Abstract:
The invention relates to the field of medical diagnostics, immunology and oncology, in particular to the new tumor markers, useful in the early diagnosis of cancer and methods for cancer diagnostics. A new antigen for ELISA to detect antibodies associated with malignancies, namely, mature human plasminogen or human plasminogen fragments containing kringles. We propose a method of diagnosing cancer at an early stage by detecting autoantibodies of type IgA, IgG, IgM to human plasminogen or its fragments in a samples of human blood plasma or sera.

Description:
CROSS-REFERENCE TO RELATED APPLICATIONS 
       [0001]    This application is a national stage application of PCT/RU2012/001148 filed on Dec. 29, 2012 which claims priority to Russian application RU2012148244 filed on Nov. 14, 2012 currently issued as a patent RU2522231. 
       FIELD OF THE INVENTION 
       [0002]    The present invention relates to the field of medical diagnostics, immunology and oncology, in particular to the discovering of new tumor markers. The invention is to the early diagnostics of cancer. More specifically, the present invention relates to one of the new universal tumor markers of neoplastic processes, namely, the autoantibodies against plasminogen or its fragments produced in the human body during the growth of tumor and its progression. The invention also relates to methods for diagnosing cancer by detecting these autoantibodies in a sample of human blood plasma. A Kit of enzyme immunoassay is proposed for detection of autoantibodies. The invention also relates to antigens, which are interacted with human autoantibodies that these antigens will use in the diagnostic of cancer. 
       Terminology 
       [0003]    Technical and scientific terms used in the description, have the same meaning and value, which are commonly used in the relevant areas of science and technology. 
         [0004]    The term “antigen” as used herein, refers to proteins or fragments thereof, capable of binding with antibodies. 
         [0005]    The term “kringle” refers to a protein domain that has a structure stabilized by three disulfide bonds. 
         [0006]    The term “domain” refers to a section of the protein, which is characterized by certain structural and functional properties. 
         [0007]    The term “immunoassay” refers to methods of identifying high-molecular compounds, comprising the steps of: (a) the step of contacting the antigen with a biological sample under conditions suitable for the formation of antigen-antibody complexes, and (b) the stage of detection of these complexes. 
         [0008]    The term “tumor marker” refers to a high-molecular compounds defined structure, revealing that in the samples of human tissue is associated with cancer. 
         [0009]    The term “epitope” in the present invention refers to a section of the protein molecule that can form a bond with the antibody. 
         [0010]    The term “human antibody” refers to an antibody having an amino acid sequence that corresponds to the amino acid sequence of the antibody produced by man. 
         [0011]    The term “autoantibodies” (auto-aggressive antibodies, autologous antibodies)—antibodies capable to react with autoantigens, that is, with the body&#39;s own antigens. 
     
    
     
       BRIEF DESCRIPTION OF DRAWINGS 
         [0012]      FIG. 1 . The primary structure of Human plasminogen. 
           [0013]      FIG. 2 . Scheme of transformation of Glu-plasminogen to the heavy (K1-5+30 r) and light chain (micro-plasmin). 
       
    
    
     DESCRIPTION OF THE RELATED ART 
       [0014]    The search of new tumor markers for the diagnosis of cancer in the early stages of the pathological process is one of important approach in the battle with cancer. The beginning of neoplastic processes may be due to various causes, so specific diagnosis requires to use many different markers with specificity to each type of tumor. Thus, the most useful in the early detection of cancer are the universal tumor markers, the detection of which is associated with a starting neoplastic process any kind. 
         [0015]    According to the studies of Folkman, the balance of angiogenic and antiangiogenic systems should be stable for normal development of the organism. During normal processes such as organogenesis in the embryo and wound healing in the adult, angiogenesis provides the necessary vascular support for the newly developing tissue. It is known that tumor tissue contains much more blood vessels and capillaries than the surrounding healthy tissue. For these vessels to the fast-growing tumor cells enter the nutrients and oxygen that need to divide. The tumor growth and lethality are dependent upon angiogenesis and that angiogenesis inhibition suppresses tumor development (Folkman J. Angiogenesis in cancer, vascular, rheumatoid and other disease. Nat Med 1995; 1:27-31, Folkman J. Tumor angiogenesis. Adv Cancer Res 1985; 43:175-203). 
         [0016]    It is known a minimal amount of cancer cells are appeared in the tissues, but organism is resistant of them, though some conditions would be suitable for their development (sufficient number of vessels, appropriate microenvironment, and their volume is greater than 2 mm 2) the tumor will begin to grow rapidly. 
         [0017]    The role of serine proteases in oncogenesis and other systemic diseases is actively investigated. Secretion of serine proteases by tumor cells leads to the destruction of normal tissue architecture, invasion of these cells into the host tissue. These enzymes, both endogenous and exogenous, can remove from the cell surface specific proteins, which leads to the change in the adhesive properties of cells. Serine proteases secreted from tumor tissue interstitial fluid, can destroy the primary capillary membrane, facilitating the penetration of tumor cells through the vessel wall to the tissues of the host and the formation of metastases. Plasminogen/plasmin takes special attention in the class of serine proteases involved in process of oncogenesis whereas it has antiangiogenic properties too. Angiostatin and some other products of degradation of plasminogen are components of antiangiogenic plasminogen system. 
         [0018]    Plasminogen is inactive precursor of plasmin. Plasmin is a endopeptidase with trypsin-like serine protease activity. Physiological action of plasmin is to control the balance of coagulation system. Plasmin is usually performed from the plasminogen by activation of streptokinase, urokinase in vitro. Native glu-plasminogen is readily converted to lys-plasminogen by plasmin hydrolysis of the Lys-77-Lys-78 bond. The system of plasminogen/plasmin is active not only in the process of fibrinolysis, but is closely associated with process of carcinogenesis. It was demonstrated a strong relationship between plasmin and metalloproteases, which are active components in carcinogenesis (Yves A. DeClerck and Walter E. Laug: Plasminogen: Structure, Activation, and Regulation, edited by David M. Waisman. Kluwer Academic/Plenum Publishers, New York, 2003). Apart from a native molecule of plasminogen (plasmin), a whole range of products of its degradation are functionally significant. Effect of degradated forms of plasmin to small molecule substrates may exceed of whole molecule (J G Klys, N V Zaitseva, A I Kizim, SV rope Proteolytic derivatives of plasminogen in the development of cancer, oncology, T 12, No 1, 2010). 
         [0019]    In the process of degradation of the molecule plasminogen (plasmin), light and heavy chains can be obtained. The light chain contains the active site of plasmin, which is characteristic for the whole class of serine proteases. The heavy chain contains five kringles (triple disulfide-linked loops) regions. Each of these kringles or combination has its functional specialization. There are variants of the existence of the kringles in plasma: K1-3; K2-3; K1-4; K1-4,5; K1-5 and some single Kringles (Perri S, Martineau D, Francois M, et al. Plasminogen kringle 5 blocks tumor progression by antiangiogenic and proinflammatory pathways, Mol Cancer Ther 2007; 6:441-9). Well known that all Kringles, and combinations thereof are actively involved in angiogenesis and tumorigenesis. (Cao et al., J. Biol. Chem. 271:29461-29467, 1996; U.S. Pat. No. 6,024,688 to Folkman et al.) The functional activity of the first four kringle (K1-4) was widely investigated. The sequence of the kringles 1-4 is named angiostatin. (Francis J. Castellino, Victoria A. Ploplis, Structure and function of the plasminogen/plasmin system, Thromb Haemost 2005; 93:647-54; C. Boccaccio and Paolo M. Comoglio Cancer Res 2005; 65(19):8579-82; Rijken D C, Lijnen H R. New insights into the molecular mechanisms of the fibrinolytic system. J Thromb Haemost 2009; 7:4-13). 
       DETAILED DESCRIPTION OF THE INVENTION 
       [0020]    Authors performed a preliminary study comparing blood samples of cancer patients and controls using two-dimensional electrophoresis followed by mass spectrometry procedure of interesting spots. Preliminarily, samples were prepared using lys-sepharose. It was found a concentrations of some proteins in cancer patients are significantly much higher than in the controls. 90% of these proteins after mass-spectrometric identification were identified as fragments of plasminogen with a molecular mass of less than 55 kDa and immunoglobulins. 
         [0021]    The inventors have suggested that in in the area of tumor growth certain peptides are formed in high concentrations that may lead to the production of autoantibodies. These autoantibodies, in turn, can have an inhibitory effect on angiostatin and other derivatives of plasminogen and thus convert the balance in favor of angiogenic system. The neovessel organization is one of the conditions leading to the rapid growth of the tumor. Experiments performed by the inventors have shown the role of human autoantibodies in early detection of of the tumor growth. There are autoantibodies against their own plasminogen and various products of its degradation, particularly angiostatin. Increased titer of autoantibodies to plasminogen and or its degradation products in the plasma is a marker of tumor at an early stage and measurement of the level of autoantibodies to plasminogen and/or its degradation products in the plasma is diagnostic factor of developing cancer. 
         [0022]    Tumors of internal organs do not have clear symptoms at early pathological growth usually. Malignant growth used to begin with chronic inflammation, without striking symptoms. Symptoms depend on the location and size of the cancer, as well as how amazed surrounding organs and tissues of the human body. Already formed a malignant tumor in stage I and II growth is painless and has no pronounced symptoms. 
         [0023]    Nonspecific symptoms were taken in attention thorough examination of the patient, including through laboratory studies of blood plasma for early detection of cancer markers of the present invention. 
         [0024]    Since autoantibodies are polyclonal and plasminogen molecule contains many epitopes, the inventors propose to use different parts of the plasminogen molecule to determine the titer of autoantibodies for early detection of cancer. Currently, there are no published sources data revealing the correlation between the appearance of autoantibodies in the blood plasma to plasminogen and/or its degradation products in cancer. 
         [0025]    Plasminogen is a single-chain glycoprotein present in plasma at a concentration of about 2 mcM (Wahl et al., Thromb. Res. 27:523-535, 1982; Kang et al., Trends Cardiovasc. Med. 90:92-102, 1999). Plasminogen contains 791 amino acid residues and 24 disulfide bonds. Protein consists of a single polypeptide chain, where N-terminal amino acid is glutamine, C-terminal asparagine. The structure of the molecule has 2%-3% of carbohydrates, which are localized in the heavy chain. Oligosaccharides attached to Asp288 and Tre345. Plasminogen is a precursor of plasmin which is formed by cleavage of plasminogen between Arg-561 and Val-562 by tissue plasminogen activator or urokinase-type plasminogen activator. In the process of activation of plasminogen bond Arg560-Val561 is cleaved and two chains are formed, light and heavy, connected by disulfide bonds. The light chain (Val561-Asn790) has an active protease center, including the amino acid sequence of serine, histidine, asparagine. The heavy chain of plasmin (Lys78-Arg560) has five triple disulfide-linked loops known as kringle regions—or kringle domains, which is a compact globular structure with a hydrophobic core. These structures are involved in the process of protein interactions in blood clotting. Both plasminogen and plasmin bind to fibrin through amino-terminal kringle regions each of which is a triple loop region formed as a result of disulfide bonds. Kringles of heavy chain named K1, K2, K3, K4, K5. Kringles 1-4 have domains, specific areas, which have a strong affinity for lysine, ε-aminocaproic acid, parabens, and other ω-carbon amino acids having antifibrinolytic properties. Lysine binding sites (LBS) play an important role in the interaction between plasmin (plasminogen) to fibrin and plasmin to inhibitor—a2-AP (antiplasmin). Any fragment of human plasminogen containing any kringles can be used to detect autoantibodies associated with cancer, there is no matter whether a product of cleavage is from natural plasminogen or produced by splitting the plasminogen in vitro (eg, by enzymatic action). 
         [0026]    Full-size human plasminogen and various products of its cleavage: light or heavy chain, and any of the fragments containing kringles, can be used as an antigens to produce a set of immunoassay for determination of autoantibodies classes IgG, IgA, and IgM in the samples of human plasma or sera. These antigens are derived from the native Glu-plasminogen, or can be obtained by using gene engineering techniques by recombinant peptide synthesis in eukaryotic and bacterial expression systems. Recombinant antigens are corresponding to amino acid sequence of human plasminogen. In particular, the antigen for the invention disclosed in the method of diagnosis, except for full-plasminogen. It may be the following fragments: Lys-plasminogen, the heavy chain (Glu-H), heavy chain (Lys-H), light chain (L), K1-4 (Tyr80-Ala440), K1-3 (Tyr80-Val338), K1-3 (Tyr80-Val354), K1-4 (Asn60-Pro447), K1-4 (Lys78-Pro447), K1-4 (Lys78- Pro446), K1-4 (Lys78-Lys468), K1-4, 5 (Lys78-Arg530), C4-5 (Val355-Phe546), K1 (Tur80-Glu164), K2-3 (Cys165-Val338), K4 (Val354-Ala440), K5 (Ser441-Fhe546), K5 (Val442-Arg561), miniplasminogen, and any combination thereof, (Table 1). 
         [0027]    The inventors discovered and experimentally first confirmed that human plasminogen or its fragments can be used as antigens to determine the titer of autoantibodies in ELISA of human plasma and the result of this reaction can be used to diagnose the presence of cancer. So, as disclosed in the present invention, method of diagnosis is based on a well-defined polypeptides with unique amino acid sequences that any other proteins with identical primary structures and amino acid sequences are identical to those disclosed in the invention. It is clear that an antigens of the present invention can be any polypeptide having partial homology (90% and above) with the claimed polypeptides, since the replacement of individual amino acids does not alter the 3-dimentional structure of Kringle and not an obstacle to the interaction of the antigen-antibody. 
         [0028]    Table 1 describes the various polypeptides—derived from human plasminogen. Kringle fragments form that can be used for preparation an enzyme immunoassay with samples of human blood plasma to detect autoantibodies associated with the growth of tumor. 
         [0000]    
       
         
               
               
               
             
               
               
               
               
             
           
               
                   
               
               
                 peptide chain 
                 Mass, kDa 
                 Name 
               
               
                   
               
             
             
               
                   
               
             
          
           
               
                 Glu 1 -Asn 791   
                 98 
                 Glu- 
                 SEQ ID NO: 1 
               
               
                   
                   
                 plasminogen 
               
               
                 Lys 78 -Asn 791   
                 83, 5-84 
                 Lys-plasminogen 
                 SEQ ID NO: 2 
               
               
                 Glu 1 -Arg 561   
                 65 
                 heavy chain (Glu- 
                 SEQ ID NO: 3 
               
               
                   
                   
                 H) 
               
               
                 Lys 78 -Arg 561   
                 59 
                 heavy chain (Lys- 
                 SEQ ID NO: 4 
               
               
                   
                   
                 H) 
               
               
                 Val 562 -Asn 791   
                 25 (26, 3) 
                 light chain (L) 
                 SEQ ID NO: 5 
               
               
                 Tyr 80 -Ala 440   
                 51-54 
                 K1-4(Tyr 80 - 
                 SEQ ID NO: 6 
               
               
                   
                   
                 Ala 440 ) 
               
               
                 Tyr 80 -Val 338   
                 41 
                 K1-3(Tyr 80 - 
                 SEQ ID NO: 7 
               
               
                   
                   
                 Val 338 ) 
               
               
                 Tyr 80 -Val 354   
                 44 
                 K1-3(Tyr 80 - 
                 SEQ ID NO: 8 
               
               
                   
                   
                 Val 354 ) 
               
               
                 Asn 60 -Pro 447   
                 55 
                 K1-4(Asn 60 - 
                 SEQ ID NO: 9 
               
               
                   
                   
                 Pro 447 ) 
               
               
                 Lys 78 -Pro 447   
                 58 
                 K1-4(Lys 78 - 
                 SEQ ID NO: 10 
               
               
                   
                   
                 Pro 447 ) 
               
               
                 Lys 78 -Pro 446   
                 58 
                 K1-4 (Lys 78 - 
                 SEQ ID NO: 11 
               
               
                   
                   
                 Pro 446 ) 
               
               
                 Lys 78 -Lys 468   
                 61 
                 K1-4(Lys 78 - 
                 SEQ ID NO: 12 
               
               
                   
                   
                 Lys 468 ) 
               
               
                 Lys 78 -Arg 530   
                 66, 60, 57 
                 K1-4,5 (Lys 78 - 
                 SEQ ID NO: 13 
               
               
                   
                   
                 Arg 530 ) 
               
               
                 Val 355 -Phe 546   
                   
                 K4-5(Val 355 - 
                 SEQ ID NO: 14 
               
               
                   
                   
                 Phe 546 ) 
               
               
                 Tyr 80 -Glu 164   
                 9, 81 
                 K1(Tyr 80 -Glu 164 ) 
                 SEQ ID NO: 15 
               
               
                 Glu 1 -Asn 791   
                 98 
                 Glu- 
                 SEQ ID NO: 1 
               
               
                   
                   
                 plasminogen 
               
               
                 Cys 165 -Val 338   
                   
                 K2-3(Cys 165 - 
                 SEQ ID NO: 16 
               
               
                   
                   
                 Val 338 ) 
               
               
                 Val 354 -Ala 440   
                 10-12 
                 K4(Val 354 -Ala 440 ) 
                 SEQ ID NO: 17 
               
               
                 Ser 441 -Fhe 546   
                 12 
                 K5(Ser 441 -Fhe 546 ) 
                 SEQ ID NO: 18 
               
               
                 Val 442 -Arg 561   
                   
                 K5(Val 442 -Arg 561 ) 
                 SEQ ID NO: 19 
               
               
                 Val 442 -Asn 791   
                 (40) 38 
                 miniplasminogen 
                 SEQ ID NO: 20 
               
               
                   
               
             
          
         
       
     
         [0029]      FIG. 1  shows the primary structure of Human plasminogen. 
         [0030]    Filled arrows identify the cleavage sites for: (a) the release of the signal peptide between residues—1 and 1, which is required for the generation of the mature form of the protein; (b) the release of the activation peptide (Glu′-Lys 77 ) resulting in the conversion of Glu′-Pg to Lys 78 -Pg or Glu′-Pm to Lys 78 -Pm; (c) the activation of Human plasminogen to plasmin (CS) at the Arg 561 -Val 562  peptide bond. Unfilled arrows identify introns in the gene sequence. Triangles locate the N-linked oligosaccharide site at sequence position 289 and the O-linked glycan at position 346. The catalytic triad, His 603 , Asp 646 , and Ser 741 , is also indicated (*). Disulfide bonds are depicted by heavy bars. ♦ phosphorylation site alanine—ala—A; arginine—arg—R; asparagine—asn—N; aspartic acid—asp—D; cysteine—cys—C; glutamine—gln—Q; glutamic acid—glu—E; glycine—gly—G; histidine—his—H; isoleucine—ile—I; leucine—leu—L; lysine—lys—K; methionine—met—M; phenylalanine—phe—F; proline—pro—P; serine—ser—S; threonine—thr—T; tryptophan—trp—W; tyrosine—tyr—Y; valine—val—V. 
         [0031]      FIG. 2  shows the scheme of transformation of Glu-plasminogen to the heavy (K1-5+30 r) and light chain (micro-plasmin). 
       BRIEF SUMMARY OF THE INVENTION 
       [0032]    Human plasminogen and its fragments disclosed herein (Table 1) were purified from blood plasma and used as antigens to create enzyme immunoassay for the determination of levels of autoantibodies class IgG, IgA, IgM to plasminogen/or its fragments in the blood of patients with various cancers, confirmed by alternative methods of diagnostics. According to the fact that the growth of all solid tumors is accompanied by increased growth of vessels and capillars in the area of the tumor the concentration of plasminogen and its products of degradation are used to increase in this area, that leads to the synthesis of autoantibodies to these products. Thus, the undoubted advantage of the present invention disclose the common marker of early diagnostic of cancer which is to the appearance of autoantibodies to plasminogen and its products of degradation in humans associated with the development of a tumor any kind. 
         [0033]    Isolation of Antigens for ELISA. 
         [0034]    The method for preparing of heavy chain (Glu-H) Glu1-Arg561 and light chain (L) Val562-Asn791 of human plasminogen. 
         [0035]    The base of the method consists in the activation of plasminogen to plasmin, followed by reduction of S-S bonds between heavy and light chains in conditions that exclude autolysis, and using affinity chromatography on Lys-Sepharose 4B for the following separation. Urokinase cleave Arg561-Val562 bond in plasminogen. The resulting plasmin cut 77-78 bond and cleaved N-terminal peptide (1-77) is out. Mercaptoethanol reduct two bonds between Cys558-Cys566 and Cys548-Cys666, linking the heavy and light chains. 
       First Step: 
       [0036]    Glu-Plasminogen was isolated from frozen donor human plasma by affinity chromatography on Lys-Sepharose 4B at 4° C., pH 8.0. Blood plasma was thawed in the presence of aprotinin, centrifuged 30 min at 4° C. and diluted 2-fold to 0.02 M phosphate buffer, pH 8.0, containing 20 KIU/ml aprotinin. Prepared plasma applied into a column with Lys-Sepharose 4B, equilibrated with 0.1 M K-phosphate buffer, pH 8.0, containing 20 KIU/ml aprotinin. The column was washed from unbound protein to 0.3 M phosphate buffer, pH 8.0, containing 20 KIU/ml aprotinin, overnight to absorbance at A280=0.05-0.01. Glu-Plasminogen was eluted with a solution of 0.2 M 6-aminocaproic acid, 0.1 M K-phosphate buffer, pH 8.0, containing 20 KIU/ml aprotinin. Fractions containing protein were pooled and subjected to further purification by precipitation (NH4) 2SO4 (0.31 g/ml protein solution). The precipitate was allowed to stand at 4° C. for 18-24 hours and then separated by centrifugation and dissolved in 0.05 M Tris-HCl buffer, pH 8.0 to a concentration of about 1.5-2.0 mg/ml. Purified Glu-Plasminogen dialyzed at 4° C. against water (pH 8.0) and lyophilized. 
       Second Step: 
       [0037]    To a solution of Glu-plasminogen (5 mg/ml) in 0.05 M Tris-HCl buffer, pH 8 8, containing 0.02 M L-lysine, 0.15 M NaCl, 20% glycerol, and 6000 KIU/ml aprotinin urokinase was added to a final concentration of 600 IU/ml and incubated for 4 h at 37° C. The complete of conversion of Glu-plasminogen to plasmin was monitored by an increase to a maximum rate of hydrolysis of plasmin specific substrate S-2251 (HD-Val-Leu-Lys p-nitroanilide, “Sigma”, USA) in samples taken from the reaction mixture. 
         [0000]    Third stage 
         [0038]    The reduction of S-S-bonds between heavy and light chains of plasmin. 
         [0039]    Mercaptoethanol was added to the solution of plasmin to a final concentration of 0.25 mM and incubated under nitrogen in the dark for 20 minutes at room temperature. In the result, free SH-groups were blocked by adding freshly prepared solution iodoacetic acid in 0.1 M Na-phosphate buffer, pH 8.0 (final concentration of 0.315 M) and incubated for 20 min. 
         [0040]    Fourth Stage: 
         [0041]    The separation of heavy and light chains of plasmin by column chromatography on Lys-Sepharose 4B. 
         [0042]    The reaction mixture was diluted to a concentration of 1 mg/ml of protein by 0.1 M Na-phosphate buffer, pH 8.0, containing 20 KIU/ml aprotinin and applied to Lys-Sepharose 4B column equilibrated with the same buffer. Chromatography was performed at 25° C. The heavy chain of plasmin has kringles 1-5 and 30 amino acid residues of the connecting peptide. It was adsorbed on the sorbent, but light chain is eluted with equilibration buffer. Heavy chain (MR˜56-57 kDa) was eluted by 0.2 M solution of 6-aminocaproic acid in 0.1 M Na-phosphate buffer, pH 8.0. The pooled fractions were dialyzed against water (pH˜8.0) and lyophilized. 
         [0043]    The purity and molecular weight of the protein was investigated by 12% SDS-polyacrylamide gel electrophoresis. 
         [0044]    Furthermore, the absence of amidase activity (for S-2251) before and after incubation with streptokinase its solution indicated that the heavy chain does not contain trace concentrations of miniplasminogen, which may not show up during electrophoresis. 
         [0045]    The purification of Lys-plasminogen (Lys78-Asn791) and heavy chain (Lys-H Lys78-Arg561) was performed by the same method, but without inhibitor -aprotinin. 
         [0046]    Isolation of miniplasminogen (Val442-Asn791). Miniplasminogen consist of K5 and light chain. Its sequence starts from Val442 to Asn791. Miniplasminogen obtained by incubation of Lys-plasminogen (Lys78-Asn791) with elastase followed by gel filtration on G-75 Sephadex. 
         [0047]    Isolation of Kringle K1-4, 5 (Lys78-Arg530) was performed according to the method described in Cao R., Wu H. L., Veitonmaki N., Linden P., Farnedo J., Shi C.Y., and Cao Y, (1999) Proc. Natl. Acad. Sci. USA, 96, 5728-5733, with some modifications. 
         [0048]    Glu-plasminogen (10 mg/ml) was activated with urokinase (600 ME/ml) in 0.05 M phosphate buffer, pH 9.0, containing 0.02 M L-lysine and 0.1 M NaCl, at 37° C. Complete conversion of plasminogen to plasmin was monitored by the increase in the amidase activity of the solution to the maximum value. An equal volume of 0.2 M glycerol buffer, pH 12.0 was added to a solution of plasmin and incubated for 18 h at 25 C, pH of 10.5. The reaction mixture was diluted 5-fold with buffer containing 0.1 M phosphate buffer, pH 8.0, and 40 KIU/ml aprotinin, and applied to a column of Lys-Sepharose 4B equilibrated with the same buffer. After the following out of microplasminogen, adsorbed K1-4,5 was eluted from the column with 0.2 M solution of 6-aminocaproic acid in 0.1 M phosphate buffer, pH 8.0 and 40 KIU/ml aprotinin, dialyzed against water and lyophilized. The purity of the substance was checked by 12% SDS-polyacrylamide gel electrophoresis. 
         [0049]    Isolation of kringle K1-4 (Tyr80-Ala440) and K1-3 (Tyr80-Val338) K4-5 (Val355-Phe546) was performed using elastase treatment of Glu-plasminogen by the method described in the works of Cao Y., Ji R. W., Davidson D., Schaller J., Marti D., Sohndel S., McCanse S. G., O&#39;Reilly M. S., Llinas M., and Folkman J. (1996) J. Biol. Chem., 271, 29461-29467. Glu-plasminogen was incubated with elastase at a ratio of 50:1 (M/M) in a buffer containing 0.05 M Tris-HCl, pH 8.5, 0.5 M NaCl, and 200 KIU aprotinin, for 5 hours at room temperature. The reaction was stopped by adding PMFS to maintain its concentration 1 mM for 40-50 min. Then gel-filtration on a column of Sephadex G-75 was performed to separate low and high molecular proteins. Protein fractions of the second peak containing K1-3, K1-4, K4-5 and miniplasminogen was applied to an affinity column with Lys-Sepharose 4B equilibrated with buffer containing 0.05 M Tris-HCl, pH 8.5, 0.15 M NaCl. After the flowing throw miniplasminogen which is not adsorbed on the Lys-Sepharose 4B, adsorbed fragments K1-3, K1-4 and K4-5 was eluted with a solution of 0.2 M 6-aminocaproic acid in the same buffer, dialyzed against a buffer containing 0.02 M Tris-HCl, pH 8.0, and applied to a column of heparin-agarose equilibrated with the same buffer. After elution of unbound fragment K1-4 and K4-5 with the buffer, the fragment K1-3 was eluted with a solution of 0.25 M KCl in the same buffer. The purified fragment K1 -3 was dialyzed against water and lyophilized. K1-4 and K4-5 were separated by gel filtration on Sephadex G-75. 
         [0050]    Kringles K5 (Ser449 (Pro452)-Fhe546), K1-3 (Tyr80-Val338), K-4 (Val335-Ala440) were prepared according to the work of Cao, Y., Chen, A., An, S. S. A., Ji, R. W., Davidson, D., and Llinas, M. (1997) J. Biol. Chem. 272, 22924-22928). The method is to digest by elastase Lys-plasminogen (Lys78-Asn791). After processing elastase mixture was applied to a column of Mono-S (Bio-Rad) equilibrated with buffer containing 20 mM NaOAc, pH 5.0. Fragments of plasminogen were eluted by gradient 1 M KCl in buffer containing 20 mM NaOAc, pH 5.0. We used a gradient of 0-20%, 20-50%, 50-70% and 70-100%. K-5 release to 50%. For this scheme, but in a another gradient K-4 (Val335-Ala440) and kringle K1-3 (Tyr80-Val354) were obtained. 
         [0051]    The method of isolation of K5 (Val442-Arg561) is to digesting by elastase of miniplazminogen (Val442-Asn791) containing 5-kringle of heavy chain following by digesting of the fragment by pepsin and then using the gel filtration and ion exchange chromatography according to the work (Theresa Thewes, Vasudevan Ramesh, Elcna L. Simplaceanu and Miguel Llinfis, Isolation, purification and I H-NMR characterization of a kringle 5 domain fragment from human plasminogen (Biochimica et Biophysica Acta 912 (1987), 254-269). 
         [0052]    Kringle K1-4 (Lys78-Pro446) and K1-4 (Lys78-Lys468) was prepared according to the method with metalloproteinases (Patterson, B. C. and Sang, Q. A. (1997) J. Biol. Chem. 272, 28823-28825). 
         [0053]    Kringle K1-4 (Asn60-Pro447) obtained by the method with metalloproteinases (Lijnen, H. R., Ugwu, F., Bini, A., and Collen, D. (1998) Biochemistry 37, 4699-4702). 
         [0054]    Kringle K1 (Tur80-Glu164) and K2-3 (Cys165-Val338) were isolated from the K1-3 (Tyr80-Val338) by treating of pepsin (or protease s.aureus V8) with a further separation on lys-Sepharose and gel filtration on Sephadex G-75. 
       Preparation of Diagnostic Test Kits for ELISA of Autoantibodies 
       [0055]    Glu-plasminogen or its fragments having at least one kringle were used as Antigens for ELISA of autoantibodies. Different types of antigens used in ELISA are listed in Table 1. Their primary amino acid sequence shown in the sequence listing. 
         [0056]    The antigen was diluted in 0.1 M carbonate-bicarbonate buffer pH 9.6 in the maximum concentration of 5 μg/ml for molecules with a molecular weight of more than 25 kDa and 10 μg/ml for molecules with a molecular weight less than 25 kDa. These dilutions of antigen were used to identify all types of immunoglobulins. 
         [0057]    PBS (phosphate buffered saline, phosphate salt solution): 
         [0058]    0.14 M NaCl; 0.003 M KCl, 0.005 M Na2HPO4, 0.002 M KH2PO4 
         [0059]    Preparation 1 L 10× PBS: 
         [0060]    80 g-NaCl 2 g-KCl 18 g-Na2HPO4 2 g-KH2PO4 
         [0061]    Substrate buffer solution (pH 4.3): 31 mM citric acid, 0.05 N NaOH, 3 mM H2O2 
         [0062]    TMB solution: 5 mM 3,3′, 5,5′-tetramethylbenzidine in 70% DMSO 
         [0063]    Chrornogenic substrate solution (prepared before use): Mixed 4 parts of the substrate buffer solution and one part of the TMB solution. 
         [0064]    To prepare a kit for ELISA was performed pre-immobilized antigen. To immobilize the antigen can be used different types of material such as nitrocellulose, glass beads or other particles that can absorb proteins, immunological plastic strips or plates. We used for experiments immunological plastic strips (Nunc). To each well was diluted into 100 μL antigen solution. Incubation was carried out for 14-16 hours at 37° C. in a humidified chamber. The antigen solution was removed by shaking out, and then wells washed twice by a solution containing a PBS with 0.05% Tweeen-20, 200 μL/well to remove unbound antigen. For the block was used solution of 1% gelatin in PBS, 200 μL/well, with incubation for 1.5-2 hours at room temperature. After incubation, the blocking fluid was removed, the plate was dried overnight at room temperature on air. 
         [0065]    Control probes and tested samples were diluted in 50-fold by diluent (0.5% gelatin, 0.001 M EDTA in PBS), then were poured 100 μL into appropriate wells and incubated for 1 h at 37° C. After incubation, the solution were removed, the plate was washed four times with washing buffer (PBS with 0.05% Tween-20), each time carefully removing the contents of the wells. Working dilution of conjugate (to determine IgG, IgA, IgM as conjugate used respectively Mab Fc IgG-peroxidase, Mab Fc IgA-peroxidase, Mab IgM-peroxidase) were added into the appropriate wells of 100 mcl/well and then incubated for 1 h at 37° C. Unbound components were removed 4 times by washing of wash solution. Then in all the wells were added 100 μL of substrate-chromogen solution, and incubated for 15 minutes at 370 C. The reaction was stopped by adding to wells 100 μL of stop solution (2M H2SO4). Photometry was performed on vertical scanning photometer “UNIPLAN” (company “Picon”, Russia) with a wavelength of 450 nm. 
         [0000]    The Diagnostic of Cancer by Enzyme Immunoassay Detection of Autoantibodies to Human Plasminogen and/or its Fragments 
         [0066]    Blood samples were taken from patients cubital vein using vacutaners with EDTA. The samples were then centrifuged at 3000 r/min for 15 min. Plasma was poured into tubes in 100 μL., Frozen and stored at −40 C. 
         [0067]    The control group has plasma samples taken from 30 healthy men and 30 healthy women. Each sample was negative in tests for hepatitis A, B and C virus, HIV, tuberculosis and syphilis. 
         [0068]    The level of autoantibodies IgG and IgA in the control samples was measured using the ELISA kit, according to the described method. Dilution of control plasma samples was chosen so that the optical density was less than 0.2. 
         [0069]    The dilution of samples in ELISA was established of 1/50 for each tested samples, which was subsequently used for the analysis of all samples. The antigen was used as a whole molecule of glu-plasminogen, as its fragments. For accuracy, the determination of each sample was tested in duplicate. After measuring 30 male and 30 female control samples was calculated the average optical density for each of the control group used for testing with various fragments of plasminogen or full plasminogen molecule as a ligand. 
         [0070]    ELISA test samples was performed with each individual antigen. Number of samples above the average in the male control group was within 2% to 5%, while in the women&#39;s within 3% to 6% when tested with all antigens investigated. For a comparative study of the control group to samples of cancer and other diseases for the control group were taken 5 samples with indicators of optical density, not more than 5% of the average. These five samples were mixed into one pooled sample—control sample (K), used as a reference level of autoantibodies to plasminogen or its fragments. The samples of control were different to the study of antibodies IgG, IgA and IgM. 
         [0071]    To assess the effectiveness of various fragments of plasminogen for the early diagnostic of cancer was used the samples of plasma of patients with various forms of cancer (Table 2). 
         [0000]    
       
         
               
             
               
               
               
               
               
             
               
             
               
               
               
               
             
               
               
               
               
             
               
             
               
               
               
               
               
             
               
             
               
               
               
               
               
             
               
             
               
               
               
               
             
           
               
                 TABLE 2 
               
               
                   
               
             
             
               
                 Prostate Cancer 
               
             
          
           
               
                 No 
                 Age 
                 Diagnosis 
                 Biopsy 
               
               
                   
               
               
                 1 
                 68 
                 T2cNoMo 
                 small acinar adenocarcinoma amid mixed 
                 PSA - 35 ng/ml 
               
               
                   
                   
                   
                 hyperplasia and chronic prostatitis. Gleason 
               
               
                   
                   
                   
                 score 2 (1 + 1) 
               
               
                 2 
                 58 
                 T2cNxMo 
                 acinar adenocarcinoma. Gleason score 6 (4 + 
                 PSA -9 ng/ml 
               
               
                   
                   
                   
                 2) 
               
               
                 3 
                 69 
                 T3NxMo 
                 adenocarcinoma 
               
               
                 4 
                 67 
                 T2aNoMo 
                 Focuses of small acinar adenocarcinoma, 
                 PSA 7 ng/ml 
               
               
                   
                   
                   
                 Gleason score 4 points (2 + 2), mixed 
               
               
                   
                   
                   
                 hyperplasia, chronic prostatitis 
               
               
                 5 
                 64 
                 T2NxMo 
                 small acinar adenocarcinoma, Gleason 
                 PSA - 26 ng/ml 
               
               
                   
                   
                   
                 score 5 (3 + 2) 
               
               
                 6 
                 63 
                 T2cNoMo 
                 small acinar adenocarcinoma, Gleason 
                             CA-12 ng/ml 
               
               
                   
                   
                   
                 score 7 (3 + 4) 
               
               
                 7 
                 62 
                 ToNoMo 
                 small acinar adenocarcinoma, Gleason 
                 PSA - 35 ng/ml 
               
               
                   
                   
                   
                 score 3 (1 + 2) 
               
               
                 8 
                 78 
                 T2NxMo 
                 small acinar adenocarcinoma, Gleason 
                 PSA - 25 ng/ml 
               
               
                   
                   
                   
                 score 5 (3 + 2) 
               
               
                 9 
                 72 
                 T2NxMo 
                 small acinar adenocarcinoma, Gleason 
                 PSA - 50 ng/ml 
               
               
                   
                   
                   
                 score 5 (3 + 2) 
               
               
                 10 
                 62 
                 T2cNoMo 
                 small acinar adenocarcinorna, Gleason 
                             CA-10 ng/ml 
               
               
                   
                   
                   
                 score 5 (2 + 3) 
               
               
                 11 
                 78 
                 T3NxMx 
                 adenocarcinoma 
                 PSA - 20 ng/ml 
               
               
                 12 
                 73 
                 T2NxMo 
                 small acinar adenocarcinoma, Gleason 
                 PSA - 55 ng/ml 
               
               
                   
                   
                   
                 score 7 (4 + 3) 
               
               
                 13 
                 75 
                 T2aN0Mo 
                 small acinar adenocarcinoma, Gleason 
                 PSA - 11 ng/ml 
               
               
                   
                   
                   
                 score 4 points (2 + 2) 
               
               
                 14 
                 73 
                 T3aNxMx 
                 small acinar adenocarcinoma, with 
                 PSA - 44 ng/ml 
               
               
                   
                   
                   
                 hyperplasia and chronic prostatitis 
               
               
                 15 
                 60 
                 T2N0M0 
                 small acinar adenocarcinoma, Gleason 
                 PSA - 67 ng/ml 
               
               
                   
                   
                   
                 score 7 (4 + 3) 
               
               
                 16 
                 66 
                 T2NxMx 
                 Large acinar adenocarcinoma, Gleason 
                 PSA - 57 ng/ml 
               
               
                   
                   
                   
                 score 5 (3 + 2) 
               
               
                 17 
                 71 
                 T2bN0M1 
                 acinar adenocarcinoma, Gleason score 7 (3 + 
                 PSA - 70 ng/ml 
               
               
                   
                   
                   
                 4) 
               
               
                 18 
                 70 
                 T3N1M0 
                 small acinar adenocarcinoma, Gleason 
                 PSA - 120 ng/ml 
               
               
                   
                   
                   
                 score 9 (4 + 5) 
               
               
                 19 
                 73 
                 T3N1M0 
                 small acinar adenocarcinoma, Gleason 
                 PSA - 30 ng/ml 
               
               
                   
                   
                   
                 score 3 (3 + 3) 
               
               
                 20 
                 78 
                 T3N1M0 
                 acinar adenocarcinoma, Gleason score 7 (3 + 
                 PSA - 70 ng/ml 
               
               
                   
                   
                   
                 4) 
               
               
                 21 
                 81 
                 T2N0M0 
                 acinar adenocarcinoma, Gleason score 7 (3 + 
                 PSA - 15 ng/ml 
               
               
                   
                   
                   
                 4) 
               
               
                 22 
                 78 
                 T2N0M0 
                 small acinar adenocarcinoma, Gleason 
                 PSA - 70 ng/ml 
               
               
                   
                   
                   
                 score 7 (4 + 3) 
               
               
                 23 
                 46 
                 T2N0M0 
                 small acinar adenocarcinoma, Gleason 
                 PSA - 35 ng/ml 
               
               
                   
                   
                   
                 score 6 (3 + 3) 
               
               
                 24 
                 71 
                 T2cN0M0 
                 Adenocarcinoma Gleason score 5 (2 + 3) 
                 PSA - 18 ng/ml 
               
               
                 25 
                 59 
                 T3NxM1 
                 Adenocarcinoma, Gleason score 8 (4 + 4) 
                 PSA - 225 ng/ml 
               
               
                 26 
                 64 
                 T2N0M0 
                 small acinar adenocarcinoma, Gleason 
                 PSA - 16 ng/ml 
               
               
                   
                   
                   
                 score 5 (3 + 2) 
               
               
                 27 
                 73 
                 T3aNxMx 
                 small acinar adenocarcinoma, with 
                 PSA - 44 ng/ml 
               
               
                   
                   
                   
                 hyperplasia and chronic prostatitis, Gleason 
               
               
                   
                   
                   
                 score 5 (3 + 2) 
               
               
                 28 
                 65 
                 T3NxM1 
                 small acinar adenocarcinoma, Gleason 
                 PSA - 163 ng/ml 
               
               
                   
                   
                   
                 score 9 (4 + 5) 
               
               
                 29 
                 62 
                 T2aNoMo 
                 Focuses small acinar adenocarcinoma 
                 PSA - 70 ng/ml 
               
               
                 30 
                 64 
                 T2aNxMx 
                 Focuses small acinar adenocarcinoma, 
                 PSA - 14 ng/ml 
               
               
                   
                   
                   
                 hyperplasia and chronic prostatitis 
               
               
                   
               
             
          
           
               
                 Lung Cancer 
               
             
          
           
               
                 No 
                 Age, sex 
                 Diagnosis 
                 Biopsy 
               
               
                   
               
               
                 1 
                 77 f 
                 Cancer of the upper lobe of the 
                 moderately differentiated 
               
               
                   
                   
                 left lung, T2N0M0, stage IB 
                 adenocarcinoma 
               
               
                 2 
                 71 f 
                 Cancer of the upper lobe of the 
                 moderately differentiated 
               
               
                   
                   
                 left lung, T2N0M0, stage IIB 
                 squamous cell carcinoma 
               
               
                 3 
                 80 M 
                 Cancer of the upper lobe of the 
                 moderately differentiated 
               
               
                   
                   
                 left lung, T2N1M0, stage IIB 
                 squamous cell carcinoma 
               
               
                 4 
                 65 M 
                 Cancer of the right upper lobe, 
                 moderately differentiated 
               
               
                   
                   
                 T2N0M0, stage IB 
                 squamous cell carcinoma 
               
               
                 5 
                 63 M 
                 Cancer of the upper lobe of the 
                 moderately differentiated 
               
               
                   
                   
                 left lung, T2N1M0, stage IIB 
                 glandular squamous cell 
               
               
                   
                   
                   
                 carcinoma 
               
               
                 6 
                 70 M 
                 Cancer of the lower lobe of the 
                 small-cell lung carcinoma 
               
               
                   
                   
                 right lung, T2N0M0, stage IB 
               
               
                 7 
                 71 M 
                 Cancer of the right upper lobe 
                 moderately differentiated 
               
               
                   
                   
                 T2N0M0, stage IB 
                 glandular squamous cell 
               
               
                   
                   
                   
                 carcinoma 
               
               
                 8 
                 71 f 
                 Cancer of the upper lobe of the 
                 small-cell lung carcinoma 
               
               
                   
                   
                 left lung, T4N2M0, stage IIIB 
               
               
                 9 
                 66 M 
                 Cancer of the lower lobe of the 
                 small-cell lung carcinoma 
               
               
                   
                   
                 right lung, T2N1M1, stage IV, 
               
               
                   
                   
                 with liver metastases 
               
               
                 10 
                 52 M 
                 Cancer of the right upper lobe 
                 small-cell lung carcinoma 
               
               
                   
                   
                 T2N2M0, stage IIIA 
               
               
                 11 
                 58 f 
                 Cancer of the lower lobe of the 
                 moderately differentiated 
               
               
                   
                   
                 right lung, T1N0M0, stage IA 
                 squamous cell carcinoma 
               
               
                 12 
                 57 f 
                 Cancer of the right upper lobe 
                 small-cell lung carcinoma 
               
               
                   
                   
                 T2N0M0, stage IB 
               
               
                 13 
                 55 M 
                 Cancer of the lower lobe of the 
                 small-cell lung carcinoma 
               
               
                   
                   
                 right lung, T2N0M0, stage IB 
               
               
                 14 
                 68 f 
                 Cancer of the upper lobe of the 
                 moderately differentiated 
               
               
                   
                   
                 left lung, T4N2M0, stage IIIB 
                 squamous cell carcinoma 
               
               
                 15 
                 65 f 
                 Cancer of the right upper lobe, 
                 Moderately differentiated 
               
               
                   
                   
                 T2N0M0, stage IB 
                 squamous cell carcinoma 
               
               
                 16 
                 65 M 
                 Cancer of the lower lobe of the 
                 small-cell lung carcinoma 
               
               
                   
                   
                 right lung, T2N0M0, stage IB 
               
               
                 17 
                 47 f 
                 Cancer of the lower lobe of the 
                 moderately differentiated 
               
               
                   
                   
                 right lung, T2N0M0, stage IA 
                 adenocarcinoma 
               
               
                 18 
                 31 M 
                 Cancer of the lower lobe of the 
                 small-cell lung carcinoma 
               
               
                   
                   
                 right lung, T2N1M1, stage 
               
               
                   
                   
                 Cancer of the upper lobe of the 
               
               
                   
                   
                 left lung, T4N2M0, stage IVB 
               
               
                   
                   
                 with metastases 
               
               
                 19 
                 54 f 
                 Cancer of the upper lobe of the 
                 small-cell lung carcinoma 
               
               
                   
                   
                 left lung, T4N2M0, stage IIIB 
               
               
                 20 
                 54 f 
                 Cancer of the right upper lobe 
                 small-cell lung carcinoma 
               
               
                   
                   
                 T2N2M0, stage IIIA 
               
               
                   
               
             
          
           
               
                 No 
                 Age 
                 Diagnosis 
                 Biopsy 
               
               
                   
               
             
          
           
               
                 Breast Cancer 
               
             
          
           
               
                 1 
                 63 
                 Cancer of the right breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 120 units/mL 
               
               
                   
                   
                 T2N0M0, stage IIB 
                 grade II (6 points) 
               
               
                 2 
                 47 
                 Cancer of the left breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 160 units/mL 
               
               
                   
                   
                 T2N1M0, stage IIA 
                 grade II (6 points)) 
               
               
                 3 
                 52 
                 Cancer of the right breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 140 units/mL 
               
               
                   
                   
                 T2N0M0, stage IIB 
                 grade II (6 points) 
               
               
                 4 
                 65 
                 Cancer of the left breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 110 units/mL 
               
               
                   
                   
                 T4N1M0, stage IIIB 
                 grade II (6 points) 
               
               
                 5 
                 59 
                 Cancer of the left breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 140 units/mL 
               
               
                   
                   
                 T2N1M0, stage IIB 
                 grade II (6 points) 
               
               
                 6 
                 72 
                 Cancer of the left breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 160 units/mL 
               
               
                   
                   
                 T2N2M0, stage IIIA 
                 grade III (8 points) 
               
               
                 7 
                 54 
                 Cancer of the left breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 90 units/mL 
               
               
                   
                   
                 T1N0M0, stage I 
                 grade I (5 points) 
               
               
                 8 
                 73 
                 Cancer of the right breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 100 units/mL 
               
               
                   
                   
                 T2N0M0, stage IIB 
                 grade II (7 points) 
               
               
                 9 
                 48 
                 Cancer of the right breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 170 units/mL 
               
               
                   
                   
                 T4N0M0, stage IIIB 
                 grade II (7 points) 
               
               
                 10 
                 42 
                 Cancer of the left breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 60 units/mL 
               
               
                   
                   
                 T4N1M0, stage IIIB 
                 grade II (7 points) 
               
               
                 11 
                 61 
                 Cancer of the right breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 130 units/mL 
               
               
                   
                   
                 T2N0M0, stage IIB 
                 grade II (6 points) 
               
               
                 12 
                 46 
                 Cancer of the left breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 150 units/mL 
               
               
                   
                   
                 T3N1M0, stage IIIA 
                 grade II (7 points) 
               
               
                 13 
                 47 
                 Cancer of the left breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 100 units/mL 
               
               
                   
                   
                 T2N0M0, stage IIA 
                 grade I (5 points) 
               
               
                 14 
                 60 
                 Cancer of the right breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 180 units/mL 
               
               
                   
                   
                 T2N0M0,            IIB 
                 grade II (7 points) 
               
               
                 15 
                 62 
                 Cancer of the right breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 90 units/mL 
               
               
                   
                   
                 T2N0M0, stage IIB 
                 grade II (6 points) 
               
               
                 16 
                 47 
                 Cancer of the left breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 140 units/mL 
               
               
                   
                   
                 T4N1M0, stage IIIB 
                 grade II (6 points) 
               
               
                 17 
                 28 
                 Cancer of the left breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 100 units/mL 
               
               
                   
                   
                 T4N1M0, stage IIIB 
                 grade II (6 points) 
               
               
                 18 
                 61 
                 Cancer of the left breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 100 units/mL 
               
               
                   
                   
                 T4N1M0, stage IIIB 
                 grade II (7 points) 
               
               
                 19 
                 69 
                 Cancer of the right breast, 
                 Infiltrative ductal Carcinoma, 
                 CA 15-3 110 units/mL 
               
               
                   
                   
                 T4N1M0, stage IIIB 
                 grade III (8 points) 
               
               
                 20 
                 50 
                 Cancer of the left breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 110 units/mL 
               
               
                   
                   
                 T2N1M0, stage IIB 
                 grade II (6 points) 
               
               
                 21 
                 48 
                 Cancer of the right breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 90 units/mL 
               
               
                   
                   
                 T2N1M0, stage IIB 
                 grade II (7 points) 
               
               
                 22 
                 80 
                 Cancer of the left breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 70 units/mL 
               
               
                   
                   
                 T1N1M0, stage IIA 
                 grade II (6 points) 
               
               
                 23 
                 60 
                 Cancer of the left breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 140 units/mL 
               
               
                   
                   
                 T2N1M0, stage IIB 
                 grade II (7 points) 
               
               
                 24 
                 62 
                 Cancer of the right breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 100 units/mL 
               
               
                   
                   
                 T2N1M0, stage IIB 
                 grade II (6 points) 
               
               
                 25 
                 54 
                 Cancer of the right breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 90 units/mL 
               
               
                   
                   
                 T4N1M0, stage IIIB 
                 grade II (7 points) 
               
               
                 26 
                 75 
                 Cancer of the left breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 120 units/mL 
               
               
                   
                   
                 T2N2M0, stage IIIA 
                 grade III (8 points) 
               
               
                 27 
                 67 
                 Cancer of the right breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 150 units/mL 
               
               
                   
                   
                 T4N1M0, stage IIIB 
                 grade II (7 points) 
               
               
                 28 
                 67 
                 Cancer of the left breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 110 units/mL 
               
               
                   
                   
                 T2N1M0, stage IIB 
                 grade II (6 points) 
               
               
                 29 
                 61 
                 Cancer of the left breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 180 units/mL 
               
               
                   
                   
                 T2N1M0, stage IIB 
                 grade II (6 points) 
               
               
                 30 
                 39 
                 Cancer of the right breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 160 units/mL 
               
               
                   
                   
                 T2N2M0, stage IIIA 
                 grade II (7 points) 
               
               
                 31 
                 57 
                 Cancer of the right breast, 
                 Infiltrative ductal carcilnoma, 
                 CA 15-3 120 units/mL 
               
               
                   
                   
                 T2N1M0, stage IIB 
                 grade III (8 points) 
               
               
                 32 
                 70 
                 Cancer of the left breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 200 units/mL 
               
               
                   
                   
                 T2N1M0, stage IIB 
                 grade II (6 points) 
               
               
                 33 
                 69 
                 Cancer of the left breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 180 units/mL 
               
               
                   
                   
                 T4N2M0, stage IIIB 
                 grade III (8 points) 
               
               
                 34 
                 49 
                 Cancer of the right breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 220 units/mL 
               
               
                   
                   
                 T2N2M0, stage IIIA 
                 grade III (8 points) 
               
               
                 35 
                 61 
                 Cancer of the left breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 140 units/mL 
               
               
                   
                   
                 T2N2M1, stage IV 
                 grade III (8 points) 
               
               
                 36 
                 43 
                 Cancer of the left breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 60 units/mL 
               
               
                   
                   
                 T4N1M0, stage IIIB 
                 grade II (7 points) 
               
               
                 37 
                 50 
                 Cancer of the right breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 200 units/mL 
               
               
                   
                   
                 T2N1M0, stage IIB 
                 grade II (7 points) 
               
               
                 38 
                 85 
                 Cancer of the left breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 250 units/mL 
               
               
                   
                   
                 T2N1M0, stage IIB 
                 grade II (7 points) 
               
               
                 39 
                 47 
                 Cancer of the left breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 150 units/mL 
               
               
                   
                   
                 T4N1M0, stage IIIB 
                 grade II (6 points) 
               
               
                 40 
                 55 
                 Cancer of the left breast, 
                 Infiltrative ductal carcinoma, 
                 CA 15-3 110 units/mL 
               
               
                   
                   
                 T4N2M0, stage IIIB 
                 grade II (6 points) 
               
             
          
           
               
                 Ovarian cancer 
               
             
          
           
               
                 1 
                 71 
                 T3cNoMo, stage IIIC 
                 Serous cystadenocarcinoma 
                 CA125-428 units/mL 
               
               
                 2 
                 62 
                 T3N1M0, stage IIIC 
                 Serous cystadenocarcinoma 
                 CA125-510 units/mL 
               
               
                 3 
                 62 
                 T3N0M0, stage IIB 
                 Serous cystadenocarcinoma 
                 CA125-120 units/mL 
               
               
                 4 
                 56 
                 T2N0M0, stage IIB 
                 Endometrioid 
                 CA125-350 units/mL 
               
               
                   
                   
                   
                 cystadenocarcinoma 
               
               
                 5 
                 64 
                 T1N0M0, stage IA 
                 serous-mucinous 
                 CA125-240 units/mL 
               
               
                   
                   
                   
                 cystadenocarcinoma 
               
               
                 6 
                 67 
                 T2N0M0, stage IIB 
                 Serous cystadenocarcinoma 
                 CA125-370 units/mL 
               
               
                 7 
                 53 
                 T3N1M0, stage IIIC 
                 Serous cystadenocarcinoma 
                 CA125-600 units/mL 
               
               
                 8 
                 46 
                 T2N0M0, stage IIB 
                 granulosa cell cancer 
                 CA125-350 units/mL 
               
               
                 9 
                 30 
                 T2N0M0, stage IIB 
                 Endometrioid 
                 CA125-540 units/mL 
               
               
                   
                   
                   
                 cystadenocarcinoma 
               
               
                 10 
                 63 
                 T1N0M0, stage IA 
                 Serous cystadenocarcinoma 
                 CA125-120 units/mL 
               
               
                 11 
                 50 
                 T2N1M0, stage IIIC 
                 Serous cystadenocarcinoma 
                 CA125-200 units/mL 
               
               
                   
               
             
          
           
               
                 Melanoma 
               
             
          
           
               
                 No 
                 Age 
                 Diagnosis 
                 Biopsy 
               
               
                   
               
               
                 1 
                 71 
                 Melanoma of the skin right foot, 
                 Epithelioid melanoma, Clark&#39;s level 5. 
               
               
                   
                   
                 T4N0M0, stage III 
                 Breslow&#39;s depth 12 MM 
               
               
                 2 
                 67 
                 Melanoma of abdominal skin, 
                 Epithelioid melanoma, Clark&#39;s level 5. 
               
               
                   
                   
                 T4N2M0, stage III 
                 Breslow&#39;s depth 20 MM 
               
               
                 3 
                 68 
                 Melanoma of the skin of the right 
                 Epithelioid melanoma, Clark&#39;s level 5. 
               
               
                   
                   
                 shoulder, T4N1M0, stage IIIB 
                 Breslow&#39;s depth 10 MM 
               
               
                 4 
                 38 
                 Melanoma skin left foot, T4N0M0, 
                 Epithelioid melanoma, Clark&#39;s level 4. 
               
               
                   
                   
                 stage III 
                 Breslow&#39;s depth 8 MM 
               
               
                 5 
                 60 
                 Melanoma skin left chest T2N0M0, 
                 Epithelioid melanoma, Clark&#39;s level 2. 
               
               
                   
                   
                 stage II 
                 Breslow&#39;s depth 4 MM 
               
               
                 6 
                 78 
                 Melanoma of the left hand and 
                 Epithelioid melanoma, Clark&#39;s level 4. 
               
               
                   
                   
                 forearm skin, T3N0M0, stage III 
                 Breslow&#39;s depth 10 MM 
               
               
                 7 
                 60 
                 Melanoma of the skin area to the 
                 Epithelioid melanoma, Clark&#39;s level 5. 
               
               
                   
                   
                 left subscapular, T3N0M0, stage III 
                 Breslow&#39;s depth 11 MM 
               
               
                 8 
                 72 
                 Melanoma skin nasolabial triangle, 
                 Epithelioid melanoma, Clark&#39;s level 2. 
               
               
                   
                   
                 T2N2M0, stage II 
                 Breslow&#39;s depth 4 MM 
               
               
                 9 
                 48 
                 Melanoma of abdominal skin, 
                 Epithelioid melanoma, Clark&#39;s level 4. 
               
               
                   
                   
                 T4N2M0, stage IV 
                 Breslow&#39;s depth 14 MM 
               
               
                 10 
                 56 
                 Melanoma of abdominal skin, 
                 Epithelioid melanoma, Clark&#39;s level 5. 
               
               
                   
                   
                 T4N2M0, stage III 
                 Breslow&#39;s depth 12 MM 
               
               
                 11 
                 58 
                 Melanoma skin armpit, T4N2M0, 
                 Epithelioid melanoma, Clark&#39;s level 5. 
               
               
                   
                   
                 stage III 
                 Breslow&#39;s depth 10 MM 
               
               
                 12 
                 63 
                 Melanoma of the skin of the neck 
                 Epithelioid melanoma, Clark&#39;s level 5. 
               
               
                   
                   
                 occipital T4N1M0, stage III 
                 Breslow&#39;s depth 15 MM 
               
               
                 13 
                 71 
                 Melanoma of abdominal skin, 
                 Epithelioid melanoma, Clark&#39;s level 5. 
               
               
                   
                   
                 T4N2M0, stage III 
                 Breslow&#39;s depth 18 MM 
               
               
                 14 
                 43 
                 Melanoma skin left chest T2N0M0, 
                 Epithelioid melanoma, Clark&#39;s level 3. 
               
               
                   
                   
                 stage III 
                 Breslow&#39;s depth 6 MM 
               
               
                   
               
             
          
         
       
     
         [0072]    The results of using various antigens in the assay system for identification of autoantibodies such as IgG and IgA were shown. 
       EXAMPLES 
     Example 1  
     Detection of IgG and IgA Autoantibodies in Prostate Cancer 
       [0073]    Diagnoses of patients with prostate cancer were established on the basis of the following indicators: clinical examination with morphological confirmation of the diagnosis and on the basis of cancer markers (PSA). Only in this group included 30 patients. 
         [0074]    Immunoassay test (ELISA) of samples taken from prostate cancer patients and a control sample was made according to the procedure described. Considered positive samples had an optical density in the ELISA by 20% or more of the optical density of the control sample. 
       Results: 
       [0075]    When used as a Glu-plasminogen antigen in ELISA, the number of positive samples in prostate cancer patients was 83% for IgG and 86% for IgA. 
         [0076]    When used as an antigen heavy chain (Glu1-Arg561) in ELISA, the number of p itive samples in prostate cancer patients was 83% for IgG and 80% for IgA. 
         [0077]    When used as an antigen light chain (Val562-Asn791 in ELISA, the number of positive samples in prostate cancer patients was 57% for IgG and 63% for IgA. 
         [0078]    When used as an antigen miniplasminogen (Val442-Asn791) in ELISA, the number of positive samples in prostate cancer patients was 69% for IgG and 68% for IgA. 
         [0079]    When used as an antigen fragment K1-4, 5 (Lys78-Arg530) in ELISA, the number of positive samples in prostate cancer patients was 83% for IgG and 74% for IgA. 
         [0080]    When used as an antigen fragment K1-4 (Tyr80-Asn440) in ELISA, the number of positive samples in prostate cancer patients was 80% for IgG and 74% for IgA 
         [0081]    When used as an antigen fragment K1-3 (Tyr80-Val338) in ELISA, the number of positive samples in prostate cancer patients was 77% for IgG and 69% for IgA. 
         [0082]    When used as an antigen fragment K2-3 (Cys165-Val338) in ELISA, the number of positive samples in prostate cancer patients was 61% for IgG and 56% for IgA. 
         [0083]    When used as an antigen fragment K4-5 (Val355-Phe546) in ELISA, the number of positive samples in prostate cancer patients was 60% for IgG and 51% for IgA. 
         [0084]    When used as an antigen fragment K1 (Tur80-Glu164) in ELISA, the number of positive samples in prostate cancer patients was 45% for IgG and 39% for IgA. 
         [0085]    When used as an antigen fragment K4 (Val354-Ala439) in ELISA, the number of positive samples in prostate cancer patients was 44% for IgG and 30% for IgA. 
         [0086]    When used as an antigen fragment K5 (Ser441-Fhe546) in ELISA, the number of positive samples in prostate cancer patients was 56% for IgG and 36% for IgA. 
       Example 2  
     Diagnosis of Lung Cancer 
       [0087]    Diagnosis of patients with lung cancer have been established on the basis of a clinical study with morphological confirmation of the diagnosis. Only in this group included 20 patients. 
         [0088]    Immunoassay test (ELISA) of samples taken from patients with lung cancer and a control sample was made according to the procedure described. Considered positive samples had an optical density in the ELISA by 20% or more of the optical density of the control sample. 
       Results. 
       [0089]    When used as a Glu-plasminogen antigen in ELISA, the number of positive samples with lung cancer patients was 80% for IgG and 55% for IgA. 
         [0090]    When used as an antigen heavy chain (Glu1-Arg561) in ELISA, the number of positive samples in patients with lung cancer was 76% for IgG and 65% for IgA. 
         [0091]    When used as an antigen light chain (Val562-Asn791) in ELISA, the number of positive samples in patients with lung cancer was 45% for IgG and 75% for IgA. 
         [0092]    When used as an antigen miniplasminogen (Val442-Asn791) in ELISA, the number of positive samples in patients with lung cancer was 55% for IgG and 75% for IgA. 
         [0093]    When used as an antigen fragment K1-4, 5 (Lys78-Arg530) in ELISA, the number of positive samples in patients with lung cancer was 76% for IgG and 75% for IgA. 
         [0094]    When used as an antigen fragment K1-4 (Tyr80-Asn440) in ELISA, the number of positive samples in patients with lung cancer was 72% for IgG and 75% for IgA. 
         [0095]    When used as an antigen fragment K1-4 (Asn60-Pro447) in ELISA, the number of positive samples in patients with lung cancer was 45% for IgG and 75% for IgA. 
         [0096]    When used as an antigen fragment K1-4 (Lys78-Pro447) in ELISA, the number of positive samples in patients with lung cancer was 77% for IgG and 75% for IgA. 
         [0097]    When used as an antigen fragment K1-4 (Lys78-Pro446) in ELISA, the number of positive samples in patients with lung cancer was 77% for IgG and 75% for IgA. 
         [0098]    When used as an antigen fragment K1-4 (Lys78-Lys468) in ELISA, the number of positive samples in patients with lung cancer was 81% for IgG and 79% for IgA. 
         [0099]    When used as an antigen fragment K1-4 (Leu74-Leu451) in ELISA, the number of positive samples in patients with lung cancer was 81% for IgG and 75% for IgA. 
         [0100]    When used as an antigen fragment K1-3 (Tyr80-Val338) in ELISA, the number of positive samples in patients with lung cancer was 68% for IgG and 65% for IgA. 
         [0101]    When used as an antigen fragment K1-3 (Tyr80-Val354) in ELISA, the number of positive samples in patients with lung cancer was 68% for IgG and 65% for IgA. 
         [0102]    When used as an antigen fragment K1-3 (Leu74-Pro349) in ELISA, the number of positive samples in patients with lung cancer was 68% for IgG and 65% for IgA. 
         [0103]    When used as an antigen fragment K2-3 (Cys165-Val354) in ELISA, the number of positive samples in patients with lung cancer was 45% for IgG and 60% for IgA. 
         [0104]    When used as an antigen fragment K4-5 (Val355-Phe546) in ELISA, the number of positive samples in patients with lung cancer was 35% for IgG and 55% fo in ELISA r IgA. 
         [0105]    When used as an antigen fragment K1 (Tur80-Glu164) in ELISA, the number of positive samples in patients with lung cancer was 35% for IgG and 45% for IgA. 
         [0106]    When used as an antigen fragment K4 (Val354-Ala439) in ELISA, the number of positive samples in patients with lung cancer was 25% for IgG and 40% for IgA. 
         [0107]    When used as an antigen fragment K5 (Ser441-Fhe546) in ELISA, the number of positive samples in patients with lung cancer was 34% for IgG and 35% for IgA. 
       Example 3  
     Diagnosis of Breast Cancer 
       [0108]    Diagnoses of patients with breast cancer have been established on the basis of clinical examination with morphological confirmation of the diagnosis and on the basis of cancer markers (CA 15-3). Only in this group included 40 patients. 
         [0109]    Immunoassay test (ELISA) of samples taken from breast cancer patients and a control sample was made according to the procedure described. Considered positive samples had an optical density in the ELISA by 20% or more of the optical density of the control sample. 
       Results. 
       [0110]    When used as a Glu-plasminogen as an antigen in ELISA, the number of positive samples in patients with breast cancer was 63% for IgG and 53% for IgA. 
         [0111]    When used as an antigen heavy chain (Glu1-Arg561) in ELISA, the number of positive samples in patients with breast cancer was 65% for IgG and 65% for IgA. 
         [0112]    When used as an antigen light chain (Val562-Asn791) in ELISA, the number of positive samples in patients with breast cancer was 50% for IgG and 60% for IgA. 
         [0113]    When used as an antigen miniplasminogen (Val442-Asn791) in ELISA, the number of positive samples in patients with breast cancer was 60% for IgG and 65% for IgA. 
         [0114]    When used as an antigen fragment K1-4, 5 (Lys78-Arg530) in ELISA, the number of positive samples in patients with breast cancer was 66% for IgG and 65% for IgA. 
         [0115]    When used as an antigen fragment K1-4 (Tyr80-Asn440) in ELISA, the number of positive samples in patients with breast cancer was 67% for IgG and 65% for IgA. 
         [0116]    When used as an antigen fragment K1-3 (Tyr80-Val338) in ELISA, the number of positive samples in patients with breast cancer was 62% for IgG and 60% for IgA. 
         [0117]    When used as an antigen fragment K2-3 (Cys165-Val338) in ELISA, the number of positive samples in patients with breast cancer was 45% for IgG and 50% for IgA. 
         [0118]    When used as an antigen fragment K4-5 (Val355-Phe546) in ELISA, the number of positive samples in patients with breast cancer was 40% for IgG and 45% for IgA. 
         [0119]    When used as an antigen fragment K1 (Tur80-Glu164) in ELISA, the number of positive samples in patients with breast cancer was 35% for IgG and 35% for IgA. 
         [0120]    When used as an antigen fragment K4 (Val354-Ala439) in ELISA, the number of positive samples in patients with breast cancer was 25% for IgG and 30% for IgA. 
         [0121]    When used as an antigen fragment K5 (Ser441-Fhe546) in ELISA, the number of positive samples in patients with breast cancer was 25% for IgG and 30% for IgA. 
       Example 4  
     Diagnosis of Ovarian Cancer 
       [0122]    Diagnoses of patients with ovarian cancer were established on the basis of the following indicators: clinical examination with morphological confirmation of the diagnosis and on the basis of cancer markers (CA 125). Only in this group had 11 patients. 
         [0123]    Immunoassay test (ELISA) of samples from patients with ovarian cancer, and a control sample was made according to the procedure described. Considered positive samples had an optical density in the ELISA by 20% or more of the optical density of the control sample 
       Results. 
       [0124]    When used as a Glu-plasminogen as an antigen in ELISA, the number of positive samples from patients with ovarian cancer was 73% for IgG and 64% for IgA. 
         [0125]    When used as an antigen heavy chain (Glu1-Arg561) in ELISA, the number of positive samples from patients with ovarian cancer was 71% for IgG and 57% for IgA. 
         [0126]    When used as an antigen light chain (Val562-Asn791) in ELISA, the number of positive samples from patients with ovarian cancer was 50% for IgG and 45% for IgA. 
         [0127]    When used as an antigen miniplasminogen (Val442-Asn791) in ELISA, the number of positive samples from patients with ovarian cancer was 55% for IgG and 45% for IgA. 
         [0128]    When used as an antigen fragment K1-4, 5 (Lys78-Arg530) in ELISA, the number of positive samples from patients with ovarian cancer was 64% for IgG and 45% for IgA. 
         [0129]    When used as an antigen fragment K1-4 (Tyr80-Asn440) in ELISA, the number of positive samples from patients with ovarian cancer was 64% for IgG and 45% for IgA 
         [0130]    When used as an antigen fragment K1-3 (Tyr80-Val338) in ELISA, the number of positive samples from patients with ovarian cancer was 54% for IgG and 36% for IgA. 
         [0131]    When used as an antigen fragment K2-3 (Cys165-Val338) in ELISA, the number of positive samples from patients with ovarian cancer was 45% for IgG and 36% for IgA. 
         [0132]    When used as an antigen fragment K4-5 (Val355-Phe546) in ELISA, the number of positive samples from patients with ovarian cancer was 45% for IgG and 36% for IgA. 
         [0133]    When used as an antigen fragment K1 (Tur80-Glu164) in ELISA, the number of positive samples from patients with ovarian cancer was 27% for IgG and 27% for IgA. 
         [0134]    When used as an antigen fragment K4 (Val354-Ala439) in ELISA, the number of positive samples from patients with ovarian cancer was 27% for IgG and 27% for IgA. 
         [0135]    When used as an antigen fragment K5 (Ser441-Fhe546) in ELISA, the number of positive samples from patients with ovarian cancer was 27% for IgG and 36% for IgA. 
       Example 5  
     Melanoma 
       [0136]    Diagnoses of patients with melanoma have been confirmed on the basis of: clinical examination with morphological confirmation of the diagnosis. Only in this group included 14 patients. Immunoassay test (ELISA) of samples taken from patients with melanoma and a control sample was made according to the procedure described. Considered positive samples had an optical density in the ELISA by 20% or more of the optical density of the control sample. 
       Results. 
       [0137]    When used as a Glu-plasminogen as an antigen in ELISA, the number of positive samples in patients with melanoma, 78% for IgG and 37% for IgA. 
         [0138]    When used as an antigen heavy chain (Glu1-Arg561) in ELISA, the number of positive samples in patients with melanoma, 78% for IgG and 37% for IgA. 
         [0139]    When used as an antigen light chain (Val562-Asn791) in ELISA, the number of positive samples from melanoma patients was 62% for IgG and 43% for IgA. 
         [0140]    When used as an antigen minipiasminogen (Val442-Asn791) in IFA, the number of positive samples from melanoma patients was 62% for IgG and 43% for IgA. 
         [0141]    When used as an antigen fragment K1-4, 5 (Lys78-Arg530) in ELISA, the number of positive samples from melanoma patients was 72% for IgG and 37% for IgA. 
         [0142]    When used as an antigen fragment K1-4 (Tyr80-Asn440) in ELISA, the number of positive samples from melanoma patients was 72% for IgG and 37% for IgA. 
         [0143]    When used as an antigen fragment K1-3 (Tyr80-Val338) in ELISA, the number of positive samples from melanoma patients was 65% for IgG and 35% for IgA. 
         [0144]    When used as an antigen fragment K2-3 (Cys165-Val338) in ELISA, the number of positive samples from melanoma patients was 57% for IgG and 35% for IgA. 
         [0145]    When used as an antigen fragment K4-5 (Val355-Phe546) in ELISA, the number of positive samples from melanoma patients was 57% for IgG and 35% for IgA. 
         [0146]    When used as an antigen fragment K1 (Tur80-Glu164) in ELISA, the number of positive samples from melanoma patients was 35% for IgG and 29% for IgA. 
         [0147]    When used as an antigen fragment K4 (Val354-Ala439) in ELISA, the number of positive samples from melanoma patients was 35% for IgG and 29% for IgA. 
         [0148]    When used as an antigen fragment K5 (Ser441-Fhe546) in ELISA, the number of positive samples from melanoma patients was 35% for IgG and 29% for IgA. 
       Sequence Listing 
       [0000]    
       
         &lt;110&gt; Goufman Evgeny Iosifovich, Iakovlev Vasily Nikolaevich, Kanaev Alexey Alexeevich, Suleymanov Rustam Raisovich. 
         &lt;120&gt; Method for the diagnosis of cancer and enzyme-linked immunoassay (ELISA) kit for its application. 
       
     
         [0000]    
       
         
               
               
             
           
               
                 &lt;160&gt; Number sequences-20 
                   
               
               
                 &lt;210&gt; 1 
               
               
                 &lt;212&gt; Protein 
               
               
                 &lt;213&gt;  Homo sapiens   
               
               
                 &lt;400&gt; Description of the sequence  
               
               
                 SEQ ID NO: 1 
                   
               
               
                 Glu 1  Pro Leu Asp Asp Tyr Val Asn Thr Gln 10  Gly Ala Ser Leu Phe Ser Val Thr Lys Lys 20  Gln Leu 
                   
               
               
                   
               
               
                 Gly Ala Gly Ser Ile Glu Glu Cys 30  Ala Ala Lys Cys Glu Glu Asp Glu Glu Phe 40  Thr Cys Arg Ala 
               
               
                   
               
               
                 Phe Gln Tyr His Ser Lys 50  Glu Gln Gln Cys Val Ile Met Ala Glu Asn 60  Arg Lys Ser Ser Ile Ile Ile 
               
               
                   
               
               
                 Arg Met Arg 70  Asp Val Val Leu Phe Glu Lys Lys 78  Val Tyr 80  Leu Ser Glu Cys Lys Thr Gly Asn 
               
               
                   
               
               
                 Gly Lys 90  Asn Tyr Arg Gly Thr Met Ser Lys Thr Lys 100  Asn Gly Ile Thr Cys Gln Lys Trp Ser 
               
               
                   
               
               
                 Ser 110  Thr Ser Pro His Arg Pro Arg Phe Ser Pro 120  Ala Thr His Pro Ser Glu Gly Leu Glu Glu 130   
               
               
                   
               
               
                 Asn Tyr Cys Arg Asn Pro Asp Asn Asp Pro 140  Gln Gly Pro Trp Cys Tyr Thr Thr Asp Pro 150  Glu 
               
               
                   
               
               
                 Lys Arg Tyr Asp Tyr Cys Asp Ile Leu 160  Glu Cys Glu Glu Glu Cys Met His Cys Ser 170  Gly Glu 
               
               
                   
               
               
                 Asn Tyr Asp Gly Lys Ile Ser Lys 180  Thr Met Ser Gly Leu Glu Cys Gln Ala Trp 80  Asp Ser Gln Ser 
               
               
                   
               
               
                 Pro His Ala His Gly Tyr 200  Ile Pro Ser Lys Phe Pro Asn Lys Asn Leu 210  Lys Lys Asn Tyr Cys Arg 
               
               
                   
               
               
                 Asn Pro Asp Arg 220  Glu Leu Arg Pro Trp Cys Phe Thr Thr Asp 230  Pro Asn Lys Arg Trp Glu Leu 
               
               
                   
               
               
                 Cys Asp Ile 240  Pro Arg Cys Thr Thr Pro Pro Pro Ser Ser 250  Gly Pro Thr Tyr Gln Cys Leu Lys Gly 
               
               
                   
               
               
                 Thr 260  Gly Glu Asn Tyr Arg Gly Asn Val Ala Val Thr Val Ser Gly His Thr Cys Gln His Trp 280  Ser 
               
               
                   
               
               
                 Ala Gln Thr Pro His Thr His Asn Arg 290  Thr Pro Glu Asn Phe Pro Cys Lys An Leu 300  Asp Glu 
               
               
                   
               
               
                 Asn Tyr Cys Arg Asn Pro Asp Gly 310  Lys Arg Ala Pro Trp Cys His Thr Thr Asn 320  Ser Gln Val 
               
               
                   
               
               
                 Arg Trp Glu Tyr Cys Lys Ile 330  Pro Ser Cys Asp Ser Ser Pro Val Ser Thr 340  Glu Gln Leu Ala Pro 
               
               
                   
               
               
                 Thr Ala Pro Pro Glu 350  Leu Thr Pro Val Val Gln Asp Cys Tyr His 360  Gly Asp Gly Gln Ser Tyr Arg 
               
               
                   
               
               
                 Gly Thr Ser 370  Ser Thr Thr Thr Thr Gly Lys Lys Cys Gln 380  Ser Trp Ser Ser Met Thr Pro His Arg 
               
               
                   
               
               
                 His 390  Gln Lys Thr Pro Glu Asn Tyr Pro Asn Ala 400  Gly Leu Thr Met Asn Tyr Cys Arg Asn Pro 410   
               
               
                   
               
               
                 Asp Ala Asp Lys Gly Pro Trp Cys Phe Thr 420  Thr Asp Pro Ser Val Arg Trp Glu Tyr Cys 430  Asn 
               
               
                   
               
               
                 Leu Lys Lys Cys Ser Gly Thr Glu Ala 440  Ser Val Val Ala Pro Pro Pro Val Val Leu 450  Leu Pro Asp 
               
               
                   
               
               
                 Val Glu Thr Pro Ser Glu Glu 460  Asp Cys Met Phe Gly Asn Gly Lys Gly Tyr 470  Arg Gly Lys Aug 
               
               
                   
               
               
                 Ala Thr Thr Val Thr Gly 480  Thr Pro Cys Gln Asp Trp Ala Ala Gln Glu 490  Pro His Arg His Ser Ile 
               
               
                   
               
               
                 Phe Thr Pro Glu 500  Thr Asn Pro Arg Ala Gly Leu Glu Lys Asn 510  Tyr Cys Arg Asn Pro Asp Gly 
               
               
                   
               
               
                 Asp Val Gly 520  Gly Pro Trp Cys Tyr Thr Thr Asn Pro Arg 530  Lys Leu Tyr Asp Tyr Cys Asp Val 
               
               
                   
               
               
                 Pro Gln 540  Cys Ala Ala Pro Ser Phe Asp Cys Gly Lys 550  Pro Gln Val Glu Pro Lys Lys Cys Pro 
               
               
                   
               
               
                 Gly Arg Val Val Gly Gly Cys Val Ala His Pro 570  His Ser Trp Pro Trp Gln Val Ser Leu Arg 580   
               
               
                   
               
               
                 Thr Aug Phe Gly Met His Phe Cys Gly Gly 590  Thr Leu Ile Ser Pro Glu Trp Val Leu Thr 600  Ala Ala 
               
               
                   
               
               
                 His Cys Leu Glu Lys Ser Pro Arg 610  Pro Ser Ser Tyr Lys Val Ile Ley Gly Ala 620  His Gln Glu Val 
               
               
                   
               
               
                 Asn Leu Glu Pro His Val 630  Gln Glu Ile Glu Val Ser Aug Leu Phe Leu 640  Glu Pro Thr Aug Lys 
               
               
                   
               
               
                 Asp Ile Ala Leu Leu 650  Lys Leu Ser Ser Pro Ala Val Ile Thr Asp 660  Lys Val Ile Pro Ala Cys Leu 
               
               
                   
               
               
                 Pro Ser Pro 670  Asn Tyr Val Val Ala Asp Aug Thr Glu Cys 680  Phe Ile Thr Gly Trp Gly Glu Thr Gln 
               
               
                   
               
               
                 Gly 690  Thr Phe Gly Ala Gly Leu Leu Lys Glu Ala 700  Gln Leu Pro Val Ile Glu Asn Lys Val Cys 710   
               
               
                   
               
               
                 Asn Arg Tyr Glu Phe Leu Asn Gly Arg Va 720 l Gln Ser Thr Glu Leu Cys Ala Gly His Lue 730  Ala 
               
               
                   
               
               
                 Gly Gly Thr Asp Ser Cys Gln Gly Asp 740  Ser Gly Gly Pro Leu Val Cys Phe Glu Lys 750  Asp Lys Tyr 
               
               
                   
               
               
                 Ile Leu Gln Gly Val Thr Ser 760  Trp Gly Leu Gly Cys Ala Aug Pro Asn Lys 770  Pro Gly Val Tyr Val 
               
               
                   
               
               
                 Aug Val Ser Aug Phe 780  Val Thr Trp Ile Glu Gly Val Met Arg As n790  Asn 
               
               
                   
               
               
                 &lt;210&gt; 2 
               
               
                 &lt;212&gt; Protein 
               
               
                 &lt;213&gt;  Homo sapiens   
               
               
                 &lt;400&gt; Description of the sequence  
               
               
                 SEQ ID NO: 2 
                   
               
               
                 Lys 78  Val Tyr 80  Leu Ser Glu Cys Lys Thr Gly Asn Gly Lys 90  Asn Tyr Aug Gly Thr Met Ser Lys Thr 
                   
               
               
                   
               
               
                 Lys 100  Asn Gly Ile Thr Cys Gln Lys Trp Ser Ser 110  Thr Ser Pro His Aug Pro Aug Phe Ser Pro 120   
               
               
                   
               
               
                 Ala Thr His Pro Ser Glu Gly Leu Glu Glu 130  Asn Tyr Cys Arg Asn Pro Asp Asn Asp Pro 140  Gln 
               
               
                   
               
               
                 Gly Pro Trp Cys Tyr Thr Thr Asp Pro 150  Glu Lys Aug Tyr Asp Tyr Cys Asp Ile Leu 160  Glu Cys Glu 
               
               
                   
               
               
                 Glu Glu Cys Met His Cys Ser 170  Gly Glu Ash Tyr Asp Gly Lys Ile Ser Lys 180  Thr Met Ser Gly Leu 
               
               
                   
               
               
                 Glu Cys Gln Ala Trp 80  Asp Ser Gln Ser Pro His Ala His Gly Tyr 200  Ile Pro Ser Lys Phe Pro Asn 
               
               
                   
               
               
                 Lys Asn Leu 210  Lys Lys Asn Tyr Cys Arg Asn Pro Asp Arg 220  Glu Leu Arg Pro Trp Cys Phe Thr 
               
               
                   
               
               
                 Thr Asp 230  Pro Asn Lys Arg Trp Glu Leu Cys Asp Ile 240  Pro Aug Cys Thr Thr Pro Pro Pro Ser 
               
               
                   
               
               
                 Ser 250  Gly Pro Thr Tyr Gln Cys Leu Lys Gly Thr 260  Gly Glu Asn Tyr Arg Gly Asn Val Ala Val Thr 
               
               
                   
               
               
                 Val Ser Gly His Thr Cys Gln His Trp 280  Ser Ala Gln Thr Pro His Thr His Asn Arg 290  Thr Pro Glu 
               
               
                   
               
               
                 Asn Phe Pro Cys Lys Asn Leu 300  Asp Glu Asn Tyr Cys Arg Asn Pro Asp Gly 310  Lys Arg Ala Pro 
               
               
                   
               
               
                 Trp Cys His Thr Thr Asn 320  Ser Gln Val Arg Trp Glu Tyr Cys Lys Ile 330  Pro Ser Cys Asp Ser Ser 
               
               
                   
               
               
                 Pro Val Ser Thr 340  Glu Gln Leu Ala Pro Thr Ala Pro Pro Glu 350  Leu Thr Pro Val Val Gln Asp Cys 
               
               
                   
               
               
                 Tyr His 360  Gly Asp Gly Gln Ser Tyr Arg Gly Thr Ser 370  Ser Thr Thr Thr Thr Gly Lys Lys Cys 
               
               
                   
               
               
                 Gln 380  Ser Trp Ser Ser Met Thr Pro His Arg His 390  Gln Lys Thr Pro Glu Asn Tyr Pro Asn Ala 400   
               
               
                   
               
               
                 Gly Leu Thr Met Asn Tyr Cys Arg Asn Pro 410  Asp Ala Asp Lys Gly Pro Trp Cys Phe Thr 420  Thr 
               
               
                   
               
               
                 Asp Pro Ser Val Arg Trp Glu Tyr Cys 430  Asn Leu Lys Lys Cys Ser Gly Thr Glu Ala 440  Ser Val Val 
               
               
                   
               
               
                 Ala Pro Pro Pro Val Val Leu 450  Leu Pro Asp Val Glu Thr Pro Ser Glu Glu 460  Asp Cys Met Phe 
               
               
                   
               
               
                 Gly Asn Gly Lys Gly Tyr 470  Arg Gly Lys Arg Ala Thr Thr Val Thr Gly 480  Thr Pro Cys Gln Asp Trp 
               
               
                   
               
               
                 Ala Ala Gln Glu 490  Pro His Arg His Ser Ile Phe Thr Pro Glu 500  Thr Asn Pro Arg Ala Gly Leu Glu 
               
               
                   
               
               
                 Lys Asn 510  Tyr Cys Arg Asn Pro Asp Gly Asp Val Gly 520  Gly Pro Trp Cys Tyr Thr Thr Asn Pro 
               
               
                   
               
               
                 Arg 530  Lys Leu Tyr Asp Tyr Cys Asp Val Pro Gln 540  Cys Ala Ala Pro Ser Phe Asp Cys Gly Lys 550   
               
               
                   
               
               
                 Pro Gln Val Glu Pro Lys Lys Cys Pro Gly 560  Arg Val Val Gly Gly Cys Val Ala His Pro 570  His Ser 
               
               
                   
               
               
                 Trp Pro Trp Gln Val Ser Leu Arg 580  Thr Arg Phe Gly Met His Phe Cys Gly Gly 590  Thr Leu Ile 
               
               
                   
               
               
                 Ser Pro Glu Trp Val Leu Thr 600  Ala Ala His Cys Leu Glu Lys Ser Pro Arg 610  Pro Ser Ser Tyr Lys 
               
               
                   
               
               
                 Val Ile Leu Gly Ala 620  His Gln Glu Val Asn Leu Glu Pro His Val 630  Gln Glu Ile Glu Val Ser Arg 
               
               
                   
               
               
                 Leu Phe Leu 640  Glu Pro Thr Arg Lys Asp Ile Ala Leu Leu 650  Lys Leu Ser Ser Pro Ala Val Ile Thr 
               
               
                   
               
               
                 Asp 660  Lys Val Ile Pro Ala Cys Leu Pro Ser Pro 670  Asn Tyr Val Val Asp Arg Thr Glu Cys 680   
               
               
                   
               
               
                 Phe Ile Thr Gly Trp Gly Glu Thr Gln Gly 690  Thr Phe Gly Ala Gly Leu Leu Lys Glu Ala 700  Gln Leu 
               
               
                   
               
               
                 Pro Val Ile Glu Asn Lys Val Cys 710  Asn Arg Tyr Glu Phe Leu Asn Gly Arg Va 720 l Gln Ser Thr Glu 
               
               
                   
               
               
                 Leu Cys Ala Gly His Leu 730  Ala Gly Gly Thr Asp Ser Cys Gln Gly Asp 740  Ser Gly Gly Pro Leu 
               
               
                   
               
               
                 Val Cys Phe Glu Lys 750  Asp Lys Tyr Ile Leu Gln Gly Val Thr Ser 760  Trp Gly Leu Gly Cys Ala Arg 
               
               
                   
               
               
                 Pro Asn Lys 770  Pro Gly Val Tyr Val Arg Val Ser Arg Phe 780  Val Thr Trp Ile Glu Gly Val Met Arg 
               
               
                   
               
               
                 Asn 790  Asn 
               
               
                   
               
               
                 &lt;210&gt; 3 
               
               
                 &lt;212&gt; protein 
               
               
                 &lt;213&gt;  Homo sapiens   
               
               
                 &lt;400&gt; Description of the sequence  
               
               
                 SEQ ID NO: 3 
                   
               
               
                 Glu 1  Pro Leu Asp Asp Tyr Val Asn Thr Gln 10  Gly Ala Ser Leu Phe Ser Val Thr Lys Lys 20  Gln Leu 
                   
               
               
                   
               
               
                 Gly Ala Gly Ser Ile Glu Glu Cys 30  Ala Ala Lys Cys Glu Glu Asp Glu Glu Phe 40  Thr Cys Arg Ala 
               
               
                   
               
               
                 Phe Gln Tyr His Ser Lys 50  Glu Gln Gln Cys Val Ile Met Ala Glu Asn 60  Arg Lys Ser Ser Ile Ile Ile 
               
               
                   
               
               
                 Arg Met Arg 70  Asp Val Val Leu Phe Glu Lys Lys 78  Val Tyr 80  Leu Ser Glu Cyc Lys Thr Gly Asn 
               
               
                   
               
               
                 Gly Lys 90  Asn Tyr Arg Gly Thr Met Ser Lys Thr Lys 100  Asn Gly Ile Thr Cys Gln Lys Trp Ser 
               
               
                   
               
               
                 Ser 110  Thr Ser Pro His Arg Pro Arg Phe Ser Pro 120  Ala Thr His Pro Ser Glu Gly Leu Glu Glu 130   
               
               
                   
               
               
                 Asn Tyr Cys Arg Asn Pro Asp Asn Asp Pro 140  Gln Gly Pro Trp Cys Tyr Thr Thr Asp Pro 150  Glu 
               
               
                   
               
               
                 Lys Arg Tyr Asp Tyr Cys Asp Ile Leu 160  Glu Cys Glu Glu Glu Cys Met His Cys Ser 170  Gly Glu 
               
               
                   
               
               
                 Asn Tyr Asp Gly Lys Ile Ser Lys 180  Thr Met Ser Gly Leu Glu Cys Gln Ala Trp 80  Asp Ser Gln Ser 
               
               
                   
               
               
                 Pro His Ala His Gly Tyr 200  Ile Pro Ser Lys Phe Pro Asn Lys Asn Leu 210  Lys Lys Asn Tyr Cys Arg 
               
               
                   
               
               
                 Asn Pro Asp Arg 220  Glu Leu Arg Pro Trp Cys Phe Thr Thr Asp 230  Pro Asn Lys Arg Trp Glu Leu 
               
               
                   
               
               
                 Cys Asp Ile 240  Pro Arg Cys Thr Thr Pro Pro Pro Ser Ser 250  Gly Pro Thr Tyr Gln Cys Leu Lys Gly 
               
               
                   
               
               
                 Thr 260  Gly Glu Asn Tyr Arg Gly Asn Val Ala Val Thr Val Ser Gly His Thr Cys Gln His Trp 280  Ser 
               
               
                   
               
               
                 Ala Gln Thr Pro His Thr His Asn Arg 290  Thr Pro Glu Asn Phe Pro Cys Lys Asn Leu 300  Asp Glu 
               
               
                   
               
               
                 Asn Tyr Cys Arg Asn Pro Asp Gly 310  Lys Arg Ala Pro Trp Cys His Thr Thr Asn 320  Ser Gln Val 
               
               
                   
               
               
                 Arg Trp Glu Tyr Cys Lys Ile 330  Pro Ser Cys Asp Ser Ser Pro Val Ser Thr 340  Glu Gln Leu Ala Pro 
               
               
                   
               
               
                 Thr Ala Pro Pro Glu 350  Leu Thr Pro Val Val Gln Asp Cys Tyr His 360  Gly Asp Gly Gln Ser Tyr Arg 
               
               
                   
               
               
                 Gly Thr Ser 370  Ser Thr Thr Thr Thr Gly Lys Lys Cys Gln 38 0 Ser Trp Ser Ser Met Thr Pro His Arg 
               
               
                   
               
               
                 His 390  Gln Lys Thr Pro Glu Asn Tyr Pro Asn Ala 400  Gly Leu Thr Met Asn Tyr Cys Arg Asn Pro 410   
               
               
                   
               
               
                 Asp Ala Asp Lys Gly Pro Trp Cys Phe Thr 420  Thr Asp Pro Ser Val Arg Trp Glu Tyr Cys 430  Asn 
               
               
                   
               
               
                 Leu Lys Lys Cys Ser Gly Thr Glu Ala 440  Ser Val Val Ala Pro Pro Pro Val Val Leu 450  Leu Pro Asp 
               
               
                   
               
               
                 Val Glu Thr Pro Ser Glu Glu 460  Asp Cys Met Phe Gly Asn Gly Lys Gly Tyr 470  Arg Gly Lys Arg 
               
               
                   
               
               
                 Ala Thr Thr Val Thr Gly 480  Thr Pro Cys Gln Asp Trp Ala Ala Gln Glu 490  Pro His Arg His Ser Ile 
               
               
                   
               
               
                 Phe Thr Pro Glu 500  Thr Asn Pro Arg Ala Gly Leu Glu Lys Asn 510  Tyr Cys Arg Asn Pro Asp Gly 
               
               
                   
               
               
                 Asp Val Gly 520  Gly Pro Trp Cys Tyr Thr Thr Asn Pro Arg 530  Lys Leu Tyr Asp Tyr Cys Asp Val 
               
               
                   
               
               
                 Pro Gln 540  Cys Ala Ala Pro Ser Phe Asp Cys Gly Lys 550  Pro Gln Val Glu Pro Lys Lys Cys Pro 
               
               
                   
               
               
                 Gly 560  Arg 
               
               
                   
               
               
                 &lt;210&gt; 3 
               
               
                 &lt;212&gt; protein 
               
               
                 &lt;213&gt;  Homo sapiens   
               
               
                 &lt;400&gt; Description of the sequence  
               
               
                 SEQ ID NO: 4 
                   
               
               
                 Lys 78  Val Tyr 80  Leu Ser Glu Cys Lys Thr Gly Asn Gly Lys 90  Asn Tyr Arg Gly Thr Met Ser Lys Thr 
                   
               
               
                   
               
               
                 Lys 100  Asn Gly Ile Thr Cys Gln Lys Trp Ser Ser 110  Thr Ser Pro His Arg Pro Arg Phe Ser Pro 120   
               
               
                   
               
               
                 Ala Thr His Pro Set Glu Gly Leu Glu Glu 130  Asn Tyr Cys Arg Asn Pro Asp Asn Asp Pro 140  Gln 
               
               
                   
               
               
                 Gly Pro Trp Cys Tyr Thr Thr Asp Pro 150  Glu Lys Arg Tyr Asp Tyr Cys Asp Ile Leu 160  Glu Cys Glu 
               
               
                   
               
               
                 Glu Glu Cys Met His Cys Ser 170  Gly Glu Asn Tyr Asp Gly Lys Ile Ser Lys 180  Thr Met Ser Gly Leu 
               
               
                   
               
               
                 Glu Cys Gln Ala Trp 80  Asp Ser Gln Ser Pro His Ala His Gly Tyr 200  Ile Pro Ser Lys Phe Pro Asn 
               
               
                   
               
               
                 Lys Asn Leu 210  Lys Lys Asn Tyr Cys Arg Asn Pro Asp Arg 220  Glu Leu Arg Pro Trp Cys Phe Thr 
               
               
                   
               
               
                 Thr Asp 230  Pro Asn Lys Arg Trp Glu Leu Cys Asp Ile 240  Pro Arg Cys Thr Thr Pro Pro Pro Ser 
               
               
                   
               
               
                 Ser 250  Gly Pro Thr Tyr Gln Cys Leu Lys Gly Thr 260  Gly Glu Asn Tyr Arg Gly Asn Val Ala Val Thr 
               
               
                   
               
               
                 Val Ser Gly His Thr Cys Gln His Trp 280  Ser Ala Gln Thr Pro His Thr His Asn Arg 290  Thr Pro Glu 
               
               
                   
               
               
                 Asn Phe Pro Cys Lys Asn Leu 300  Asp Glu Asn Tyr Cys Arg Asn Pro Asp Gly 310  Lys Arg Ala Pro 
               
               
                   
               
               
                 Trp Cys His Thr Thr Asn 320  Ser Gln Val Arg Trp Glu Tyr Cys Lys Ile 330  Pro Ser Cys Asp Ser Ser 
               
               
                   
               
               
                 Pro Val Ser Thr 340  Glu Gln Leu Ala Pro Thr Ala Pro Pro Glu 350  Leu Thr Pro Val Val Gln Asp Cys 
               
               
                   
               
               
                 Tyr His 360  Gly Asp Gly Gln Ser Tyr Arg Gly Thr Ser 370  Ser Thr Thr Thr Thr Gly Lys Lys Cys 
               
               
                   
               
               
                 Gle 380  Ser Trp Ser Ser Met Thr Pro His Arg His 390  Gln Lys Thr Pro Glu Asn Tyr Pro Asn Ala400 
               
               
                   
               
               
                 Gly Leu Thr Met Asn Tyr Cys Arg Asn Pro 410  Asp Ala Asp Lys Gly Pro Trp Cys Phe Thr 420  Thr 
               
               
                   
               
               
                 Asp Pro Ser Val Arg Trp Glu Tyr Cys 430  Asn Leu Lys Lys Cys Ser Gly Thr Glu Ala 440  Ser Val Val 
               
               
                   
               
               
                 Ala Pro Pro Pro Val Val Leu 450  Leu Pro Asp Val Glu Thr Pro Ser Glu Glu 460  Asp Cys Met Phe 
               
               
                   
               
               
                 Gly Asn Gly Lys Gly Tyr 470  Arg Gly Lys Arg Ala Thr Thr Val Thr Gly 480  Thr Pro Cys Gln Asp Trp 
               
               
                   
               
               
                 Ala Ala Gln Glu 490  Pro His Arg His Ser Ile Phe Thr Pro Glu 500  Thr Asn Pro Arg Ala Gly Leu Glu 
               
               
                   
               
               
                 Lys Asn 510  Tyr Cys Arg Asn Pro Asp Gly Asp Val Gly 520  Gly Pro Trp Cys Tyr Thr Thr Asn Pro 
               
               
                   
               
               
                 Arg 530  Lys Leu Tyr Asp Tyr Cys Asp Val Pro Gln 540  Cys Ala Ala Pro Ser Phe Asp Cys Gly Lys550 
               
               
                   
               
               
                 Pro Gln Val Glu Pro Lys Lys Cys Pro Gly 560  Arg 
               
               
                   
               
               
                 &lt;210&gt; 5 
               
               
                 &lt;212&gt; protein 
               
               
                 &lt;213&gt;  Homo sapiens   
               
               
                 &lt;400&gt; Description of the sequence  
               
               
                 SEQ ID NO: 5 
                   
               
               
                 Val Val Gly Gly Cys Val Ala His Pro 570  His Ser Trp Pro Trp Gln Val Ser Leu Arg 580  Thr Arg Phe 
                   
               
               
                   
               
               
                 Gly Met His Phe Cys Gly Gly 590  Thr Leu Ile Ser Pro Glu Trp Val Leu Thr 600  Ala Ala His Cys Leu 
               
               
                   
               
               
                 Glu Lys Ser Pro Arg 610  Pro Ser Ser Tyr Lys Val Ile Leu Gly Ala 620  His Gln Glu Val Asn Leu Glu 
               
               
                   
               
               
                 Pro His Val 630  Gln Glu Ile Glu Val Ser Arg Leu Phe Leu 640  Glu Pro Thr Arg Lys Asp Ile Ala Leu 
               
               
                   
               
               
                 Leu 650  Lys Leu Ser Ser Pro Ala Val Ile Thr Asp 660  Lys Val Ile Pro Ala Cys Leu Pro Set Pro 670   
               
               
                   
               
               
                 Asn Tyr Val Val Ala Asp Arg Thr Glu Cys 680  Phe Ile Thr Gly Trp Gly Glu Thr Gln Gly 690  Thr Phe 
               
               
                   
               
               
                 Gly Ala Gly Leu Leu Lys Glu Ala 700  Gln Leu Pro Val Ile Glu Asn Lys Val Cys 710  Asn Arg Tyr Glu 
               
               
                   
               
               
                 Phe Leu Asn Gly Arg Va 720 l Gln Ser Thr Glu Leu Cys Ala Gly His Leu 730  Ala Gly Gly Thr Asp 
               
               
                   
               
               
                 Ser Cys Gln Gly Asp 740  Ser Gly Gly Pro Leu Val Cyc Phe Glu Lys 750  Asp Lys Tyr Ile Leu Gln Gly 
               
               
                   
               
               
                 Val Thr Ser 760  Trp Gly Leu Gly Cys Ala Arg Pro Asn Lys 770  Pro Gly Val Tyr Val Arg Val Ser Arg 
               
               
                   
               
               
                 Phe 780  Val Thr Trp Ile Glu Gly Val Met Arg Asn 790  Asn 
               
               
                   
               
               
                 &lt;210&gt; 6 
               
               
                 &lt;212&gt; protein 
               
               
                 &lt;213&gt;  Homo sapiens   
               
               
                 &lt;400&gt; Description of the sequence  
               
               
                 SEQ ID NO: 6 
                   
               
               
                 Tyr 80  Leu Ser Glu Cys Lys Thr Gly Asn Gly Lys 90  Asn Tyr Arg Gly Thr Met Ser Lys Thr Lys 100   
                   
               
               
                   
               
               
                 Asn Gly Ile Thr Cys Gln Lys Trp Ser Ser 110  Thr Ser Pro His Arg Pro Arg Phe Ser Pro 120  Ala Thr 
               
               
                   
               
               
                 His Pro Ser Glu Gly Leu Glu Glu 130  Asn Tyr Cys Arg Asn Pro Asp Asn Asp Pro 140  Gln Gly Pro 
               
               
                   
               
               
                 Trp Cys Tyr Thr Thr Asp Pro 150  Glu Lys Arg Tyr Asp Tyr Cys Asp Ile Leu 160  Glu Cys Glu Glu Glu 
               
               
                   
               
               
                 Cys Met His Cys Ser 170  Gly Glu Asn Tyr Asp Gly Lys Ile Ser Lys 180  Thr Met Ser Gly Leu Glu Cys 
               
               
                   
               
               
                 Gln Ala Trp 80  Asp Ser Gln Ser Pro His Ala His Gly Tyr 200  Ile Pro Ser Lys Phe Pro Asn Lys Asn 
               
               
                   
               
               
                 Leu 210  Lys Lys Asn Tyr Cys Arg Asn Pro Asp Arg 220  Glu Leu Arg Pro Trp Cys Phe Thr Thr 
               
               
                   
               
               
                 Asp 230  Pro Asn Lys Arg Trp Glu Leu Cys Asp Ile 240  Pro Arg Cys Thr Thr Pro Pro Pro Ser Ser 250   
               
               
                   
               
               
                 Gly Pro Thr Tyr Gln Cys Leu Lys Gly Thr 260  Gly Glu Asn Tyr Arg Gly Asn Val Ala Val Thr Val 
               
               
                   
               
               
                 Ser Gly His Thr Cys Gln His Trp 280  Ser Ala Gln Thr Pro His Thr His Asn Arg 290  Thr Pro Glu An 
               
               
                   
               
               
                 Phe Pro Cys Lys Asn Leu 300  Asp Glu Asn Tyr Cys Arg Asn Pro Asp Gly 310  Lys Arg Ala Pro Trp 
               
               
                   
               
               
                 Cys His Thr Thr Asn 320  Ser Gln Val Arg Trp Glu Tyr Cys Lys Ile 330  Pro Ser Cys Asp Ser Ser Pro 
               
               
                   
               
               
                 Val Ser Thr 340  Glu Gln Leu Ala Pro Thr Ala Pro Pro Glu 350  Leu Thr Pro Val Val Gln Asp Cys Tyr 
               
               
                   
               
               
                 His 360  Gly Asp Gly Gln Ser Tyr Arg Gly Thr Ser 370  Ser Thr Thr Thr Thr Gly Lys Lys Cys Gln 380   
               
               
                   
               
               
                 Ser Trp Ser Ser Met Thr Pro His Arg His 390  Gln Lys Thr Pro Glu Asn Tyr Pro Asn Ala 400  Gly 
               
               
                   
               
               
                 Leu Thr Met Asn Tyr Cys Arg Asn Pro 410  Asp Ala Asp Lys Gly Pro Trp Cys Phe Thr 420  Thr Asp 
               
               
                   
               
               
                 Pro Ser Val Arg Trp Glu Tyr Cys 430  Asn Leu Lys Lys Cys Ser Gly Thr Glu Ala 440   
               
               
                   
               
               
                 &lt;210&gt; 7 
               
               
                 &lt;212&gt; protein 
               
               
                 &lt;213&gt;  Homo sapiens   
               
               
                 &lt;400&gt; Description of the sequence  
               
               
                 SEQ ID NO: 7 
                   
               
               
                 Tyr 80  Leu Ser Glu Cys Lys Thr Gly Asn Gly Lys 90  Asn Tyr Arg Gly Thr Met Ser Lys Thr Lys 100   
                   
               
               
                   
               
               
                 Asn Gly Ile Thr Cys Gln Lys Trp Ser Ser 110  Thr Ser Pro His Arg Pro Arg Phe Ser Pro 120  Ala Thr 
               
               
                   
               
               
                 His Pro Ser Glu Gly Leu Glu Glu 130  Asn Tyr Cys Arg Asn Pro Asp Asn Asp Pro 140  Gln Gly Pro 
               
               
                   
               
               
                 Trp Cys Tyr Thr Thr Asp Pro 150  Glu Lys Arg Tyr Asp Tyr Cys Asp Ile Leu 160  Glu Cys Glu Glu Glu 
               
               
                   
               
               
                 Cys Met His Cys Ser 170  Gly Glu Asn Tyr Asp Gly Lys Ile Ser Lys 180  Thr Met Ser Gly Leu Glu Cys 
               
               
                   
               
               
                 Gln Ala Trp 80  Asp Ser Gln Ser Pro His Ala His Gly Tyr 200  Ile Pro Ser Lys Phe Pro Asn Lys Asn 
               
               
                   
               
               
                 Leu 210  Lys Lys Asn Tyr Cys Arg Asn Pro Asp Arg 220  Glu Leu Arg Pro Trp Cys Phe Thr Thr 
               
               
                   
               
               
                 Asp 230  Pro Asn Lys Arg Trp Glu Leu Cys Asp Ile 240  Pro Arg Cys Thr Thr Pro Pro Pro Ser Ser 250   
               
               
                   
               
               
                 Gly Pro Thr Tyr Gln Cys Leu Lys Gly Thr 260  Gly Glu Asn Tyr Arg Gly Asn Val Ala Val Thr Val 
               
               
                   
               
               
                 Ser Gly His Thr Cys Gln His Trp 280  Ser Ala Gln Thr Pro His Thr His Asn Arg 290  Thr Pro Glu Asn 
               
               
                   
               
               
                 Phe Pro Cys Lys Asn Leu 300  Asp Glu Asn Tyr Cys Arg Asn Pro Asp Gly 310  Lys Arg Ala Pro Trp 
               
               
                   
               
               
                 Cys His Thr Thr Asn 320  Ser Gln Val Arg Trp Glu Tyr Cys Lys Ile 330  Pro Ser Cys Asp Ser Ser Pro 
               
               
                   
               
               
                 Val 
               
               
                   
               
               
                 &lt;210&gt; 8 
               
               
                 &lt;212&gt; protein 
               
               
                 &lt;213&gt;  Homo sapiens   
               
               
                 &lt;400&gt; Description of the sequence  
               
               
                 SEQ ID NO: 8 
                   
               
               
                 Tyr 80  Leu Ser Glu Cys Lys Thr Gly Asn Gly Lys 90  Asn Tyr Arg Gly Thr Met Ser Lys Thr Lys 100   
                   
               
               
                   
               
               
                 Asn Gly Ile Thr Cys Gln Lys Trp Ser Ser 110  Thr Ser Pro His Arg Pro Arg Phe Ser Pro 120  Ala Thr 
               
               
                   
               
               
                 His Pro Ser Glu Gly Leu Glu Glu 130  Asn Tyr Cys Arg Asn Pro Asp Asn Asp Pro 140  Gln Gly Pro 
               
               
                   
               
               
                 Trp Cys Tyr Thr Thr Asp Pro 150  Glu Lys Arg Tyr Asp Tyr Cys Asp Ile Leu 160  Glu Cys Glu Glu Glu 
               
               
                   
               
               
                 Cys Met His Cys Ser 170  Gly Glu Asn Tyr Asp Gly Lys Ile Ser Lys 180  Thr Met Ser Gly Leu Gln Cys 
               
               
                   
               
               
                 Gln Ala Trp 80  Asp Ser Gln Ser Pro His Ala His Gly Tyr 200  Ile Pro Ser Lys Phe Pro Asn Lys Asn 
               
               
                   
               
               
                 Leu 210  Lys Lys Asn Tyr Cys Arg Asn Pro Asp Arg 220  Glu Leu Arg Pro Trp Cys Phe Thr Thr 
               
               
                   
               
               
                 Asp 230  Pro Asn Lys Arg Trp Gln Leu Cys Asp Ile 240  Pro Arg Cys Thr Thr Pro Pro Pro Ser Ser 250   
               
               
                   
               
               
                 Gly Pro Thr Tyr Gln Cys Leu Lys Gly Thr 260  Gly Glu Asn Tyr Arg Gly Asn Val Ala Val Thr Val 
               
               
                   
               
               
                 Ser Gly His Thr Cys Gln His Trp 280  Ser Ala Gln Thr Pro His Thr His Asn Arg 290  Thr Pro Gln Asn 
               
               
                   
               
               
                 Phe Pro Cys Lys Asn Leu 300  Asp Gln Asn Tyr Cys Arg Asn Pro Asp Gly 310  Lys Arg Ala Pro Trp 
               
               
                   
               
               
                 Cys His Thr Thr Asn 320  Ser Gln Val Arg Trp Gln Tyr Cys Lys Ile 330  Pro Ser Cys Asp Ser Ser Pro 
               
               
                   
               
               
                 Val Ser Thr 340  Gln Gln Leu Ala Pro Thr Ala Pro Pro Glu 350  Leu Thr Pro Val 
               
               
                   
               
               
                 &lt;210&gt; 9 
               
               
                 &lt;212&gt; protein 
               
               
                 &lt;213&gt;  Homo sapiens   
               
               
                 &lt;400&gt; Description of the sequence  
               
               
                 SEQ ID NO: 9 
                   
               
               
                 Asn 60  Arg Lys Ser Ser Ile Ile Ile Arg Met Arg 70  Asp Val Val Leu Phe Glu Lys Lys 78  Val Tyr 80  Leu 
                   
               
               
                   
               
               
                 Ser Glu Cys Lys Thr Gly Asn Gly Lys 90  Asn Tyr Arg Gly Thr Met Ser Lys Thr Lys 100  Asn Gly Ile 
               
               
                   
               
               
                 Thr Cys Gln Lys Trp Ser Ser 110  Thr Ser Pro His Arg Pro Arg Phe Ser Pro 120  Ala Thr His Pro Ser 
               
               
                   
               
               
                 Glu Gly Leu Glu Glu 130  Asn Tyr Cys Arg Asn Pro Asp Asn Asp Pro 140  Gln Gly Pro Trp Cys Tyr 
               
               
                   
               
               
                 Thr Thr Asp Pro 150  Glu Lys Arg Tyr Asp Tyr Cys Asp Ile Leu 160  Gln Cys Gln Glu Glu Cys Met 
               
               
                   
               
               
                 His Cys Ser 170  Gly Gln Asn Tyr Asp Gly Lys Ile Ser Lys 180  Thr Met Ser Gly Leu Gln Cys Gln Ala 
               
               
                   
               
               
                 Trp 80  Asp Ser Gln Ser Pro His Ala His Gly Tyr 200  Ile Pro Ser Lys Phe Pro Asn Lys Asn Leu 210   
               
               
                   
               
               
                 Lys Lys Asn Tyr Cys Arg Asn Pro Asp Arg 220  Glu Leu Arg Pro Trp Cys Phe Thr Thr Asp 230  Pro 
               
               
                   
               
               
                 Asn Lys Arg Trp Gly Leu Cys Asp Ile 240  Pro Arg Cys Thr Thr Pro Pro Pro Ser Ser 250  Gly Pro Thr 
               
               
                   
               
               
                 Tyr Gln Cys Leu Lys Gly Thr 260  Gly Gln Asn Tyr Arg Gly Asn Val Ala Val Thr Val Ser Gly His Thr 
               
               
                   
               
               
                 Cys Gln His Trp 280  Ser Aia Gln Thr Pro His Thr His Asn Arg 290  Thr Pro Gln Asn Phe Pro Cys Lys 
               
               
                   
               
               
                 Asn Leu 300  Asp Gln Asn Tyr Cys Arg Asn Pro Asp Gly 310  Lys Arg Ala Pro Trp Cys His Thr Thr 
               
               
                   
               
               
                 Asn 320  Ser Gln Val Arg Trp Gln Tyr Cys Lys Ile 330  Pro Ser Cys Asp Ser Ser Pro Val Ser Thr 340   
               
               
                   
               
               
                 Gln Gln Leu Ala Pro Thr Ala Pro Pro Glu 350  Leu Thr Pro Val Val Gln Asp Cys Tyr His 360  Gly Asp 
               
               
                   
               
               
                 Gly Gln Ser Tyr Arg Gly Thr Ser 370  Ser Thr Thr Thr Thr Gly Lys Lys Cys Gln 380  Ser Trp Ser Ser 
               
               
                   
               
               
                 Met Thr Pro His Arg His 390  Gln Lys Thr Pro Gln Asn Tyr Pro Asn Ala 400  Gly Leu Thr Met Asn 
               
               
                   
               
               
                 Tyr Cys Arg Asn Pro 410  Asp Ala Asp Lys Gly Pro Trp Cys Phe Thr 420  Thr Asp Pro Ser Val Arg 
               
               
                   
               
               
                 Trp Gln Tyr Cys 430  Asn Leu Lys Lys Cys Ser Gly Thr Gln Ala 440  Ser Val Val Ala Pro Pro Pro 
               
               
                   
               
               
                 &lt;210&gt; 10 
               
               
                 &lt;212&gt; protein 
               
               
                 &lt;213&gt;  Homo sapiens   
               
               
                 &lt;400&gt; Description of the sequence  
               
               
                 SEQ ID NO: 10 
                   
               
               
                 Lys 78  Val Tyr 80  Leu Ser Glu Cys Lys Thr Gly Asn Gly Lys 90  Asn Tyr Arg Gly Thr Met Ser Lys Thr 
                   
               
               
                   
               
               
                 Lys 100  Asn Gly Ile Thr Cys Gln Lys Trp Ser Ser 110  Thr Ser Pro His Arg Pro Arg Phe Ser Pro 120   
               
               
                   
               
               
                 Ala Thr His Pro Ser Glu Gly Leu Glu Glu 130  Asn Tyr Cys Arg Asn Pro Asp Asn Asp Pro 140  Gln 
               
               
                   
               
               
                 Gly Pro Trp Cys Tyr Thr Thr Asp Pro 150  Glu Lys Arg Tyr Asp Tyr Cys Asp Ile Leu 160  Glu Cys Glu 
               
               
                   
               
               
                 Glu Glu Cys Met His Cys Ser 170  Gly Glu Asn Tyr Asp Gly Lys Ile Ser Lys 180  Thr Met Ser Gly Leu 
               
               
                   
               
               
                 Glu Cys Gln Ala Trp 80  Asp Ser Gln Ser Pro His Ala His Gly Tyr 200  Ile Pro Ser Lys Phe Pro Asn 
               
               
                   
               
               
                 Lys Asn Leu 210  Lys Lys Asn Tyr Cys Arg Asn Pro Asp Arg 220  Glu Leu Arg Pro Trp Cys Phe Thr 
               
               
                   
               
               
                 Thr Asp 230  Pro Asn Lys Arg Trp Glu Leu Cys Asp Ile 240  Pro Arg Cys Thr Thr Pro Pro Pro Ser 
               
               
                   
               
               
                 Ser 250  Gly Pro Thr Tyr Gln Cys Leu Lys Gly Thr 260  Gly Glu Asn Tyr Arg Gly Asn Val Ala Val Thr 
               
               
                   
               
               
                 Val Ser Gly His Thr Cys Gln His Trp 280  Ser Ala Gln Thr Pro His Thr His Asn Arg 290  Thr Pro Glu 
               
               
                   
               
               
                 Asn Phe Pro Cys Lys Asn Leu 300  Asp Glu Asn Tyr Cys Arg Asn Pro Asp Gly 310  Lys Arg Ala Pro 
               
               
                   
               
               
                 Trp Cys His Thr Thr Asn 320  Ser Gln Val Arg Trp Glu Tyr Cys Lys Ile 330  Pro Ser Cys Asp Ser Ser 
               
               
                   
               
               
                 Pro Val Ser Thr 340  Glu Gln Leu Ala Pro Thr Ala Pro Pro Glu 350  Leu Thr Pro Val Val Gln Asp Cys 
               
               
                   
               
               
                 Tyr His 360  Gly Asp Gly Gln Ser Tyr Arg Gly Thr Ser 370  Ser Thr Thr Thr Gly Lys Lys Cys 
               
               
                   
               
               
                 Gln 380  Ser Trp Ser Ser Met Thr Pro His Arg His 390  Gln Lys Thr Pro Glu Asn Tyr Pro Asn Ala 400   
               
               
                   
               
               
                 Gly Leu Thr Met Asn Tyr Cys Arg Asn Pro 410  Asp Ala Asp Lys Gly Pro Trp Cys Phe Thr 420  Thr 
               
               
                   
               
               
                 Asp Pro Ser Val Arg Trp Glu Tyr Cys 430  Asn Leu Lys Lys Cys Ser Gly Thr Glu Ala 440  Ser Val Val 
               
               
                   
               
               
                 Ala Pro Pro Pro 
               
               
                   
               
               
                 &lt;210&gt; 11 
               
               
                 &lt;212&gt; protein 
               
               
                 &lt;213&gt;  Homo sapiens   
               
               
                 &lt;400&gt; Description of the sequence  
               
               
                 SEQ ID NO: 11 
                   
               
               
                 Lys 78  Val Tyr 80  Leu Ser Glu Cys Lys Thr Gly Asn Gly Lys 90  Asn Tyr Arg Gly Thr Met Ser Lys Thr 
                   
               
               
                   
               
               
                 Lys 100  Asn Gly Ile Thr Cys Gln Lys Trp Ser Ser 110  Thr Ser Pro His Arg Pro Arg Phe Ser Pro 120   
               
               
                   
               
               
                 Ala Thr His Pro Ser Glu Gly Leu Glu Glu 130  Asn Tyr Cys Arg Asn Pro Asp Asn Asp Pro 140  Gln 
               
               
                   
               
               
                 Gly Pro Trp Cys Tyr Thr Thr Asp Pro 150  Glu Lys Arg Tyr Asp Tyr Cys Asp Ile Leu 160  Glu Cys Glu 
               
               
                   
               
               
                 Glu Glu Cys Met His Cys Ser 170  Gly Glu Asn Tyr Asp Gly Lys Ile Ser Lys 180  Thr Met Ser Gly Leu 
               
               
                   
               
               
                 Glu Cys Gln Ala Trp 80  Asp Ser Gln Ser Pro His Ala His Gly Tyr 200  Ile Pro Ser Lys Phe Pro Asn 
               
               
                   
               
               
                 Lys Asn Leu 210  Lys Lys Asn Tyr Cys Arg Asn Pro Asp Arg 220  Glu Leu Arg Pro Trp Cys Phe Thr 
               
               
                   
               
               
                 Thr Asp 230  Pro Asn Lys Arg Trp Glu Leu Cys Asp Ile 240  Pro Arg Cys Thr Thr Pro Pro Pro Ser 
               
               
                   
               
               
                 Ser Gly Pro Thr Tyr Gln Cys Leu Lys Gly Thr 260  Gly Glu Asn Tyr Arg Gly Asn Val Ala Val Thr 
               
               
                   
               
               
                 Val Ser Gly His Thr Cys Gln His Trp 280  Ser Ala Gln Thr Pro His Thr His Asn Arg 290  Thr Pro Glu 
               
               
                   
               
               
                 Asn Phe Pro Cys Lys Asn Leu 300  Asp Glu Asn Tyr Cys Arg Asn Pro Asp Gly 310  Lys Arg Ala Pro 
               
               
                   
               
               
                 Trp Cys His Thr Thr Asn 320  Ser Gln Val Arg Trp Glu Tyr Cys Lys Ile 330  Pro Ser Cys Asp Ser Ser 
               
               
                   
               
               
                 Pro Val Ser Thr 340  Glu Gln Leu Ala Pro Thr Ala Pro Pro Glu 350  Leu Thr Pro Val Val Gln Asp Cys 
               
               
                   
               
               
                 Tyr His 360  Gly Asp Gly Gln Ser Tyr Arg Gly Thr Ser 370  Ser Thr Thr Thr Thr Gly Lys Lys Cys 
               
               
                   
               
               
                 Gln 380  Ser Trp Ser Ser Met Thr Pro His Arg His 390  Gln Lys Thr Pro GIU Asn Tyr Pro Asn Ala 400   
               
               
                   
               
               
                 Gly Leu Thr Met Asn Tyr Cys Arg Asn Pro 410  Asp Ala Asp Lys Gly Pro Trp Cys Phe Thr 420  Thr 
               
               
                   
               
               
                 Asp Pro Ser Val Arg Trp Glu Tyr Cys 430  Asn Leu Lys Lys Cys Ser Gly Thr Glu Ala 440  Ser Val Val 
               
               
                   
               
               
                 Ala Pro Pro 
               
               
                   
               
               
                 &lt;210&gt; 12 
               
               
                 &lt;212&gt; protein 
               
               
                 &lt;213&gt;  Homo sapiens   
               
               
                 &lt;400&gt; Description of the sequence  
               
               
                 SEQ ID NO: 12 
                   
               
               
                 Lys 78  Val Tyr 80  Leu Set Glu Cys Lys Thr Gly Asn Gly Lys 90  Asn Tyr Arg Gly Thr Met Ser Lys Thr 
                   
               
               
                   
               
               
                 Lys 100  Asn Gly Ile Thr Cys Gln Lys Trp Ser Ser 110  Thr Ser Pro His Arg Pro Arg Phe Ser Pro 120   
               
               
                   
               
               
                 Ala Thr His Pro Set Glu Gly Leu Glu Glu 130  Asn Tyr Cys Arg Asn Pro Asp Asn Asp Pro 140  Gln 
               
               
                   
               
               
                 Gly Pro Trp Cys Tyr Thr Thr Asp Pro 150  Glu Lys Arg Tyr Asp Tyr Cys Asp Ile Leu 160  Glu Cys Glu 
               
               
                   
               
               
                 Glu Glu Cys Met His Cys Ser 170  Gly Glu Asn Tyr Asp Gly Lys Ile Ser Lys 180  Thr Met Ser Gly Leu 
               
               
                   
               
               
                 Glu Cys Gln Ala Trp 80  Asp Ser Gln Ser Pro His Ala His Gly Tyr 200  Ile Pro Ser Lys Phe Pro Asn 
               
               
                   
               
               
                 Lys Asn Leu 210  Lys Lys Asn Tyr Cys Arg Asn Pro Asp Arg 220  Glu Leu Arg Pro Trp Cys Phe Thr 
               
               
                   
               
               
                 Thr Asp 230  Pro Asn Lys Arg Trp Glu Leu Cys Asp Ile 240  Pro Arg Cys Thr Thr Pro Pro Pro Ser 
               
               
                   
               
               
                 Ser 250  Gly Pro Thr Tyr Gln Cys Leu Lys Gly Thr 260  Gly Glu Asn Tyr Arg Gly Asn Val Ala Val Thr 
               
               
                   
               
               
                 Val Ser Gly His Thr Cys Gln His Trp 280  Ser Ala Gln Thr Pro His Thr His Asn Arg 290  Thr Pro Glu 
               
               
                   
               
               
                 Asn Phe Pro Cys Lys Asn Leu 300  Asp Glu Asn Tyr Cys Arg Asn Pro Asp Gly 310  Lys Arg Ala Pro 
               
               
                   
               
               
                 Trp Cys His Thr Thr Asn 320  Set Gln Val Arg Trp Glu Tyr Cys Lys Ile 330  Pro Ser Cys Asp Ser Ser 
               
               
                   
               
               
                 Pro Val Ser Thr 340  Glu Gln Leu Ala Pro Thr Ala Pro Pro Glu 350  Leu Thr Pro Val Val Gln Asp Cys 
               
               
                   
               
               
                 Tyr His 360  Gly Asp Gly Gln Ser Tyr Arg Gly Thr Ser 370  Ser Thr Thr Thr Thr Gly Lys Lys Cys 
               
               
                   
               
               
                 Gln 380  Ser Trp Ser Ser Met Thr Pro His Arg His 390  Gln Lys Thr Pro Glu Asn Tyr Pro Asn Ala 400   
               
               
                   
               
               
                 Gly Leu Thr Met Asn Tyr Cys Arg Asn Pro 410  Asp Ala Asp Lys Gly Pro Trp Cys Phe Thr 420  Thr 
               
               
                   
               
               
                 Asp Pro Ser Val Arg Trp Glu Tyr Cys 430  Asn Leu Lys Lys Cys Set Gly Thr Glu Ala 440  Ser Val Val 
               
               
                   
               
               
                 Ala Pro Pro Pro Val Val Leu 450  Leu Pro Asp Val Glu Thr Pro Ser Glu Glu 460  Asp Cys Met Phe 
               
               
                   
               
               
                 Gly Asn Gly Lys 
               
               
                   
               
               
                 &lt;210&gt; 13 
               
               
                 &lt;212&gt; protein 
               
               
                 &lt;213&gt;  Homo sapiens   
               
               
                 &lt;400&gt; Description of the sequence  
               
               
                 SEQ ID NO: 13 
                   
               
               
                 Lys 78  Val Tyr 80  Leu Ser Glu Cys Lys Thr Gly Asn Gly Lys 90  Asn Tyr Arg Gly Thr Met Ser Lys Thr 
                   
               
               
                   
               
               
                 Ly s100  Asn Gly Ile Thr Cys Gln Lys Trp Ser Ser 110  Thr Ser Pro His Arg Pro Arg Phe Ser Pro 120   
               
               
                   
               
               
                 Ala Thr His Pro Ser Glu Gly Leu Glu Glu 130  Asn Tyr Cys Arg Asn Pro Asp Asn Asp Pro 140  Gln 
               
               
                   
               
               
                 Gly Pro Trp Cys Tyr Thr Thr Asp Pro 150  Glu Lys Arg Tyr Asp Tyr Cys Asp Ile Leu 160  Glu Cys Glu 
               
               
                   
               
               
                 Glu Glu Cys Met His Cys Ser 170  Gly Glu Asn Tyr Asp Gly Lys Ile Ser Lys 180  Thr Met Ser Gly Leu 
               
               
                   
               
               
                 Glu Cys Gln Ala Trp 80  Asp Ser Gln Ser Pro His Ala His Gly Tyr 200  Ile Pro Ser Lys Phe Pro Asn 
               
               
                   
               
               
                 Lys Asn Leu 210  Lys Lys Asn Tyr Cys Arg Asn Pro Asp Arg 220  Glu Leu Arg Pro Trp Cys Phe Thr 
               
               
                   
               
               
                 Thr Asp 230  Pro Asn Lys Arg Trp Glu Leu Cys Asp Ile 240  Pro Arg Cys Thr Thr Pro Pro Pro Ser 
               
               
                   
               
               
                 Ser 250  Gly Pro Thr Tyr Gln Cys Leu Lys Gly Thr 260  Gly Glu Asn Tyr Arg Gly Asn Val Ala Val Thr 
               
               
                   
               
               
                 Val Ser Gly His Thr Cys Gln His Tr p280  Ser Ala Gln Thr Pro His Thr His Asn Arg 290  Thr Pro Glu 
               
               
                   
               
               
                 Asn Phe Pro Cys Lys Asn Leu 300  Asp Glu Asn Tyr Cys Arg Asn Pro Asp Gly 310  Lys Arg Ala Pro 
               
               
                   
               
               
                 Trp Cys His Thr Thr Asn 320  Ser Gln Val Arg Trp Glu Tyr Cys Lys Ile 330  Pro Ser Cys Asp Ser Ser 
               
               
                   
               
               
                 Pro Val Ser Thr 340  Glu Gln Leu Ala Pro Thr Ala Pro Pro Glu 350  Leu Thr Pro Val Val Gln Asp Cys 
               
               
                   
               
               
                 Tyr His 360  Gly Asp Gly Gln Ser Tyr Arg Gly Thr Ser 370  Ser Thr Thr Thr Thr Gly Lys Lys Cys 
               
               
                   
               
               
                 Gln 380  Ser Trp Ser Ser Met Thr Pro His Arg His 390  Gln Lys Thr Pro Glu Asn Tyr Pro Asn Ale 400   
               
               
                   
               
               
                 Gly Leu Thr Met Asn Tyr Cys Arg Asn Pro 410  Asp Ala Asp Lys Gly Pro Trp Cys Phe Thr 420  Thr 
               
               
                   
               
               
                 Asp Pro Ser Val Arg Trp Glu Tyr Cys 430  Asn Leu Lys Lys Cys Ser Gly Thr Glu Ala 440  Ser Val Val 
               
               
                   
               
               
                 Ala Pro Pro Pro Val Val Leu 450  Leu Pro Asp Val Glu Thr Pro Ser Glu Glu 460  Asp Cys Met Phe 
               
               
                   
               
               
                 Gly Asn Gly Lys Gly Tyr 470  Arg Gly Lys Arg Ala Thr Thr Val Thr Gly 480  Thr Pro Cys Gln Asp Trp 
               
               
                   
               
               
                 Ala Ala Gln Glu 490  Pro His Arg His Ser Ile Phe Thr Pro Glu 500  Thr Asn Pro Arg Ala Gly Leu Glu 
               
               
                   
               
               
                 Lys Asn 510  Tyr Cys Arg Asn Pro Asp Gly Asp Val Gly 520  Gly Pro Trp Cys Tyr Thr Thr Asn Pro 
               
               
                   
               
               
                 Arg 530   
               
               
                   
               
               
                 &lt;210&gt; 14 
               
               
                 &lt;212&gt; protein 
               
               
                 &lt;213&gt;  Homo sapiens   
               
               
                 &lt;400&gt; Description of the sequence  
               
               
                 SEQ ID NO: 14 
                   
               
               
                 Val Gln Asp Cys Tyr His 360  Gly Asp Gly Gln Ser Tyr Arg Gly Thr Ser 370  Ser Thr Thr Thr Thr Gly 
                   
               
               
                   
               
               
                 Lys Lys Cys Gln 380  Ser Trp Ser Ser Met Thr Pro His Arg His 390  Gln Lys Thr Pro Glu Asn Tyr Pro 
               
               
                   
               
               
                 Asn Ala 400  Gly Leu Thr Met Asn Tyr Cys Arg Asn Pro 410  Asp Ala Asp Lys Gly Pro Trp Cys Phe 
               
               
                   
               
               
                 Thr 420  Thr Asp Pro Ser Val Arg Trp Glu Tyr Cys 430  Asn Leu Lys Lys Cys Ser Gly Thr Glu Ala 440   
               
               
                   
               
               
                 Ser Val Val Ala Pro Pro Pro Val Val Leu 450  Leu Pro Asp Val Glu Thr Pro Ser Glu Glu 460  Asp Cys 
               
               
                   
               
               
                 Met Phe Gly Asn Gly Lys Gly Tyr 470  Arg Gly Lys Arg Ala Thr Thr Val Thr Gly 480  Thr Pro Cys Gln 
               
               
                   
               
               
                 Asp Trp Ala Ala Gln Glu 490  Pro His Arg His Ser Ile Phe Thr Pro Glu 500  Thr Asn Pro Arg Ala Gly 
               
               
                   
               
               
                 Leu Glu Lys Asn 510  Tyr Cys Arg Asn Pro Asp Gly Asp Val Gly 520  Gly Pro Trp Cys Tyr Thr Thr 
               
               
                   
               
               
                 Asn Pro Arg 530  Lys Leu Tyr Asp Tyr Cys Asp Val Pro Gln 540  Cys Ala Ala Pro Ser Phe 
               
               
                   
               
               
                 &lt;210&gt; 15 
               
               
                 &lt;212&gt; protein 
               
               
                 &lt;213&gt;  Homo sapiens   
               
               
                 &lt;400&gt; Description of the sequence  
               
               
                 SEQ ID NO: 15 
                   
               
               
                 Tyr 80  Leu Ser Glu Cys Lys Thr Gly Asn Gly Lys 90  Asn Tyr Arg Gly Thr Met Ser Lys Thr Lys 100   
                   
               
               
                   
               
               
                 Asn Gly Ile Thr Cys Gln Lys Trp Ser Ser 110  Thr Ser Pro His Arg Pro Arg Phe Ser Pro 120  Ala Thr 
               
               
                   
               
               
                 His Pro Ser Glu Gly Leu Glu Glu 130  Asn Tyr Cys Arg Asn Pro Asp Asn Asp Pro 140  Gln Gly Pro 
               
               
                   
               
               
                 Trp Cys Tyr Thr Thr Asp Pro 150  Glu Lys Arg Tyr Asp Tyr Cys Asp Ile Leu 160  Glu Cys Glu Glu 
               
               
                   
               
               
                 &lt;210&gt; 16 
               
               
                 &lt;212&gt; protein 
               
               
                 &lt;213&gt;  Homo sapiens   
               
               
                 &lt;400&gt; Description of the sequence  
               
               
                 SEQ ID NO: 16 
                   
               
               
                 Cys Met His Cys Ser 170  Gly Glu Asn Tyr Asp Gly Lys Ile Ser Lys 180  Thr Met Ser Gly Leu Glu Cys 
                   
               
               
                   
               
               
                 Gln Ala Trp 80  Asp Ser Gln Ser Pro His Ala His Gly Tyr 200  Ile Pro Ser Lys Phe Pro Asn Lys Asn 
               
               
                   
               
               
                 Leu Lys Lys Asn Tyr Cys Arg Asn Pro Asp Arg 220  Glu Leu Arg Pro Trp Cys Phe Thr Thr 
               
               
                   
               
               
                 Asp 230  Pro Asn Lys Arg Trp Glu Leu Cys Asp Ile 240  Pro Arg Cys Thr Thr Pro Pro Pro Ser Ser 250   
               
               
                   
               
               
                 Gly Pro Thr Tyr Gln Cys Leu Lys Gly Thr 260  Gly Glu Asn Tyr Arg Gly Asn Val Ala Val Thr Val 
               
               
                   
               
               
                 Ser Gly His Thr Cys Gln His Trp 280  Ser Ala Gln Thr Pro His Thr His Asn Arg 290  Thr Pro Glu Asn 
               
               
                   
               
               
                 Phe Pro Cys Lys Asn Leu 300  Asp Glu Asn Tyr Cys Arg Asn Pro Asp Gly 310  Lys Arg Ala Pro Trp 
               
               
                   
               
               
                 Cys His Thr Thr Asn 320  Ser Gln Val Arg Trp Glu Tyr Cys Lys Ile 330  Pro Ser Cys Asp Ser Ser Pro 
               
               
                   
               
               
                 Val 
               
               
                   
               
               
                 &lt;210&gt; 17 
               
               
                 &lt;212&gt; protein 
               
               
                 &lt;213&gt;  Homo sapiens   
               
               
                 &lt;400&gt; Description of the sequence  
               
               
                 SEQ ID NO: 17 
                   
               
               
                 Val Gln Asp Cys Tyr His 360  Gly Asp Gly Gln Ser Tyr Arg Gly Thr Ser 370  Ser Thr Thr Thr Thr Gly 
                   
               
               
                   
               
               
                 Lys Lys Cys Gln 380  Ser Trp Ser Ser Met Thr Pro His Arg His 390  Gln Lys Thr Pro Glu Asn Tyr Pro 
               
               
                   
               
               
                 Asn Ala 400  Gly Leu Thr Met Asn Tyr Cys Arg Asn Pro 410  Asp Ala Asp Lys Gly Pro Trp Cys Phe 
               
               
                   
               
               
                 Thr 420  Thr Asp Pro Ser Val Arg Trp Glu Tyr Cys 430  Asn Leu Lys Lys Cys Ser Gly Thr Glu Ala 440   
               
               
                   
               
               
                 &lt;210&gt; 18 
               
               
                 &lt;212&gt; protein 
               
               
                 &lt;213&gt;  Homo sapiens   
               
               
                 &lt;400&gt; Description of the sequence  
               
               
                 SEQ ID NO: 18 
                   
               
               
                 Ser Val Val Ala Pro Pro Pro Val Val Leu 450  Leu Pro Asp Val Glu Thr Pro Ser Glu Glu 460  Asp Cys 
                   
               
               
                   
               
               
                 Met Phe Gly Asn Gly Lys Gly Tyr 470  Arg Gly Lys Arg Ala Thr Thr Val Thr Gly 480  Thr Pro Cys Gln 
               
               
                   
               
               
                 Asp Trp Ala Ala Gln Glu 490  Pro His Arg His Ser Ile Phe Thr Pro Glu 500  Thr Asn Pro Arg Ala Gly 
               
               
                   
               
               
                 Leu Glu Lys Asn 510  Tyr Cys Arg Asn Pro Asp Gly Asp Val Gly 520  Gly Pro Trp Cys Tyr Thr Thr 
               
               
                   
               
               
                 Asn Pro Arg 530  Lys Leu Tyr Asp Tyr Cys Asp Val Pro Gln 540  Cys Ala Ala Pro Ser Phe 
               
               
                   
               
               
                 &lt;210&gt; 19 
               
               
                 &lt;212&gt; protein 
               
               
                 &lt;213&gt;  Homo sapiens   
               
               
                 &lt;400&gt; Description of the sequence  
               
               
                 SEQ ID NO: 19 
                   
               
               
                 Val Val Ala Pro Pro Pro Val Val Leu 450  Leu Pro Asp Val Glu Thr Pro Ser Glu Glu 460  Asp Cys 
                   
               
               
                   
               
               
                 Met Phe Gly Asn Gly Lys Gly Tyr 470  Arg Gly Lys Arg Ala Thr Thr Val Thr Gly 480  Thr Pro Cys Gln 
               
               
                   
               
               
                 Asp Trp Ala Ala Gln Glu 490  Pro His Arg His Ser Ile Phe Thr Pro Glu 500  Thr Asn Pro Arg Ala Gly 
               
               
                   
               
               
                 Leu Glu Lys Asn 510  Tyr Cys Arg Asn Pro Asp Gly Asp Val Gly 520  Gly Pro Trp Cys Tyr Thr Thr 
               
               
                   
               
               
                 Asn Pro Arg 530  Lys Leu Tyr Asp Tyr Cys Asp Val Pro Gln 540  Cys Ala Ala Pro Ser Phe Asp Cys 
               
               
                   
               
               
                 Gly Lys 550  Pro Gln Val Glu Pro Lys Lys Cys Pro Gly 560  Arg 
               
               
                   
               
               
                 &lt;210&gt; 20 
               
               
                 &lt;212&gt; 6enoκ 
               
               
                 &lt;213&gt;  Homo sapiens   
               
               
                 &lt;400&gt; Description of the sequence  
               
               
                 SEQ ID NO: 20 
                   
               
               
                 Val Val Ala Pro Pro Pro Val Val Leu 450  Leu Pro Asp Val Glu Thr Pro Ser Glu Glu 460  Asp Cys 
                   
               
               
                   
               
               
                 Met Phe Gly Asn Gly Lys Gly Tyr 470  Arg Gly Lys Arg Ala Thr Thr Val Thr Gly 480  Thr Pro Cys Gln 
               
               
                   
               
               
                 Asp Trp Ala Ala Gln Glu 490  Pro His Arg His Ser Ile Phe Thr Pro Glu 500  Thr Asn Pro Arg Ala Gly 
               
               
                   
               
               
                 Leu Glu Lys Asn 510  Tyr Cys Arg Asn Pro Asp Gly Asp Val Gly 520  Gly Pro Trp Cys Tyr Thr Thr 
               
               
                   
               
               
                 Asn Pro Arg 530  Lys Leu Tyr Asp Tyr Cys Asp Val Pro Gln 540  Cys Ala Ala Pro Ser Phe Asp Cys 
               
               
                   
               
               
                 Gly Lys 550  Pro Gln Val Glu Pro Lys Lys Cys Pro Gly 560  Arg Val Val Gly Gly Cys Val Ala His 
               
               
                   
               
               
                 Pro 570  His Ser Trp Pro Trp Gln Val Ser Leu Arg 580  Thr Arg Phe Gly Met His Phe Cys Gly Gly 590   
               
               
                   
               
               
                 Thr Leu Ile Ser Pro Glu Trp Val Leu Thr 600  Ala Ala His Cys Leu Glu Lys Ser Pro Arg 610  Pro Ser 
               
               
                   
               
               
                 Ser Tyr Lys Val Ile Leu Gly Ala 620  His Gln Glu Val Asn Leu Glu Pro His Val 630  Gln Glu Ile Glu 
               
               
                   
               
               
                 Val Ser Arg Leu Phe Leu 640  Glu Pro Thr Arg Lys Asp Ile Ala Leu Leu 650  Lys Leu Ser Ser Pro Ala 
               
               
                   
               
               
                 Val Ile Thr Asp 660  Lys Val Ile Pro Ala Cys Leu Pro Ser Pro 670   Asn Tyr Val Val Ala Asp Arg Thr 
               
               
                   
               
               
                 Glu Cys 680  Phe Ile Thr Gly Trp Gly Glu Thr Gln Gly 690  Thr Phe Gly Ala Gly Leu Leu Lys Glu 
               
               
                   
               
               
                 Ala 700  Gln Leu Pro Val Ile Glu Asn Lys Val Cys 710  Asn Arg Tyr Glu Phe Leu Asn Gly Arg Va 720 l 
               
               
                   
               
               
                 Gln Ser Thr Glu Leu Cys Ala Gly His Leu 730  Ala Gly Gly Thr Asp Ser Cys Gln Gly Asp 740  Ser 
               
               
                   
               
               
                 Gly Gly Pro Leu Val Cys Phe Glu Lys 750  Asp Lys Tyr He Leu Gln Gly Val Thr Ser 760  Trp Gly Leu 
               
               
                   
               
               
                 Gly Cys Ala Arg Pro Asn Lys 770  Pro Gly Val Tyr Val Arg Val Ser Arg Phe 780  Val Thr Trp Ile Glu 
               
               
                   
               
               
                 Gly Val Met Arg Asn 790  Asn 
               
             
          
         
       
     
       INDUSTRIAL APPLICABILITY 
       [0151]    The present invention is embodied with multipurpose equipment extensively employed by the industry.