prior to the clinical testing of new drugs in humans. In addition, the monoclonal antibody of the present invention is capable, not only of inhibiting the free enzyme in the bloodstream or exudate, but also of releasing the phospholipase A2 of type I I bound to the cell. Phospholipase A2 of type I I binds to the cell via heparin sulfate, etc. , on the surface of the cell, and it is known to bind to hepatocyte, endothelial cells, etc. , which have heparin sulfate, etc. , on the surface of the cell. The monoclonal antibody of the present invention is characteristic not only for inhibiting free type I I phospholipase A2, in a bloodstream or exudate, but also for releasing the enzyme bound to the cell from the cell membrane. Additionally, the present invention includes a protein consisting of a portion of the monoclonal antibody with the properties described above and having the antigen binding capacity equivalent to that of said monoclonal antibody. Also, the present invention includes reduced alkylated derivatives of said protein, and variant proteins produced by the addition, elimination, substitution or mutation of one or more amino acid residues in the amino acid sequence of the monoclonal antibody obtained by immunizing animals, provided that they retain the antigen binding capacity equivalent to that of said monoclonal antibody. further, the inventors isolated the cDNAs encoding the variable regions of the H and L chains of the "hybridoma 1.4" and "hybridoma 10.1" antibodies, generating the antibody comprised in the present invention, and determining the sequence of complete base. This allows not only the production of the antibody on a large scale using engineering techniques by engineering, but also the easy modification of said antibody of the present invention using engineering techniques of the antibody. [Base sequences determined with "hybridoma 1 .4" are shown in SEC I D NO: 1 (L chain) and SEC I D NO: 3 (H chain). Also, the putative amino acid sequences corresponding to said base sequences are shown in SEQ ID NO: 2 (L chain) and SEQ ID NO: 4 (H chain). Base sequences determined with "hybridoma 10.1" are shown in SEQ ID NO: 5 (L chain) and SEQ ID NO: 7 (H chain). Also, the putative amino acid sequences corresponding to said base sequences are shown in SEC I D NO: 6 (L chain) and SEC I D NO: 8 (H chain)]. That is, the present invention comprises the following: 1) A monoclonal antibody capable of inhibiting the activity of human type II phospholipase A2, as well as that type II phospholipase A2 derived from monkey and / or mouse, or a protein comprising a part thereof having said inhibitory activity, 2) A monoclonal antibody capable of releasing type II phospholipase A2 bound to the cell membrane, or a protein comprising a part thereof having said activity, 3) The monoclonal antibody or protein according to 2), wherein said phospholipase A2 is derived from the human being, 4) A monoclonal antibody capable not only of inhibiting the activity of human type II phospholipase A2, as well as that of type II phospholipase A2 derived from mono and or mouse, but also to release type II phospholipase A2 bound to the cell membrane, or a protein comprising a part thereof having said activity, 5) a monoclonal antibody prod ucido through any of the hybridomas 12 H5 (FERM BP-5300), 1.4 (FERM BP-5297) and 10.1 (FERM BP-5297), or a protein comprising a part thereof, or a monoclonal or a protein comprising a part thereof, having the activity against type II phospholipase A2 equivalent to that of said monoclonal antibody or protein, 6) The monoclonal antibody or protein comprising a part thereof according to 1) or 2) , wherein said monoclonal antibody or protein comprises a protein having the amino acid sequence represented by any of SEQ ID NO: 2, 4, 6 or 8, or those modified through substitution, elimination or insertion of one or more amino acid residues contained in said sequence, 7) The monoclonal antibody or protein comprising a part thereof according to 1) or 2), wherein said monoclonal body or protein comprises a protein having the amino acid sequence represents gives by any of SEQ ID NO: 9 to 20, or those modified through the substitution, deletion or insertion of one or more amino acid residues contained in said sequence, 8) A cell producing the monoclonal antibody or protein according to any from 1) to 7), 9) The cell according to 8), wherein said cell is a hybridoma, 10) The cell according to 8) wherein said cell is a cell transformed with recombinant DNA, 1 1) A method for producing the monoclonal antibody or protein according to any of 1) to 7), wherein said method comprises the methods of culturing the cell according to 8), and recovering monoclonal antibodies or proteins from the supernatant of the growth medium, ) A DNA encoding the monoclonal antibody or protein according to any one of 1) to 7), 13) The DNA encoding 12), wherein said DNA comprises the base sequence with any of SEQ ID NO: 1, 3, 5 or 7, or that modified by the substitution, deletion, or insertion of one or more amino acid residues contained in said sequence, 14) A recombinant vector comprising the DNA according to 12) or 13), 15) A medical composition that comprises the monoclonal antibody or protein according to any one of 1) to 7), and a pharmaceutically acceptable carrier, 16) A type II phospholipase A2 inhibitor, comprising the monoclonal antibody according to any one of 1) to 7). The "protein" of the present invention is not limited in the number of amino acid residues, including the so-called "peptide" with less amino acid residues. Also, "the monoclonal antibody or protein producing cell" of the present invention includes each of the cell types such as bacteria, yeast, mammalian cell, etc. , provided that it is capable of producing the monoclonal antibody or protein of the present invention. The monoclonal antibody of the present invention is produced by culturing a mouse hybridoma in a growth medium or mouse peritoneal fluid. Also, parts of it can be obtained, such as F (ab ') 2, Fab, Fab', etc. , through the digestion of the antibody produced with a proteolytic enzyme selected from a group consisting of trypsin, papain and pepsin, followed by an appropriate purification. The hybridomas of the present invention, such as the hybridomas 12 H5, 1.4 and 10.1, can be obtained by fusing cells of the BALB / c mouse spleen, which has been sensitized with type II phospholipase A2 normal humans, at increased frequencies and during a period of elongated immunization with mouse myeloma P3x63Ag8 / U 1 (P3U 1) cells through a normal method, for example, the cell fusion method described by Kohier and Milstein (see example later). Hybridomas obtained by the inventors herein, and included in the present invention, are deposited as follows. Deposit of the hybridoma 12H5: a) Name and address of the depository Name: National Institute of Bioscience and Human-Technology, Agency of Industrial Science and Technology, Ministry of International Trade and Industry Address: Higashi 1 -1 -3. Tsukuba-shi, Ibaragi-ken 305, Japan b) Date of deposit (original date of deposit) November 22, 1994 c) Access No. Seimeiken-Jo-Ki No. 5300 (FERM BP-5300) Deposit of Hybridoma 1 .4: a) Name and address of depository Name: National Institute of Bioscience and Human-Technology, Agency of Industrial Science and Technology, Ministry of International Trade and Industry Address: Higashi 1 - 1 -3. Tsukuba-shi, Ibaragi-ken 305, Japan b) Date of deposit (original date of deposit) November 22, 1994 c) Accession No. Seimeiken-Jo-Ki No. 5297 (FERM BP-5297) Deposit of hybridoma 10.1 : a) Name and address of the depository Name: National Institute of Bioscience and Human-Technology, Agency of Industrial Science and Technology, Ministry of International Trade and Industry Address: Higashi 1 -1 -3. Tsukuba-shi, Ibaragi-ken 305, Japan b) Date of deposit (original date of deposit) November 22, 1994 c) Access No. Seimeiken-Jo-Ki No. 5298 (FERM B P-5298). As the culture medium of said hybridomas, minimal essential means of Eagle modified with Dalbecco (hereinafter abbreviated as DMEM) were supplemented with fetal bovine serum, L-glutamine, glucose, sodium pyruvate, 2-mercaptoethanol and antibiotic (for example, penicillin G, streptomycin, gentamicin, etc.), etc. Hybridomas are usually cultured in the medium at 37 ° C in an air atmosphere of 5% CO2-95% air, for 2 to 4 days, or in the peritoneal cavity of a BALB / c mouse, pretreated with 2, 6, 10, 14-tetramethylpentadecane (e.g., pristan, Aldrich) for about 10 to 20 days, to produce the antibody in an amount sufficient for purification. further, the monoclonal antibody or protein comprising a part thereof of the present invention, can be obtained by inserting all or part of the gene encoding the antibody of the present invention or a part thereof into an expression vector, and transferring them to cells of E. coli, yeast or mammal, which will produce them. The monoclonal antibody thus produced can be purified by fractionating the supernatant from the culture medium or peritoneal fluid through normal methods to isolate and purify proteins. These methods include, for example, centrifugation, dialysis, ammonium sulfate precipitation, column chromatography on DEAE-cellulose, hydroxylapatite, and protein-A agarose, etc.

From the antibody thus purified, its active fragments, for example, F (ab ') 2, Fab Fab' Fv, can be obtained through the digestion of said antibody with proteolytic enzymes such as pepsin, papain, etc. , followed by the isolation and purification of digestion using normal methods. In addition, reduced alkylated derivatives of the antibody may be obtained, wherein the H and L chains are bound only by the non-covalent bond, through reductive alkylation of the disulfide bonds linking the H to H chain and / or the H chain. to L using dithiothreitol and iodoacetamide, etc. (Useful Immunological Experimental Method, p39, Kodansha Scientific Book). In addition, in order to suppress effector activity, it is possible to modify the amino acid sequence of the Fe moiety of the monoclonal antibody or a portion thereof of the present invention (Duncan A. R. Et al., Nature, 332: 738, 1988 Tao M. et al., J. Exp. Med., 178: 661-667, 1993; Lund J., J. Immunology, 147: 2657, 1991). It is also possible to replace the Fe fragment with another protein. In addition, it is possible to fuse the Fe fragment with another protein to induce a novel function. When the monoclonal antibody or protein comprising a part thereof of the present invention are used for the treatment of humans, it is preferred to use the antibody or protein comprising a part thereof, which contains portions derived from human beings in large proportions. Said monoclonal antibodies are illustrated by, 1) the so-called "chimeric antibody" consisting of the amino acid sequence derived from animal, such as mouse, etc. , only as the variable region and that derived for human beings as the constant region and 2) the so-called "humanized antibody" consisting of the amino acid sequence derived from animal, such as mouse, only in the region of complementary determination (from hereinafter abbreviated as CDR, or hypervariable region) and that derived from human beings in other regions. These antibodies of such types are also included in the present invention. These "chimeric antibody" or "humanized antibody" can be readily prepared by those skilled in the art, by isolating the gene corresponding to the variable region or CDR of the hybridoma of the present invention, recombining it with the human antibody gene, and transducing of the resulting gene to the host cells that will be expressed (see Int J Cancer, 44, 424-433 (1989), Proc Nati Acad Sci USA, 87, 8095-8099 (1990)). Also, once the base sequence of the gene is determined, it is possible to use the gene comprising the corresponding synthesized sequence, or the gene obtained from the hybridoma or human antibody producing cells combined with said synthesized gene. Also, as the gene corresponding to the variable region or CDR, base sequences shown in either SEQ ID NO: 1, 3, 5 or 7, or DNAs comprising the base sequence of their CDR regions, are highly preferred, or DNAs are preferably used which comprise base sequences encoding the amino acid sequence shown in any of SEQ ID NO: 2, 4, 6 or 8, or that of its C DR region, DNA genes comprising the prepared base sequences modifying one or more bases of said DNA through substitution, elimination or insertion, or parts thereof. Mutagenesis of the gene by substitution, deletion or insertion can be done by known methods (Gillamn et al., Gene, 8, 81-97 (1979), Roberts et al., Nature, 328, 731-734 (1987)). To classify the variant base sequences thus obtained for the sequence encoding the amino acid sequence with preferable properties, methods including the phage display method can be used (Ann.Rev. Immunol., 12, 433-455 (1994)) . Specifically, the "humanized antibody" can be readily prepared by those skilled in the art using known methods (Nature, 321, 522-525 (1986)).; Science, 239, 1534-1536 (1988); Proc. Nati Acad. Sci. USA, 86, 10029-10033 (1989); Proc. Nati Acad. Sci. USA, 88, 2869-2873 (1991)). Furthermore, according to these literatures, for the preparation of the "humanized antibody", it is preferable to use a human antibody, which is highly homologous to that of an animal, other than a human being, from which said CDR region is derived. will be transplanted, since the human antibody will be substituted only in the CDR region with that animal derivative, other than a human being, (ie, the antibody that constitutes the basic skeleton of the "humanized antibody", termed "human receptor antibody" ). As shown in Example 7, human antibodies highly homologous to the mouse antibody, obtained by the present inventors and included in the present invention, were, for example, the Pag-1 antibody (H ughes-Johns N. C , and others, Biochem. J., 268, 135-140 (1990)), the WEA antibody (Goni F., et al., Proc. Nati, Acad. Sci, USA, 80, 4837-4841 (1983)); the ITH5-2 antibody (Chin L. T., et al., Immunol.Let, 44, 25-30 (1995)); the ITC48 antibody (Ohlin M., et al., Mol.Immunol., 31, 983-991 (1994); and these antibodies are conceivably preferable as a "human receptor antibody" to prepare the "humanized antibody." The "chimeric antibody" "can be readily prepared by those skilled in the art using known methods (Nature, 314, 268-270 (1995); Proc, Nati, Acad. Sci. USA, 84, 3439-3443 (1987); Proc. Nati. Acad. Sci. USA, 84, 214-218 (1987), Proc. Nati, Acad. Sci. USA, 81, 6851-6855 (1984).) The preferred monoclonal antibody or proteins included in the present invention, are illustrated by of monoclonal antibodies 12H5 or 1.4, prepared by sensitizing animals with human type II phospholipase A2, as described above, and their fragments such as F (ab '), Fab, Fab', etc. Also, monoclonal antibodies are included which comprise the variable or hypervariable regions of these monoclonal antibodies or their fragments and other reg ions derived from the human being. The medical compositions of the present invention are effective especially through parenteral administration, i.e., subcutaneous, intramuscular or intravenous administration. The composition for parenteral administration is usually composed of a solution containing a monoclonal antibody or protein comprising its fragment dissolved in an administrable vehicle, preferably an aqueous vehicle. Various aqueous vehicles such as water, regular water pH, 0.4% saline, 0.3% glycine solution, 5% glucose solution, human albumin solution, etc. can be used. These solutions must be sterilized, and generally do not contain any particle-forming material. These compositions can be sterilized by conventional and well-known sterilization techniques. These compositions may contain pharmaceutically acceptable supplements to bring them closer to physiological conditions, such as pH and isotonic regulating agents including, for example, sodium acetate, sodium chloride, potassium chloride, calcium chloride, sodium lactate, citrate of sodium, etc. The actual preparation of the parenterally administrable compositions can be performed by those skilled in the art, using established or well known techniques, described, for example, in "Remington's Pharmaceutical Science, 15th Edition, Mack Publishing Company, Easton, PA (1980). " The monoclonal antibodies, proteins or medical compositions of the present invention can be stored frozen or lyophilized, and dissolved in an appropriate solvent when used. Lyophilization and freezing and dissolution of the compositions can be performed by those skilled in the art using known methods.

BRIEF DESCRIPTION OF THE DRAWINGS Figure 1 shows the result of ELISA for 12H5, 10.1 and 1.4, respectively, with recombinant human type I phospholipase A2. Figure 2 shows the result of ELISA for 12H 5, 10.1 and 1.4, respectively, with phospholipase A2 derived from rat platelets. Figure 3 shows the inhibition of 12H 5, 10.1 and 1.4, respectively, against the activity of recombinant human type I phospholipase A2. Figure 4 shows the inhibition of 12H5, 10.1 and 1.4, respectively, against rat phospholipase A2 activity of type I. Figure 5 shows the inhibition of 12H5 against phospholipase A2 derived from platelets of several animals. Figure 6 shows the inhibition of 10.1 against phospholipase A2 derived from platelets of several animals. Figure 7 shows the inhibition of 1.4 against phospholipase A2 derived from platelets of several animals. Figure 8 shows the activity of 12H5, 10.1 and 1.4, respectively, to release phospholipase A2 bound to BRL-3A cells.

Figure 9 shows the anchored PCR method for cDNA cloning of variable regions of the H and L chains of the 1.4 antibody. Figure 10 shows the A DNc sequence of the variable region of the L chain of the 1.4 antibody and the translated amino acid sequence. The putative sequences that correspond to the CDR sequence stand out. Figure 11 shows the cDNA sequence of the variable region of the H chain of the 1.4 antibody and the translated amino acid sequence. The putative sequences that correspond to the CDR sequence stand out. Figure 12 shows the method for cloning cDNA in the variable region of the H and L chains of the antibody, through PC R. Figure 13 shows the cDNA sequence of the variable region of the L chain of antibody 10.1 and the sequence of amino acid translated. The putative sequences that correspond to the CDR sequence stand out. Figure 14 shows the cDNA sequence of the variable region of the H chain of the 10.1 antibody and the translated amino acid sequence. The putative sequences that correspond to the CDR sequence stand out.

BEST MODE FOR CARRYING OUT THE INVENTION Next, examples of the present invention are described without limiting the same in any way.

EXAMPLE 1 Preparation of Hybridomas 12.5. 1.4 and 10.1 a) Preparation of Recombinant Human Type II Phospholipase A2 Monoclonal antibodies of the present invention can be obtained by immunizing animals with a recombinant human type II phospholipase A2, which was prepared completely according to the method described by Tokkai in H5 -192167. b) Preparation of sensitized spleen cells Recombinant human type II phospholipase A2 was dissolved, purified as in Example 1 a), (20 μg per animal), in physiological saline (0.1 ml), mixed with an equal volume of auxiliary Freund's complete to emulsify, and BALB / c mice were injected peritoneally (6 weeks of age at the time of immunization initiation) (first immunization). Then, an emulsion of the same amount of phospholipase A2 mixed with the equal volume of Freund's complete adjuvant was administered as a booster seven times at an interval of 2 to 3 weeks. Twenty-four days after the 7th. immunization, was administered peritoneally (final immunization) phospholipase A2 (20 μg per animal) dissolved in the physiological saline (0.2 ml). Three days after the final immunization, cells were collected from the spleen of a mouse, and suspended in the DMEM medium. c) Preparation of 12H5 hybridomas. 1.4 v 10.1 Spleen cells prepared, as specified above, were fused (1 x 108 cells) with P3 x 63Ag8 «U 1 (P3U 1) (2 x 107 cells) of mouse myeloma, according to the Kohier method and Milstein (see Nature, 256, 495 (1975)). That is, the basal cells and P3U 1 cells were washed several times with DMEM, placed together in a 50 ml plastic centrifuge tube and mixed thoroughly. right away, after the medium was removed through centrifugation, DMEM (1 ml) which contained 50% (w / v) of polyethylene glycol (Sigma, average molecular weight 3350) was slowly added, with stirring for 1 minute, and preheated to 37 ° C. Then, the DMEM (10 ml), preheated to 37 ° C, was added dropwise to the above mixture to complete the melting. The reaction mixture thus obtained was centrifuged to remove the supernatant, and to the residue was added HAT medium [DMEM medium containing 10% feral bovine serum supplemented with hypoxanthine (1 x 10"4 M), aminopterin (4). x 10"7 M) and thymidine (1.6 x 10" 5 M)] to adjust the concentration of 6 x 105 spleen cells / ml This cell suspension was distributed in cavities of a 96-well plastic plate 200 μl per cavity (1.2 x 105 spleen cells) Eleven days later, half volume of the medium was removed by aspiration, and the HAT medium described above was added to the residue seven to eleven days after the fusion. In the cell, proliferation of the hybridoma was observed in about half of the cavities.The activity of the antibody in the supernatant was analyzed by the method described in d), below.The positive clones were transferred to a plastic plate of 48%. cavities, and then au A 24 cavity plate, where they were grown in the same medium, except without containing aminopterin. The ability to reproduce the activity of the antibody was confirmed by using the culture supernatant in a 24-well plate through the method described in d), and at the same time, the inhibitory activity of the enzyme was determined by the method described in FIG. e). In order to know that the three clones, which show the inhibitory activity of enzyme, are monoclonal, a successive cloning was carried out limiting the dilution method. The monoclonal antibodies generated by the clones thus obtained were designated as 12H5, 1.4 and 10.1, respectively. d) Determination of the activity of the antibody through ELISA Purified, recombinant human type I phospholipase A2 was dissolved in saline regulated at its pH with 20 mM Tris (TBS, pH 7. 4) at 0.05 μg / ml. An aliquot (100 μl each) of the enzyme solution was placed in each well of a 96-well flat bottom microtiter plate and incubated in a humid chamber at 4 ° C overnight. Then, after discarding the solution, TBS containing 1% BSA (200 μl each) was added to each well, and left at 37 ° C for 1 hour to block the non-absorbed fraction in each well. Then, each cavity was washed several times with TBS containing 0.05% Tween 20 (TBS-Tween). The supernatant of the culture medium for the hybridoma culture or peritoneal cavity fluid, or a purified antibody preparation were diluted with TBS-Tween containing 0.2% BSA (BSA-TBS-Tween), and an aliquot (100 μl) was added. each) of the solution to each cavity, and incubated at room temperature for 2 hours. After washing the cavities with TBS-Tween similarly as described above, anti-mouse antibody conjugated with horseradish peroxidase (Zymed) diluted, 000 times with BSA-TBS-Twee was added to each cavity. (100 μl each), and incubated at room temperature for 2 hours. After the reaction, the wells were washed with TBS-Tween in a similar manner as above, and, as the substrate enzyme, were added to each well, pH buffer of 0.1 M sodium acetate-citrate (pH 5.8). ) (100 μl each) containing 0.006% hydrogen peroxide and 3,3 ', 5,5'-tetramethylbenzidine (0.1 mg / ml). After being left at room temperature for 30 minutes, 3 M of sulfuric acid (25 μl each) was added to terminate the reaction, then the optical density of the reaction mixture was measured at 450 nm. e) Assay of phospholipase A2 inhibitory activity The inhibitory activity of the antibody against phospholipase A2 was determined by a slight modification of the method described in Journal of Biological Chemistry (Pepinsky RB et al., J. Biol. Chem. 261 (9) , 4239-4246 (1986)) as follows. A purified preparation of recombinant human type II phospholipase A2 (0.5 ng) and the supernatant of a hybridoma growth medium, or an antibody preparation, was incubated in a pH regulator of 125 M Tris (pH 8.0) (100 μl) containing 150 mM sodium chloride, 12.5 mM calcium chloride and bovine serum albumin ((250 μg / ml) at room temperature for 2 hours, then an autoclave preparation of Escherichia coli SN 17, to which tritium-labeled oleic acid was added, was added to the reaction mixture (25 μl, 50,000 to 100,000 cpm), and the resulting mixture was incubated at 37 ° C. for 30 minutes. ice, and the reaction was terminated by adding 4 N hydrochloric acid (25 μl). After the reaction was finished, bovine serum albumin (40 mg / ml) was added to the mixture and mixed, and the mixture was added to the mixture. The result was allowed to stand on the ice for 30 minutes. After centrifugation at 15,000 rpm for 2 minutes, the radioactivity of the supernatant was measured. From the radioactivity thus obtained, this was removed from the negative control (radioactivity of a sample containing neither phospholipase A2 nor antibody), the resulting value was used to express the activity of phospholipase A2. The inhibitory relationship against the antibody was expressed according to the following calculation based on the activity of the phospholipase A2 of the positive control (phospholipase A2 activity of a sample containing said enzyme but without antibody). Inhibitory relationship against phospholipase A2 =. { 1 - (phospholipase A2 activity incubated with the supernatant of the culture medium or purified antibody) / (phospholipase A2 activity of the positive control)} x 100 According to the method described by Tokkai in H5-192167, p8, an autoclave preparation of £ was formed. coli, incorporated with tritium-labeled oleic acid, as a substrate for phospholipase A2 activity.

EXAMPLE 2 Generation v Preparation of the Monoclonal Antibody a) Generation of the monoclonal antibody 2.6, 10, 14-tetramethylpentadecane (pristan, Sigma) (0.5 ml) was injected into the peritoneal cavity of a mouse of the BALB / c strain (5 to 6 weeks after birth), and, 14 to 21 days later, a hybridoma (5 x 106) suspended in the physiological saline (0.5 ml) was injected peritoneally. Ten to twenty-one days later, the peritoneal fluid produced from the sacrificed and ventrotomized mouse was collected. The peritoneal fluid containing the monoclonal antibody was recovered by adding from 5 to 10 ml of a mouse. b) Preparation of the monoclonal antibody After centrifugation of the peritoneal fluid to remove the insoluble materials, an equal volume of a saturated ammonium sulfate solution was added, and the mixture was stirred for 1 hour on ice. The precipitate formed was collected by centrifugation, dissolved in a small volume of 0.1 M phosphate pH regulator (pH 7.4) containing 0.9% NaCl, and dialyzed against 100 volumes of the same pH regulator at 4 ° C. overnight to obtain a raw gamma globulin fraction. From this fraction, IgG was purified using a mouse monoclonal antibody MAPS-I I purification kit (BioRad Laboratories). That is, an equal volume of the binding pH regulator was added to the gamma globulin fraction and mixed. The resulting mixture was loaded onto a column filled with "Protein A-Sepharose CL4B (Pharmacia)", which was fully equilibrated with the same binding pH regulator (20 ml of gel bed volume), and the column was washed with 3 volumes of said binding pH regulator. IgG was eluted with approximately 3 column volumes of the elution pH buffer provided in the kit. Thus, the eluted IgG was dialyzed against, for example, 20 mM saline regulated at its pH with Tris (pH 7.4) to obtain the antibody preparation. Usually, 5 to 10 mg of IgG per 1 ml of peritoneal fluid was obtained.

EXAMPLE 3 Determination of the Antibody Subclass The subclass of antibody IgG in the supernatant of the hybridoma culture medium was determined using a mouse monoclonal antibody typing kit (Amersham). Through this method based on ELISA, it was determined that the subclasses of 12H 5, 1 .4 and 10.1 are IgG2a, IgG2a and IgG1, respectively.

EXAMPLE 4 Cross Reactivity of Antibodies a) Purification of rat type II phospholipase A2 To the rat PRP (platelet rich plasma) was added 1/5 volume of an ACD solution (2% (w / v) of a glucose solution containing 65 mM citric acid and 85 mM sodium citrate), and the mixture was centrifuged at 2,500 xg for 10 minutes. To the platelet thus obtained, a mixture of the medium of ACD / F10 (1: 5) (F 10, Gibco) was added to make a suspension containing 2 x 109 platelets per ml, and the resulting suspension was centrifuged at 2500 xg during 10 minutes. Then, the platelets thus precipitated were resuspended in the medium F 10 at the density of 2 x 109 cells per ml, and then, to the resulting suspension, 1 M of calcium chloride and thrombin was added successively (2, 500 units). / ml) at the final concentration of 2 mM and 2.5 units, respectively. The resulting mixture was incubated at 37 ° C for 5 minutes, centrifuged at 3,000 x g for 15 minutes to recover the supernatant (rat platelet supernatant stimulated by thrombin), which was subjected to the last procedure. Phospholipase A2 was purified through a slight modification of the method described by Tokkai in H 5-192167, p 12. That is, the supernatant of the rat platelet stimulated by thrombin was passed through a column (Econocolumn, column size of 20 ml; BioRad) of sulphated cellulofine (Seikagaku-Kogyo), which had been equilibrated with 0.1 M acetate pH regulator (pH 6.0). After the column was thoroughly washed with 200 ml of washing pH buffer (0.1 M acetate pH buffer containing 0.5 M NaCl (pH 6.0)), this was made with 30 ml of the pH regulator (0.1 M acetate pH regulator containing 1.5 M NaCl (pH 6.0)) to obtain the binding fraction of the sulfated cellulofine, which was fractionated through of H PLC. H PLC was performed in an H PLC LC-4A system (Shimazu Seisaku-sho) using a CAPCELL PAK C 18 column (4.6 mm ID x 25 cm, Shiseido) all at the flow rate of 1 ml / minute. The binding fraction of sulphated cellulofine was passed through the column, which was then washed by passing 0.1% trifluoroacetic acid for 60 minutes. Then, phospholipase A2 was eluted with a linear gradient of acetonitrile from 0 to 50% in the presence of 0.1% trifluoroacetic acid. Phospholipase A2, eluted with a peak of about 30% acetonitrile, was recovered. In SDS-polyacrylamide gel electrophoresis, the reaction thus recovered showed an individual protein band through silver staining, confirming this fraction as rat type I phospholipase A2. b) Preparation of type II phospholipase A2 derived from platelets of human, monkey, dog, rabbit, cat and mouse The type II phospholipase A2 derived from platelets of various animal species was purified according to the method for preparing platelet sonicate. of human being described in Journal Biological Chemistry (J. Biol. Chem., 264 (10), 5768-5775 (1989)). A PRP (platelet rich plasma) prepared from human, rhesus monkey, dog, rabbit, mouse and cat, EDTA was added to a final concentration of 2 mM, and the mixture was centrifuged at 2500 xg for 10 minutes to recover the cells The cells were suspended in pH buffer of 30 mM Tris HCl containing 120 mM NaCl and 2 mM EDTA (pH 7.4), and centrifuged at 2,500 x g for 10 minutes. After repeating the procedure twice, the cells thus obtained were resuspended in the pH buffer of 30 mM Tris HCl containing 120 mM NaCl and 2 mM EDTA (pH 7.4) at a density of 2 x 10 cells / ml, and frozen in liquid nitrogen. The frozen cell suspension was thawed, mixed with an equal volume of 0.36 N of sulfuric acid, and the sound was applied to the mixture using a BIOMC 7040 U LRA SON IC PROCESSOR (Seiko, output 80, 15 s, 6 times). The cell sonicate thus obtained was left on ice for 60 minutes, and then centrifuged at 4 ° C and 10,000 x g for 30 minutes to recover the supernatant. The precipitate was resuspended in 0.18 N of sulfuric acid, the volume of which was 1/3 of the volume of the supernatant, was left on ice for 60 minutes, and then centrifuged at 4 ° C and 10,000 xg for 30 minutes to recover the supernatant. Both supernatants were combined, and dialyzed against pH buffer of 50 mM acetate containing 200 mM NaCl (pH 4.5) overnight. The dialyzed sonicate was centrifuged at 4 ° C and 15,000 x g for 40 minutes to recover the supernatant, which was used as platelet-derived phospholipase A2.

ELISA Activity Purified preparations of various monoclonal antibodies obtained in Example 2b) were subjected to the following experiments. The IgG preparation was adjusted to a concentration of 10 μg / ml with BSA-TBS-Tween, and serially diluted 3 times. ELISA was performed with these serially diluted preparations according to Example 1 d), to examine the cross-reactivity to human type II phospholipase A2, recombinant and rat type II phospholipase A2. ELISA for rat type II phospholipase A2 was performed by replacing recombinant human type II phospholipase A2 in Example 1 d) with rat type I phospholipase A2. The results are shown in Figures 1 and 2, with the concentration of the antibody shown on the abscissa and the absorption at 450 nm on the ordinate. Cross reactivity was observed towards rat phospholipase A2, with 12H5 and 1.4, but not with 1 0. 1. d) Inhibitory activity against phospholipase A2 derived from several animals The purified preparations of several monoclonal antibodies obtained in Example 2b) were subjected to the following experiments. 1) Inhibitory activity against recombinant human type I I phospholipase A2. Using the recombinant human type I phospholipase A2, described in Example 1 a (actual, and in accordance with Example 1 e), the inhibitory activity of antibodies against phospholipase A2 was determined. The inhibitory ratios of each antibody to recombinant human type I phospholipase A2 are shown in Figure 3, with the concentrations of antibody samples pre-incubated with phospholipase A2 shown on the abscissa. 12H5 and 1 .4 almost completely inhibited the activity of phospholipase A2, at a concentration of 10 μg / ml. On the other hand, the inhibitory ratio of 10.1 reached 80%, at a concentration of 3 μg / ml. 2) Inhibitory activity against phospholipase A2 of rat type II. The inhibitory activity of rat type II phospholipase A2 was analyzed by replacing the human type II phospholipase A2, recombinant in Example 1e) with the purified preparation of rat type II phospholipase A2 (0.5 ng) in Example 4a), real. The inhibitory ratios of each antibody to the rat type II phospholipase A2 are shown in Figure 4, with the concentrations of antibodies pre-incubated with the phospholipase A2 sample shown on the abscissa. 12H5 and 1.4 also inhibited rat type II phospholipase A2, and the antibody concentration, required for 50% inhibition of phospholipase A2 activity, was 5 μg / ml with 12H5. 3) Inhibitory activity against type II phospholipase A2 derived from platelets of human, monkey, dog, rabbit, cat and mouse. The inhibitory activity against phospholipase A2 was analyzed by replacing recombinant human type II phospholipase A2 in real Example 1e) with a predetermined amount (20 to 40 μg / ml) of each platelet sonicate. The inhibitory ratios of each antibody against phospholipase A2 are shown in Figures 5 to 7, with the concentrations of the antibodies pre-incubated with the platelet sonicate shown on the abscissa. 12H5, 1.4 and 10.1 inhibited human platelet-derived phospholipase A2, similarly as recombinant human type II phospholipase A2, indicating that the recombinant human type II phospholipase A2 antibody of the present invention exhibited the activity of inhibiting recombinant human type II phospholipase A2. On the other hand, the antibodies of the present invention inhibited phospholipase A2 derived from rhesus monkey and mouse platelets, partially inhibited those derived from dog platelet, but not those derived from rabbit and cat tags.

EXAMPLE 5 Antibody Activity to Release Phospholipase A2 Type II Attached to the Cell Hepatocyte (BRL-3A, purchased from Dainippon Pharmaceutical) was cultured on a 96-well flat bottom plate (Falcon Co.) using F12K / 10% FCS (F12K, Dainihon Pharmaceutical Co.). After removing the culture medium from cells developed for co-incubation, F12K / 10% FCS (50 μl), containing recombinant human type II phospholipase A2, labeled with 125 I (4 ng), was added to the cells in the wells, and incubated at 37 ° C to bind the phospholipase A2 to the cells. After 1 hour, F12K / 10% FCS, containing a purified preparation of the antibody or F12K / 10% FCS (50 μl, each) was added only to the wells and incubated for another 2 hours. After incubation, the culture medium was recovered to measure its radioactivity. At the same time, the radioactivity of phospholipase A2 labeled with 12 SI (4 ng) was measured separately, which was added to the reaction system. The activity of the antibody to release phospholipase A2 from the cell was expressed as a percentage of the radioactivity of the medium incubated with various concentrations of antibody, by radioactivity of phospholipase A2 added minus radioactivity of the medium F 12 K / 10% FCS, which it was added only after cell binding to phospholipase A2, and incubated. Recombinant type I phospholipase A2 was labeled with 125 I by the following method. To a pH buffer of 100 mM Tris HCl (7.4) (100 μl), containing recombinant human type II phospholipase A2 (30 μg), placed in a glass test tube coated with IODOGEN (10 μg, Pierce ), 6 μl of Na125l (100 mCi / ml, ICN) was added, and the mixture was left at room temperature for 20 minutes. The reaction was terminated by bringing the reaction mixture out of the tube, and unreacted Na125l was removed by gel filtration. Phospholipase A2-linked cells were incubated with varying concentrations of the purified antibody preparation, and the radioactivity of the incubation medium was measured. The activity of each antibody to release the phospholipase A2 bound to the cell is shown in Figure 8, with the concentration of the antibody added in the medium, shown on the abscissa. The mouse antibody (control) did not release the phospholipase A2 from the cell, not at all, while 12H5, 10.1 and 1.4, depending on the concentration, released said enzyme from the cell.

EXAMPLE 6 Cloning of cDNA in the Variable Region of the H and L Chains of the Antibody Messenger RNA was prepared for the cloning of cDNA from about 10 7 hybridoma cells using a Micro QuickPrep mRNA Purification Kit (Pharmacia). That is, the cells were suspended in extraction pH regulator, from which the mRNA was isolated using Oligo (dT) -Cellulose. CDNA was prepared, corresponding to the variable region of the H and L chains of the 1.4 antibody, using a RACE System Kit (Gibco) according to the procedure shown in Figure 9. That is, using primers, which hybridize with a gene of constant region (C) and reverse transcriptase (Reverse Transcriptase SU PER SCRI PT II) with messenger RNA (1 μg) as a template, cDNA of individual chain structure was synthesized. The group of primers used were [GSP1 L (5'-GGCACCTCCAGATGTTAACTGC-3 ') / SEQ ID NO: 21] for the L chain, and [GSP 1 H (5'-GGAA (AG) TA (AGC) CCCTTGACCAGGC-3 ') / SEQ ID NO: 22] for the H chain. The sequences in parentheses in those of GSP 1 H, represent bases that correspond to a degeneration. Then, after the AR N m, which was a template, was digested with RNase H, cDNA of individual chain structure was purified using "GLASS MAX Spin Cartridge". The poly (dC) terminus was attached to the 3 'terminus of the individual structure chain A DNc using dCTP and terminal deoxynucleotide transferase. The variable region (V) genes of the H and L chains were amplified using an anchor primer, which hybridizes with the poly (dC) end, (5'-CUACUACUACUAGGCCACGCGTCGACTAGTACGGGI IGGGI IGGGIIG-37SEC ID NO: 23) and a initiator, which hybridizes to the constant region gene (C), [GSP2 L (5'- TATAGAGCTCAAGCTTGGATGGTGGGAAGATGGATACAGTTGGTGC-3 ') / SEQ ID NO: 24] in the case of the L chain, or [GSP2 H (5 * -TATAGAGCTCAAGCTTCCAGTGGATAGAC (CAT) GATGGGG (GC) TGT (TC) GTTTTGGC-3 ') / SEQ ID NO: 25] in the case of the H chain with LA Taq (Takara Shuzo). The sequences in parentheses, in those of GSP2H, represent bases that correspond to a degeneration. In a 1.2% agarose gel electrophoresis, these PCR products showed individual bands of both H and L chains, corresponding to around 550 to 570 base pairs. Also, DNcs corresponding to variable regions of the H and L chains of antibody 10.1 were prepared, according to the procedure shown in Figure 12, using an Amplification Kit of A DNc Marathon (Clontec). That is, cDNA of individual chain structure was synthesized using a Marathon cDNA synthesis primer and a reverse transcriptase with messenger RNA (1 μg) as a template. This reaction solution was further reacted with RNase H, DNA polymerase I and DNA ligase, to synthesize the double structure chain cDNA. Then, this reaction solution was reacted with DNA T4 polymerase to shave the term from the double-stranded chain cDNA. The Marathon cDNA adapter was ligated to the double-stranded chain cDNA using T4 ligase from A DN. The variable region (V) genes of the H and L chains were amplified using the adapter primer 1, which hybridizes with the adapter and initiator GSP2L (in the case of the L chain) or GSP2H (in the case of the H string) with LATaq (Takara Shuzo). In a 1.2% agarose gel electrophoresis, these PCR products showed individual bands corresponding to both H and L chains around 550 to 570 base pairs. For sequence, fragments amplified by PCR were integrated into the pCR ™ II vector using a TA Cloning Kit (Invitrogen). The competition cells of Escherichia coli JM 109 (Takara-shuzo) were transformed with the plasmid pCR ™ M inserted into the fragment. Using the colonies formed on the plate and [5 'UD primer (5'-ACCGAGCTCGGATCCACTAG-3') / SEQ ID NO: 26], which hybridizes with pCR ™ ll, and [3 'DU primer (5'-ATGCATGCTCGAGCGGCCGCC -3 ') / SEQ ID NO: 27], the colony PCR was performed with Taq polymerase (Takara-shuzo). Analyzing fragments of amplified DNA in 1.2% agarose gel electrophoresis, among colonies comprising amplified fragments of approximately 600 to 620 base pairs, three clones were selected for the L chain and four clones for the H chain of the 1.4 antibody, and two clones for the L chain and four clones for the H chain of the 2.1 antibody. The £ was cultivated. coli from each clone, and plasmid DNA was prepared using a QIAwell-8 plasmid purification kit (Qiagen). Using the plasmid DNA prepared from each clone, a UD primer, DU primer, GSP2L for the L chain, or GSP2 H for the H chain with a DNA Sequencing Kit (Parkin-Elmer), a sequencing reaction was performed, and it was analyzed by means of a Sequencer 373DNA (ABI). Three specific clones of the L chain and four specific clones of the H chain of the 1.4 antibody, and two specific clones of the L chain and four specific clones of the H chain comprised the same sequence, respectively. Figure 10 (SEQ ID NO: 1 and 2) which refers to the L chain of the antibody 1 .4, and Figure 1 1 (SEQ ID NO: 3 and 4) which refers to the H chain of the same antibody, show the cDNA sequence in the variable region and its putative amino acid sequence, respectively. Figure 13 (SEQ ID NO: 5 and 6) which refers to the L chain of the antibody 10.1, and Figure 14 (SEQ ID NO: 7 and 8) which refers to the H chain of the same antibody, show the sequence of A DNc in the variable region and its putative amino acid sequence, in each figure, the putative sequence corresponding to the CDR sequence stands out. (The putative CDR sequences of the L chain of the 1.4 antibody, the H chain of the 1.4 antibody, the L chain of the 10.1 antibody and the H chain of the 10.1 antibody are shown under SEQ ID NO: 9 to 11, SEC ID NO: 12 to 14, SEQ ID NO: 15 to 17, and SEQ ID NO: 18 to 20, respectively).

EXAMPLE 7 Homology Search of the Amino Acid Sequence in the Variable Region of the H and L Chains between Antibodies 1.4 and 10.1 and the Human Antibody The human antibody, which has a high homology to the amino acid sequences of the variable regions of the chains H and L of antibodies 1.4 and 10.1, obtained in Example 6, were searched in GenBank, databases EMBL, SWISS-PROT, PIR and PRF.

The homology search was performed using a BLASTP program 1. 4.8MP (Altschul S.F. et al, J. Mol. Biol., 215, 403-410 (1990)). As a result, antibody 1.4 was found to be highly homologous to, for example, Pag-1 antibody (Hughes-Jones N.C. et al., Biochem. J., 268, 135-140 (1990)), and WEA antibody (Goni).

F. et al., Proc. Nati Acad. Sci. USA; 80, 4837-4841 (1983)).

Also, antibody 10.1 was found to have high homology to, for example, ITH5-2 antibody (Chin LT et al., Immunol.Let, 44, 25-30 (1995)) and to antibody ITC48 (Ohlin M., And others, Mol. Immunol., 31, 983-991 (1994)).

Industrial Applicability The monoclonal antibody and the protein comprising a part thereof of the present invention are reactive with human type I phospholipase A2, and highly inhibitors against said enzyme. Since said antibody and protein are cross-reactive with type II phospholipase A2 derived from monkey and / or mouse platelet, and can be applied to these experimental animals, they can be advantageously used in the experiment with animals with monkey and / or mouse. before the pharmacological test in humans. In addition, since the monoclonal antibody and the protein comprising a part thereof of the present invention are capable of releasing type II phospholipase A2 bound to the cell, they are especially inhibitors for type II phospholipase A2, and presumably not induce side effects caused by complement attack and / or effector cell. The monoclonal antibody and the protein comprising a part thereof of the invention, can be applied as therapeutic agents for various diseases, which are conceived to be caused or exacerbated by human type II phospholipase A2 (including cardiac infarction, cerebral infarction). , acute renal failure, allergic disease, such as asthma, chronic rheumatism, osteoarthritis, septic shock, pancreatitis, psoriasis, multiple organ failure (MOF), acute respiratory distress syndrome (ARDS), Crohn's disease and ulcerative colitis, uveitis, syndrome of respiratory distress of the newborn (SVR), bronchopulmonary dysplasia (BPD), etc.).

SEQUENCE LIST INFORMATION FOR SEQ ID NO: 1 SEQUENCE CHARACTERISTICS: LENGTH: 393 TYPE: nucleic acid CHAIN STRUCTURE: double TOPOLOGY: linear TYPE OF MOLECULE: cDNA to mRNA ORIGINAL SOURCE: CELL LINE: 1.4 ASPECT: NAME / KEY: CDS LOCATION: 1 ... 393 IDENTIFICATION METHOD: SE DESCRIPTION SEQUENCE: SEQ ID NO: 1: ATG GAG ACÁ GAC ACÁ CTC CTG CTA TGG GTG CTG CTG CTC TGG GTT CCA 48 Met Glu Thr Asp Thr Leu Leu Leu Trp Val Leu Leu Leu Trp Val Pro 1 5 10 15 GGT TCC ACÁ GGT GAC ATT GTG CTG ACC CA TCT CCA GCT TCC TTG GCT 96 Gly Ser Thr Gly Asp lie Val Leu Thr Gln Ser Pro Wing Ser Leu Wing 20 25 30 GTG TCT TTA GGG CAG AGG GCC ACC ATA TCC TGC AGA GCC AGT GAA AGT 144 Val Ser Leu Gly Gln Arg Wing Thr lie Cys Arg Wing Ser Glu Ser 35 40 45 GTT GAT AGT TAT GGC ATT AGT TTT ATG CAC TGG TAT CAG CAG AAA CCA 192 Val Asp Ser Tyr Gly lie Ser Phe Met His Trp Tyr Gln Gln Lys Pro 50 55 60 GGA CAG CCC CCC AAA CTC CTC ATT TAT CGT GCA TCC AAC CTA GAA TCT 240 Gly Gln Pro Pro Lys Leu Leu lie Tyr Arg Ala Ser Asn Leu Glu Ser 65 70 75 80 GGG ATC CCT GCC AGG TTT AGT GGC AGT GGG TCT AGG ACA GAA TTC ACC 288 Gly lie Pro Wing Arg Phe Ser Gly Ser Gly Ser Arg Thr Glu Phe Thr 85 90 95 CTC ACC ATT AAT CCT GTG GAG GCT GAT GAT GTT GCA ACC TAT CAC TGT 336 Leu Thr lie Asn Pro Val Glu Wing Asp Asp Val Wing Thr Tyr His Cys 100 105 110 CAG CAA AGT AAT GAG GAT CCA TTC ACG TTC GGC TCG GGG AA TTG 384 Gln Gln Ser Asn Glu Asp Pro Phe Thr Phe Gly Ser Gly Thr Lys Leu 115 120 125 GAA ATA AAA 393 Glu lie Lys 130 INFORMATION FOR SEQ ID NO: 2 SEQUENCE CHARACTERISTICS: LENGTH: 131 TYPE: amino acid TOPOLOGY: linear TYPE OF MOLECULE: protein ORIGINAL SOURCE: CELL LINE: 1.4 SEQUENCE OF DESCRIPTION: SEQ ID NO: 2: Met Glu Thr Asp Thr Leu Leu Leu Trp Val Leu Leu Leu Trp Val Pro 1 5 10 15 Gly Ser Thr Gly Asp lie Val Leu Thr Gln Ser Pro Wing Ser Leu Wing 20 25 30 Val Ser Leu Gly Gln Arg Wing Thr lie Ser Cys Arg Wing Being Glu Being 40 45 Val Asp Ser Tyr Gly lie Ser Phe Met His Trp Tyr Gln Gln Lys Pro 50 55 60 Gly Gln Pro Pro Lys Leu Leu lie Tyr Arg Ala Ser Asn Leu Glu Ser 65 70 75 80 Gly Lie Pro Wing Arg Phe Ser Gly Ser Gly Ser Arg Thr Glu Phe Thr 85 90 95 Leu Thr lie Asn Pro Val Glu Wing Asp Asp Val Wing Thr Tyr His Cys 100 105 110 Gln Gln Ser Asn Glu Asp Pro Phe Thr Phe Gly Ser Gly Thr Lys Leu 115 120 125 Glu He Lys INFORMATION FOR SEQ ID NO: 3 SEQUENCE CHARACTERISTICS: LENGTH: 408 TYPE: nucleic acid CHAIN STRUCTURE: double TOPOLOGY: linear TYPE OF MOLECULE: cDNA to mRNA ORIGINAL SOURCE: CELL LINE: 1.4 ASPECT: NAME / KEY: CDS LOCATION: 1 ... 408 IDENTIFICATION METHOD: E SEQUENCE OF DESCRIPTION: SEQ ID NO: 3: ATG AGA GTC CTC ATT CTT TTC TGG CTC TTC ACÁ GCC TTT CCT GGT TTC 48 Met Arg Val Leu He Leu Leu Trp Leu Phe Thr Wing Phe Pro Gly Phe CTG TCT GAT GTC CAG CTT CAG GAA TCG GGA CCT GGC CTG GTG AAA CCT 96 Leu Ser Asp Val Gln Leu Gln Glu Ser Gly Pro Gly Leu Val Lys Pro 20 25 30 TCT CAA TCT CTG TCC CTC ACC TGC ATC GTC ACT GGC TAC TCA ATC ACC 144 Ser Gln Ser Leu Ser Leu Thr Cys Met Val Thr Gly Tyr Ser He Thr 35 40 45 AGT GAT TAT GCC TGG AAC TGG ATC CGG CAG TTT CCG GGA AAC AAA CTG 192 Ser Asp Tyr Wing Trp Asn Trp He Arg Gln Phe Pro Gly Asn Lys Leu 50 55 60 GAG CGG ATG GGA TAC ATA AGG TAC AGT GGT TAC ACT AGC TAC AAC CCA 240 Glu Arg Met Gly Tyr He Arg Tyr Ser Gly Tyr Thr Ser Tyr Asn Pro 65 70 75 80 TCT CTC AAA AGT CGA ATC TTT ATC ACG CGA GAC ACÁ TCC CAG AAC CAG 288 Ser Leu Lys Ser Arg He Phe He Thr Arg Asp Thr Ser Gln Asn Gln 85 90 95 TTC TTC CTA CAT TTG ACT TCT GTG ACT ACT GAG GAC ACÁ GCC ACA TAT 336 Phe Phe Leu His Leu Thr Ser Val Thr Thr Glu Asp Thr Ala Thr Tyr 100 105 110 TAC TGT ACÁ AGA GAC TTG GAC GCC TGG TAC TTC GAT GTT TGG GGC GCA 384 Tyr Cys Thr Arg Asp Leu Asp Wing Trp Tyr Phe Asp Val Trp Gly Wing 115 120 125 GGG ACÁ ACG GTC ACC GTC TCC TCA 408 Gly Thr Thr Val Thr Val Ser Ser 130 135 INFORMATION FOR SEQ ID NO: 4 SEQUENCE CHARACTERISTICS: LENGTH: 136 TYPE: amino acid TOPOLOGY: linear TYPE OF MOLECULE: cDNA to mRNA ORIGINAL SOURCE: CELL LINE: 1.4 SEQUENCE OF DESCRIPTION: SEQ ID NO: 4: Met Arg Val Leu He Leu Leu Trp Leu Phe Thr Wing Phe Pro Gly Phe 1 5 10, 15 Leu Ser Asp Val Gln Leu Gln Glu Ser Gly Pro Gly Leu Val Lys Pro 20 25 30 Ser Gln Ser Leu Ser Leu Thr Cys Met Val Thr Gly Tyr Ser He Thr 40 45 Ser Asp Tyr Ala Trp Asn Trp He Arg Gln Phe Pro Gly Asn Lys Leu 50 55 60 Glu Arg Met Gly Tyr He Arg Tyr Ser Gly Tyr Thr Ser Tyr Asn Pro 65 70 75 80 Be Leu Lys Be Arg He Phe He Thr Arg Asp Thr Ser Gln Asn Gln 85 90 95 Phe Phe Leu His Leu Thr Ser Val Thr Thr Glu Asp Thr Ala Thr Tyr 100 105 110 Tyr Cys Thr Arg Asp Leu Asp Wing Trp Tyr Phe Asp Val Trp Gly Wing H5 120 125 Gly Thr Thr Val Thr Val Ser Ser INFORMATION FOR SEQ ID NO: 5 SEQUENCE CHARACTERISTICS: LENGTH: 381 TYPE: nucleic acid STRUCTURE OF CHAIN: double TOPOLOGY: linear TYPE OF MOLECULE: cDNA to mRNA ORIGINAL SOURCE: CELL LINE: 10.1 ASPECT: NAME / KEY: CDS LOCATION: 1 ... 381 IDENTIFICATION METHOD: E SEQUENCE OF DESCRIPTION: SEQ ID NO: 5: ATG GAA TCA CAG ACT CTG GTC TTC ATA TCC ATA CTC CTC TGG TTA TAT 48 Met Glu Ser Gln Thr Leu Val Phe He Ser He Leu Leu Trp Leu Tyr 1 5 10 15 GGA GCT GAT GGG AAC ATT GTA ATG ACC CA TCT CCC AAA TCC ATG TCC 96 Gly Wing Asp Gly Asn He Val Met Thr Gln Ser Pro Lys Ser Met Ser 20 25 30 ATG TCA GTA GGA GAG AGG GTC ACC TTG ACC TGC AAG GCC AGT GAG AAT 144 Met Ser Val Gly Glu Arg Val Thr Leu Thr Cys Lys Wing Ser Glu Asn 35 40 45 GTG GTT ACT TAT GTT TCC TGG TAT CAA CAG AAA CCA GAG CAG TCT CCT 192 Val Val Thr Tyr Val Ser Trp Tyr Gln Gln Lys Pro Glu Gln Ser Pro 50. 55 60 AAA CTC CTG ATA TAC GGG GCA TCC AAC CGG TAC ACT GGG GTC CCC GAT 240 Lys Leu Leu He Tyr Gly Ala Ser Asn Arg Tyr Thr Gly Val Pro Asp 65 70 75 80 CGC TTC ACA GGC AGT GGA TCT GCA ACA GAT TTC ACT CTC ACC ATC AGC 288 Arg Phe Thr Gly Ser Gly Ser Wing Thr Asp Phe Thr Leu Thr He Ser 85 90 95 AGT GTG CAG GCT GAA GAC CTT GCA GAT TAT CAC TGT GGA CAG GGT TAC 336 Ser Val Gln Wing Glu Asp Leu Wing Asp Tyr His Cys Gly Gln Gly Tyr 100 105 110 AGC TAT CCG TGA ACG TTC GGA GGG GGG ACC AAG CTG GAA ATA AAA 381 Ser Tyr Pro Tyr Thr Phe Gly Gly Gly Thr Lys Leu Glu He Lys 115 120 125 INFORMATION FOR SEQ ID NO: 6 SEQUENCE CHARACTERISTICS: LENGTH: 127 TYPE: amino acid TOPOLOGY: linear TYPE OF MOLECULE: protein ORIGINAL SOURCE: CELL LINE: 10.1 SEQUENCE OF DESCRIPTION: SEQ ID NO: 6: Met Glu Ser Gln Thr Leu Val Phe He Ser He Leu Leu Trp Leu Tyr 1 5 10 15 Gly Wing Asp Gly Asn He Val Met Thr Gln Ser Pro Lys Ser Met Ser 20 25 30 Met Ser Val Gly Glu Arg Val Thr Leu Thr Cys Lys Wing Ser Glu Asn 40 45 Val Val Thr Tyr Val Ser Trp Tyr Gln Gln Lys Pro Glu Gln Ser Pro 50 55 60 Lys Leu Leu He Tyr Gly Ala Ser Asn Arg Tyr Thr Gly Val Pro Asp 65 70 75 80 Arg Phe Thr Gly Ser Gly Be Wing Thr Asp Phe Thr Leu Thr He Ser 85 90 95 Ser Val Gln Wing Glu Asp Leu Wing Asp Tyr His Cys Gly Gln Gly Tyr 100 105 110 Ser Tyr Pro Tyr Thr Phe Gly Gly Gly Thr Lys Leu Glu He Lys 115 120 125 INFORMATION FOR SEQ ID NO: 7 SEQUENCE CHARACTERISTICS: LENGTH: 414 TYPE: nucleic acid CHAIN STRUCTURE: double TOPOLOGY: linear TYPE OF MOLECULE: cDNA to mRNA ORIGINAL SOURCE: CELL LINE: 10.1 ASPECT: NAME / KEY: CDS LOCATION: 1 ... 414 IDENTIFICATION METHOD: E SEQUENCE OF DESCRIPTION: SEQ ID NO: 7: ATC GCT GTC CTG GCA TTA CTT TTT TGC CTG GTA ACA TTC CCA AGC TGT 48 Met Wing Val Leu Wing Leu Leu Phe Cys Leu Val Thr Phe Pro Ser Cys 1 5 10 15 ATC CTT TCC CAG GTG CAG CTG AAG GAG TCA GGA CCT GGC CTG GTG GCG 96 He Leu Ser Gln Val Gln Leu Lys Glu Ser Gly Pro Gly Leu Val Wing 20 25 30 CCC TCA CAG AGC CTG TCC ATC ACE TGT ACC GTC TCA GGG TTC TCA TCA 144 Pro Ser Gln Ser Leu Ser He Thr Cys Thr Val Ser Gly Phe Ser Leu 35 40 45 ACC GAC TTT GGT ATA TGG GTT CGC CAG CCT CCA GGA AAG GGT CTG 192 Thr Asp Phe Gly Val Asn Trp Val Arg Gln Pro Pro Gly Lys Gly Leu 50 55 60 GAG TGG CTG GGA ATG ATA TGG ACT GAT GGA ATC ACA GAC TAT AAT TCA 240 Glu Trp Leu Gly Met He Trp Thr Asp Gly He Thr Asp Tyr Asn Ser 65 • 70 75 80 GTT CTC AAA TCC AGA CTG AGC ATC AGC AAG GAC ACC TCC AAG AGC CAA 288 Val Leu Lys Ser Arg Leu Ser He Ser Lys Asp Thr Ser Lys Ser Gln 85 90 95 GTT TTC TTC AAA ATG AAC AAT CTG CAA ACT GAT GAC ACÁ GCC AGG TAC 336 Val Phe Leu Lys Met Asn Asn Leu Gln Thr Asp Asp Thr Wing Arg Tyr 100 105 110 TAC TGT GCC AGA GAT GCA TAC TAC GGC TTC TAT TAC GCT TAC TGG 384 Tyr Cys Wing Arg Asp Wing Tyr Tyr Gly Phe Tyr Wing Met Asp Tyr Trp 115 120 125 GGT CAA GGA ACC TCA GTC ACC GTC TCC TCA 414 Gly Gln Gly Thr Ser Val Thr Val Ser Ser 130 135 INFORMATION FOR SEQ ID NO: 8 SEQUENCE CHARACTERISTICS: LENGTH: 138 TYPE: amino acid TOPOLOGY: linear TYPE OF MOLECULE: protein ORIGINAL SOURCE: CELL LINE: 10.1 SEQUENCE OF DESCRIPTION: SEC ID NO: 8: Met Wing Val Leu Wing Leu Leu Phe Cys Leu Val Thr Phe Pro Ser Cys 1 5 10 15 He Leu Ser Gln Val Gln Leu Lys Glu Ser Gly Pro Gly Leu Val Wing 20 25 30 Pro Ser Gln Ser Leu Ser He Thr Cys Thr Val Ser Gly Phe Ser Leu 35 40 45 Thr Asp Phe Gly Val Asn Trp Val Arg Gln Pro Pro Gly Lys' Gly Leu 50 55 60 Glu Trp Leu Gly Met He Trp Thr Asp Gly He Thr Asp Tyr Asn Ser 65 70 75 80 Val Leu Lys Ser Arg Leu Ser He Ser Lys Asp Thr Ser Lys Ser Gln 85 90 95 Val Phe Leu Lys Met Asn Asn Leu Gln Thr Asp Asp Thr Wing Arg Tyr 100 105 110 Tyr Cys Wing Arg Asp Wing Tyr Tyr Gly Phe Tyr Wing Met Asp Tyr Trp 115 120 125 Gly Gln Gly Thr Ser Val Thr Val Ser Being INFORMATION SEQ ID NO: 9 SEQUENCE CHARACTERISTICS: LENGTH: 15 TYPE: amino acid TOPOLOGY: linear TYPE OF MOLECULE: protein ORIGINAL SOURCE: CELL LINE: 1.4 SEQUENCE OF DESCRIPTION: SEQ ID NO: 9: Arg Ala Ser Glu Ser Val Asp Ser Tyr Gly lie Ser Phe Met His 1 5 10 15 INFORMATION FOR SEQ ID NO: 10 SEQUENCE CHARACTERISTICS: LENGTH: 7 TYPE: amino acid TOPOLOGY: linear TYPE OF MOLECULE: protein ORIGINAL SOURCE: CELL LINE: 1.4 SEQUENCE OF DESCRIPTION: SEQ ID NO: 10: Arg Ala Ser Asn Leu Glu Ser 1 5 INFORMATION FOR SEQ ID NO: 11 SEQUENCE CHARACTERISTICS: LENGTH: 9 TYPE: amino acid TOPOLOGY: linear TYPE OF MOLECULE: protein ORIGINAL SOURCE: CELL LINE: 1.4 SEQUENCE OF DESCRIPTION: SEQ ID NO: 11 Gln Gln Ser Asn Glu Asp Pro Phe Thr 1 5 INFORMATION FOR SEQ ID NO: 12 SEQUENCE CHARACTERISTICS: LENGTH: 6 TYPE: amino acid TOPOLOGY: linear TYPE OF MOLECULE: protein ORIGINAL SOURCE: CELL LINE: 1.4 SEQUENCE OF DESCRIPTION: SEQ ID NO: 12: Ser Asp Tyr Ala Trp Asn 1 5 INFORMATION FOR SEQ ID NO: 13 SEQUENCE CHARACTERISTICS: LENGTH: 16 TYPE: amino acid TOPOLOGY: linear TYPE OF MOLECULE: protein ORIGINAL SOURCE: CELL LINE: 1.4 SEQUENCE OF DESCRIPTION: SEQ ID NO: 13: Tyr lie Arg Tyr Ser Gly Tyr Thr Ser Tyr Asn Pro Ser Leu Lys Ser 1 5 10 15 INFORMATION FOR SEQ ID NO: 14 SEQUENCE CHARACTERISTICS: LENGTH: 9 TYPE: amino acid TOPOLOGY: linear TYPE OF MOLECULE: protein ORIGINAL SOURCE: CELL LINE: 1.4 SEQUENCE OF DESCRIPTION: SEQ ID NO: 14: Asp Leu Asp Ala Trp yr Phe Asp Val 1 5 INFORMATION FOR SEQ ID NO: 15 SEQUENCE CHARACTERISTICS: LENGTH: 11 TYPE: amino acid TOPOLOGY: linear TYPE OF MOLECULE: protein ORIGINAL SOURCE: CELL LINE: 10.1 SEQUENCE OF DESCRIPTION: SEQ ID NO: 15: Lys Ala Ser Glu Asn Val Val Thr Tyr Val Ser 1 5 10 INFORMATION FOR SEC ID NO: 16 SEQUENCE CHARACTERISTICS: LENGTH: 7 TYPE: amino acid TOPOLOGY: linear TYPE OF MOLECULE: protein ORIGINAL SOURCE: CELL LINE: 10.1 SEQUENCE OF DESCRIPTION: SEQ ID NO: 16: Gly Ala Ser Asn Arg Tyr Thr 1 5 INFORMATION FOR SEQ ID NO: 17 SEQUENCE CHARACTERISTICS: LENGTH: 9 TYPE: amino acid TOPOLOGY: linear TYPE OF MOLECULE: protein ORIGINAL SOURCE: CELL LINE: 10.1 SEQUENCE OF DESCRIPTION: SEQ ID NO: 17: Gly Gln Gly Tyr Ser Tyr Pro Tyr Thr 1 5 INFORMATION FOR SEQ ID NO: 18 SEQUENCE CHARACTERISTICS: LENGTH: 5 TYPE: amino acid TOPOLOGY: linear TYPE OF MOLECULE: protein ORIGINAL SOURCE: CELL LINE: 10.1 SEQUENCE OF DESCRIPTION: SEQ ID NO: 18: Asp Phe Gy Val Asn 1 5 INFORMATION FOR SEQ ID NO: 19 SEQUENCE CHARACTERISTICS: LENGTH: 16 TYPE: amino acid TOPOLOGY: linear TYPE OF MOLECULE: protein ORIGINAL SOURCE: CELL LINE: 10.1 SEQUENCE OF DESCRIPTION: SEQ ID NO: 19: Met lie Trp Thr Asp Gly lie Thr Asp Tyr Asn Ser Val Leu Lys Ser 1 5 10 15 INFORMATION FOR SEC ID NO: 20 SEQUENCE CHARACTERISTICS: LENGTH: 11 TYPE: amino acid TOPOLOGY: linear TYPE OF MOLECULE: protein ORIGINAL SOURCE: CELL LINE: 10.1 SEQUENCE OF DESCRIPTION: SEQ ID NO: 20: Asp Ala Tyr Tyr Gly Phe Tyr Ala Met Asp Tyr 1 5 10 INFORMATION FOR SEQ ID NO: 21 SEQUENCE CHARACTERISTICS: LENGTH: 22 TYPE: nucleic acid TOPOLOGY: linear TYPE OF MOLECULE: other nucleic acid (synthetic) SEQUENCE OF DESCRIPTION: SEQ ID NO: 21: GGCACCTCCA GATGTTAACT GC 22 INFORMATION FOR SEQ ID NO: 22 SEQUENCE CHARACTERISTICS: LENGTH: 20 TYPE: nucleic acid TOPOLOGY: linear TYPE OF MOLECULE: other nucleic acid (synthetic) SEQUENCE OF DESCRIPTION: SEQ ID NO: 22: GAARTAVCCC TTGACCAGGC 20 INFORMATION FOR SEQ ID NO: 23 SEQUENCE CHARACTERISTICS: LENGTH: 48 TYPE: nucleic acid TOPOLOGY: linear TYPE OF MOLECULE: other nucleic acid (synthetic) ASPECT: LOCATION: 36,37,41,42,46,47 OTHER INFORMATION: N it is inosine (I).

SEQUENCE OF DESCRIPTION: SEQ ID NO: 23: CUACUACUAC UAGGCCACGC GTCGACTAGT ACGGGNNGGG NNGGGNNG 48 INFORMATION FOR SEQ ID NO: 24 SEQUENCE CHARACTERISTICS: LENGTH: 46 TYPE: nucleic acid TOPOLOGY: linear TYPE OF MOLECULE: other nucleic acid (synthetic) SEQUENCE OF DESCRIPTION: SEQ ID NO: 24:AAGCTTGGAT GGTGGGAAGA TGGATACAGT TGGTGC 46 INFORMATION FOR SEQ ID NO: 25 SEQUENCE CHARACTERISTICS: LENGTH: 50 TYPE: nucleic acid TOPOLOGY: linear TYPE OF MOLECULE: other nucleic acid (synthetic) SEQUENCE OF DESCRIPTION: SEQ ID NO: 25: TATAGAGCTC AAGCTTCCAG TGGATAGACH GATGGGGSTG TYGTTTGGC 50 INFORMATION FOR SEQ ID NO: 26 SEQUENCE CHARACTERISTICS: LENGTH: 20 TYPE: nucleic acid TOPOLOGY: linear TYPE OF MOLECULE: other nucleic acid (synthetic) SEQUENCE OF DESCRIPTION: SEQ ID NO: 26: ACCGAGCTCG GATCCACTAG_20_INFORMATION FOR SEQ ID NO: 27 SEQUENCE CHARACTERISTICS: LENGTH: 21 TYPE: nucleic acid TOPOLOGY: linear TYPE OF MOLECULE: other nucleic acid (synthetic) SEQUENCE OF DESCRIPTION: SEQ ID NO: 27: ATGCATGCTC GAGCGGCCGC C 21

Claims (5)

1. CLAIMS 1 .- A monoclonal antibody capable of inhibiting the activity of human type II phospholipase A2, as well as that of type II phospholipase A2 derived from monkey and / or mouse, or a protein comprising a part thereof having said inhibitory activity .
2. 2. A monoclonal antibody capable of releasing type II I phospholipase A2 bound to the cell membrane, or a protein comprising a part thereof having said activity.
3. 3. The monoclonal antibody or protein according to claim 2, wherein said phospholipase A2 is derived from being human.
4. 4. A monoclonal antibody capable of inhibiting not only the human type II phospholipase A2 activity, as well as that of type II phospholipase A2 derived from monkey and / or mouse, but also of releasing bound type II phospholipase A2 to the membrane of the cell, or a protein comprising a part thereof having said activity.
5. 5. A monoclonal antibody produced through any of the hybridomas 12H5 (FERM BP-5300), 1.4 (FERM BP-5297) and 10.1 (FERM BP-5298), or a protein comprising a part thereof, or a monoclonal antibody or a protein comprising a part thereof having activity against type II phospholipase A2 equivalent to that of said monoclonal antibody or protein. 6. - The monoclonal antibody or protein comprising a part thereof according to claim 1 or 2, wherein said monoclonal antibody or protein comprises a protein having the amino acid sequence represented by any of SEQ ID NO: 2, 4, 6 or 8, or those modified by substitution, deletion or insertion of one or more amino acid residues contained in said sequence. 7. The monoclonal antibody or protein comprising a part thereof according to claim 1 or 2, wherein said monoclonal antibody or protein comprises a protein having the amino acid sequence represented by any of SEQ ID NO: 9. to 20, or those modified by substitution, elimination or insertion of one or more amino acid residues contained in said sequence. 8. A cell that produces the monoclonal antibody or protein according to any of claims 1 to 7. 9. The cell according to claim 8, wherein said cell is a hybridoma. 10. The cell according to claim 8, wherein said cell is a cell transformed with recombinant DNA. 1 - A method for producing the monoclonal antibody or protein according to any of claims 1 to 7, wherein said method comprises methods for cu ivating the cell according to claim 8, and recovering the monoclonal antibodies or proteins of the cell. supernatant of the growth medium. 12. A DNA encoding the monoclonal antibody or protein according to any of claims 1 to 7. 13. The DNA according to claim 12, wherein said DNA comprises the base sequence with any of SEQ ID NO. : 1, 3, 5 or 7, or that modified by substitution, elimination or insertion of one or more amino acid residues contained in said sequence. 14. A recombinant vector comprising the DNA according to claim 12 or 13. 15. A medical composition comprising the monoclonal antibody or protein according to any of claims 1 to 7, and a pharmacologically acceptable carrier. 16. An inhibitor of type II phospholipase A2 comprising the monoclonal antibody or protein according to any of claims 1 to 7.