Title of Invention

MONOCLONAL ANTIBODY TO LOOP 9 IN HUMAN GPVI DOMAIN 2

Abstract

The present invention provides an antibody which has the following features, its active fragment, or a derivative thereof: a) It specifically binds to human platelet membrane glycoprotein VI (GPVI); b)The function to activate a platelet and/or the function to induce a thrombocytopenia in vivo are low; and c) It at least partially depletes GPVI on the platelet membrane by contacting with a platelet.

Full Text

FORM 2 THE PATENT ACT 1970 (39 of 1970) The Patents Rules, 2003 COMPLETE SPECIFICATION (See Section 10, and rule 13) 1. TITLE OF INVENTION ANTI-PLATELET MEMBRANE GLYCOPROTEIN VI MONOCLONAL ANTIBODY 2. APPLICANT(S) a) Name b) Nationality c) Address MOCHIDA PHARMACEUTICAL CO., LTD, JAPANESE Company 7, YOTSUYA 1-CHOME, SHINJUKU-KU, TOKYO 160-8515 JAPAN 3. PREAMBLE TO THE DESCRIPTION The following specification particularly describes the invention and the manner in which it is to be performed : - ENGLISH TRANSLATION VARIFICATION CERTIFICATE u/r. 20(3)(b) I, Mr. HIRAL CHANDRAKANT JOSHI, an authorized agent for the applicant, MOCHIDA PHARMACEUTICAL CO., LTD. do hereby verify that the content of English translated complete specification filed in pursuance of PCT International application No. PCT/JP2006/ 309431 thereof is correct and complete. HIRAL CHANDRAKANT JOSHI AGENT FOR MOCHIDA PHARMACEUTICAL CO., LTD. DESCRIPTION TECHNICAL FIELD The present invention relates to an antibody to platelet membrane glycoprotein VI (hereinafter, sometimes abbreviated as GPVI) and a recognition region of the antibody. BACKGROUND ART The platelet plays a very important role in the clot formation and the biophylaxis, and its concerning in various clinical conditions is being elucidated from the physiological role. In particular, it is remarkable about the function that the platelet forms a hemostatic plug. For example, when the vascular endothelial cell suffers damage, the collagen that is the major matrix protein of the subcutaneous vascular endothelium is exposed and the platelet adheres thereto. Next, the platelet is activated by the signal from the collagen and when finally, the platelet agglutinates through the fibrinogen. Then, since this fact causes morbidity such as thromboembolic disease depending on the situation, it is remarkable as a target for therapy. In the past, for the purpose of the treatment and the prevention of the thrombosis based on the platelet aggregation, an anti-platelet agent such as aspirin, ticlopidine, GPIIb/IIIa antagonist and the like has been used. However, a lot of problems are pointed out from the aspect of the effectiveness and the side effects such as bleeding. Therefore, the excellent platelet inhibitor having the enough safety and / the sure and appropriate function without the above-mentioned problems is desired to arrive. GPVI that is present on the platelet membrane is the collagen receptor of the platelet, and it has been elucidated that the GPVI plays a central role for an activation of the platelet by collagen stimulation (see, Hiroshi Takayama, The Japanese Journal of Thrombosis and Hemostasis, 2003, volume 14, No. 2, pp. 75-81). That is, Sugiyama et al. has been reported that the membrane protein of 62kDa is specifically depleted in the platelet of a patient with autoimmune thrombocytopenia and the platelet aggregation by collagen cannot be detected (see, Tateo Sugiyama and five other members, Blood (USA), 1987, volume 69, No. 6, pp. 1712-1720), and further that the protein that has been deleted in the platelet of the patient is GPVI and the Fab fragment of the antibody purified from the serum of the patient suppresses a collagen-induced platelet aggregation (see, Tateo Sugiyama and five other members, Blood (USA), 1987, volume 69, No. 6, pp. 1712-1720, and Masaaki Moroi and three other members, The Journal of Clinical Investigation (USA), 1989, volume 84, No. 5, pp. 1440-1445). So far, Sugiyama et al. (see, Tateo Sugiyama and five other members, Blood (USA), 1987, volume 69, No. 6, pp. 1712-1720) and Takahashi et al. (see, Hoyu Takahashi and one other member, American Journal of Hematology (USA), 2001, volume 67, No. 4, pp. 262-267) have been reported about the anti-human GPVI autoantibody 2 derived from the patient with autoimmune disease. However, since, according to the report by Sugiyama et al., the anti-human GPVI autoantibody purified from the patient's plasma has a function to induce platelet aggregation, it cannot be applied for medicaments immediately. Takahashi et al. (Hoyu Takahashi and one other member, American Journal of Hematology (USA), 2001, volume 67, No. 4, pp. 262-267) describes that an autoantibody to the protein of ca. 62kDa that is speculated to be GPVI is present, and that this antibody induces a platelet aggregation. In addition, to apply these anti-GPVI antibodies derived from the patient clinically as a medicament, the antibody with high safety must be produced in quantities in stable quality. However, the method of producing industrially is not yet established. The anti-GPVI antibody, which has been prepared by the present, includes a monoclonal rat antibody to mouse GPVI (see, Publication number 1228768 of the European Patent Application) and a monoclonal mouse antibody to human GPVI (see, Publication number 01/00810 of International Patent Application and Publication number 02/080968 of International Patent Application, Thromb Haemost. 2003 Jun; 89(6): 996-1003). In addition, human single-stranded antibody (scFv: single chain Fv) that recognizes human GPVI has been prepared using the phage display method (see, Publication number 01/00810 of International Patent Application, Publication number 02/080968 of International Patent Application, and Peter A Smethurst and 15 other members, Blood (USA), 2002, volume 100, number 11, p. 474a). These single-stranded antibodies are the antibody that combines heavy chain variable region (VH) and light chain variable region (VL) of the human antibody by the peptide linker and has a variable region derived from the human. However, compared with normal immunoglobulin that the cell produces, it generally possesses a low affinity to an antigen thereof and is short in the half-Ufe in vivo, too. Further, Smethurst analyzed an epitope on GPVI in the term of clone 10B12 that suppresses platelet aggregation among the single-stranded antibodies, and suggested that lysine at the 59th position (Lys59) has a possibility to involve ( Hoyu Takahashi and one other member, American Journal of Hematology (USA), 2001, volume 67, No. 4, pp. 262-267; and Peter A Smethurst and 15 other members, Blood (USA), 2002, volume 100, number 11, p. 474a). As described above, most of the antibodies to human GPVI that have been reported until now, including the aforementioned human auto antibody, possess an activity of activating the platelet only by the antibody in vitro, and/or an activity of inducing or enhancing platelet aggregation. Thus, when administering the same in vivo, a probability to cause thrombocytopenia is considered. In fact, Nieswandt et al. has reported several monoclonal antibodies (JAQ1, JAQ2 and JAQ3) that deplete GPVI on the platelet in vivo. All antibodies caused thrombocytopenia after administration. Recently, there have been reported that collagen, the agonist for GPVI, convulxin and CRP, and an antibody (9012.2) that inhibits the binding of collagen to GPVI 3 activate the platelet, followed by, a shedding of GPVI from the platelet occurs by metalloprotease-mediated cleavage (Stephens G and four other members, Blood, 2005, 105(1): 186-191; Gardiner EE and four other members, Blood, 2004, 104: 3611-3617; Bergmeier W and six other members, Thromb Haemost, 2004, 91: 951-958). Further, the prediction of the amino acid residues on GPVI associated with the interaction with collagen (Val34, Leu36) was performed using the 9012.2 antibody, etc. (Lecut C and seven other members, J Biol Chem, 2004, 279: 52293-52299). In addition, Takayama et al. cloned anti-human GPVI antibodies using lymphocytes from the patient with autoimmune disease, and investigated about the feature of the antibodies in vitro ( Publication number 05/007800 of International Patent Application). However, in all reports that have been published until now, no antibody having the activity to deplete GPVI on the platelet membrane without activating the platelet and/or without inducing thrombocytopenia in vivo has been disclosed. DISCLOSURE OF THE INVENTION In the context of requirement for a medicament that has high safety and excellent efficacy, and is easy-to-use as an anti-platelet agent as described above, an anti-GPVI antibody applicable in vivo is desired. The present invention intends to provide a novel antibody that specifically binds to GPVI, which is the glycoprotein that exists on a platelet such as a mammalian platelet, specifically a platelet from human, monkey, rat or mouse, especially the human platelet membrane, preferably a monoclonal antibody. Particularly, there is provided the anti-GPVI antibody that is applicable in vivo, has efficacy and no problem in the aspect of side effects such as thrombocytopenia. Also, the invention provides the antibody that specifically binds to GPVI, especially human GPVI and comprises a novel CDR sequence. Moreover, the cell that produces these antibodies is provided. To solve the above-mentioned problems, the inventors have conceived of establishing a lot of mouse hybridomas that produce an antibody to GPVI and analyzing properties of the antibodies produced by the hybridomas. Based on the conception, the present inventors have made extensive investigations and have succeeded to obtain the hybridomas that produce the antibody having a binding ability to GPVI and an activity for decreasing the collagen-induced platelet aggregation. Then the inventors further analyzed recognition region on GPVI of each antibody to obtain useful information about epitope of GPVI. As a result of further investigation after isolating the clones, the inventors succeeded in obtaining a gene encoding the antibody. In addition, it was found that the amino acid sequence of CDR of the antibody is a novel one. Moreover, the inventors have completed the present invention by preparing recombinant antibodies using gene recombination technology. 4 In addition, throughout the specification, an antibody that is produced by a hybridoma, e.g. clone F1232-18 is sometimes referred to as F1232-18 antibody. The first embodiment of the present invention is an antibody which exhibits a specific function or feature, and specifically binds to GPVI such as a mammalian GPVI, specifically GPVI from human, monkey, rat or mouse, especially the human GPVI, preferably a monoclonal antibody (hereinafter, sometimes referred to as anti-human GPVI antibody and anti-human GPVI monoclonal antibody, respectively), its active fragment, or derivatives thereof. Specifically, it includes as follows. (1) An antibody having the following features, its active fragment, or derivatives thereof: a) It specifically binds to GPVI, especially human platelet membrane glycoprotein VI (GPVI); b) The action to activate a platelet and/ or the action to induce a thrombocytopenia in vivo are weak; and c) It at least partially depletes GPVI on the platelet membrane by contacting with a platelet. (2) An anti-GPVI antibody, its active fragment, or derivatives thereof which at least partially depletes GPVI on the platelet membrane by contacting with a platelet without shedding of platelet GPVI, especially, shedding of platelet GPVI by methalloprotease-mediated cleavage accompanying platelet activation, especially an antibody having the features of (1) above, its active fragment, or derivatives thereof. (3) An anti-GPVI antibody, its active fragment, or derivatives thereof which at least partially depletes GPVI on the platelet membrane via internalization of the platelet GPVI (4) The antibody of (1) to (3) , its active fragment, or derivatives thereof which at least partially depletes GPVI on the platelet membrane via internalization of the platelet GPVI. (5) The antibody of (1) to (4), its active fragment or derivatives thereof, which at least partially depletes GPVI on the platelet membrane by contacting with a platelet in vivo. (6) The antibody of (1) to (5), its active fragment or derivatives thereof, which decreases or deletes an ability of platelet to aggregate responsive to collagen by contacting with a platelet by administered in vivo. (7) The antibody of (1) to (6), its active fragment, or derivatives thereof, wherein its action to prolong bleeding time is weak. 5 (8) The antibody of (1) to (7), its active fragment or derivatives thereof, wherein the dissociation constant with GPVI is equal to or less than 4 x lf>8 M. The antibody of the aforementioned (1) to (8) is preferably an antibody which does not induce human platelet aggregation solely. Adequate examples of the antibody include antibody clones listed in Tables 6 and 11, preferably an antibody that recognizes loop 9 of GPVI, or a chimeric antibody or a humanized antibody, wherein the above antibody is recombined with human IgG, more preferably human IgG4. Further, the antibody of the present invention is an antibody, wherein the dissociation constant (Kd value) between GPVI, especially human GPVI and the antibody is preferably equal to or less than 1(H M, more preferably equal to or less than 4 x 10~9 M. The antibody, its active fragment or derivatives thereof of the present invention encompass, as long as they have a binding ability to GPVI, for example, a chimeric antibody and a humanized antibody, Fab (Fragment of antigen binding), Fab', F(ab')2, a single-chain antibody (scFv), a disulfide stabilized antibody (dsFv), diabody, nanobody and a peptide comprising CDR, and a labeled antibody, a conjugated antibody and a antibody-fused protein, etc. Moreover, the antibody of the first embodiment of the present invention, or the like, is preferably an antibody which specifically binds to GPVI such as a mammalian GPVI, specifically GPVI from human, monkey, rat or mouse, especially the human GPVI and specifically decreases platelet aggregability against collagen, but has no effect on the aggregability against other agonists, e.g. ADP or thrombin. Preferably, the antibody cannot induce human platelet aggregation solely. The antibody cannot significantly induce human platelet aggregation solely in the concentration or the dosage equivalent to those, preferably 10-fold, more preferably 100-fold, further preferably 1000-fold of those, in which it suppresses collagen-induced human platelet aggregation. Herein, the antibody of the aforementioned (1) to (8) may be an antibody which inhibits the binding of GPVI, especially human GPVI to collagen as long as it has its properties, preferably an antibody which inhibits the binding of GPVI, especially human GPVI to collagen with the dissociation constant (Kd) equal to or less than 10-8 M, more preferably equal to or less than 10'9 M, further preferably equal to or less than lO-io M. The antibody of the present invention is not necessarily limited to the specific clone, and the antibody having a similar function to that of the preferred examples of the present invention (or antibody, etc.) is encompassed within the scope of the present invention. The existence or non-existence of the function of the antibody of the present invention can be confirmed by the method shown in Examples or the publicly known method. In addition, an antibody, wherein its recognition region, binding site or epitope on GPVI is the same or at least partially common to those of the preferred antibody of the present invention, for example, an antibody which competes with each other for binding with GPVI, is included within the scope of the present invention. Whether 6 an antibody has a commonality in recognition region or binding site with the antibody of the present invention or not can be confirmed according to the method described in EXAMPLES or by publicly known methods. That is, the present invention provides an antibody which competes with a specified antibody of the present invention for GPVI binding. In the classification based on the competition experiment in EXAMPLES of the present invention, antibodies classified into eight groups in Table 1, preferably groups d, e or h, more preferably group d or e are exemplified as the antibody of the present invention. Preferred examples of the antibody of (1) to (8) aforementioned include an antibody that recognizes at least a part of loop 9 of GPVI, especially human GPVI. The second embodiment of the present invention is an anti-GPVI antibody which is defined by novel recognition region, binding site or epitope on GPVI such as mammalian GPVI, specifically GPVI from human, monkey, rat or mouse, especially human GPVI, preferably a monoclonal antibody. Specifically, it includes: (9) An antibody, its active fragment or derivatives thereof, which spcifically recognizes an amino acid sequence or a structure on the GPVI comprising at least a part of loop 2, loop 3 and loop 5, or loop 4 and loop 5 in GPVI, especially human GPVI domain 1, or loop 9, or loop 9 and loop 11 in domain 2, preferably loop 9, or loop 9 and loop 11 in domain 2 or loop 2 in domain 1, more preferably loop 9, or loop 9 and loop 11 in domain 2, further preferably loop 9 in domain 2; (10) The antibody of (9), its active fragment or derivatives thereof, wherein at least a part of loop 2, loop 3 and loop 5, or loop 4 and loop 5 in GPVI domain 1, or loop 9, or loop 9 and loop 11 in domain 2 are E21, K22 and P23 of loop 2 of the human GPVI, G33 of loop 3 and A57, K59 and L62 of loop 5, or S43, S44, S45, R46, and E48 of loop 4 and A57, K59 and L62 of loop 5, or T116, R117, G119 and Q122 of loop 9 or T116, R117, G119, and Q122 of loop 9 and R139 of loop 11; (11) The antibody of (9) or (10), its active fragment or derivatives thereof, which specifically binds to loop 2, loop 3 and loop 5, or loop 4 and loop 5 in GPVI, especially human GPVI domain 1, or loop 9, or loop 9 and loop 11 in domain 2, preferably loop 9, or loop 9 and loop 11 in domain 2 or loop 2 in domain 1, more preferably loop 9, or loop 9 and loop 11 in domain 2, further preferably loop 9 in domain 2; more preferably, (12) The antibody of (1) to (8), its active fragment or derivatives thereof, which recognizes an amino acid sequence or a structure on the GPVI comprising at least a part of loop 2, loop 3 and loop 5, or loop 4 and loop 5 in GPVI, especially human GPVI domain 1, or loop 9, or loop 9 and loop 11 in domain 2, preferably loop 9, or loop 9 and loop 11 in domain 2 or loop 2 in domain 1, more preferably loop 9, or loop 9 and loop 11 in domain 2, further preferably loop 9 in domain 2; (13) The antibody of (1) to (8), its active fragment or derivatives thereof, which specifically recognizes an amino acid sequence or a structure on the GPVI comprising E21, K22 and P23 of loop 2, G33 of loop 3 and A57, K59 and L62 of loop 5, or S43, S44, S45, R46, and E48 of loop 4 and A57, K59 and L62 of loop 5, or T116, 7 R117, G119 and Q122 of loop 9 or T116, R117, G119, and Q122 of loop 9 and R139 of loop 11; and (14) The antibody of (1) to (8), its active fragment or derivatives thereof, which specifically binds to loop 2, loop 3 and loop 5, or loop 4 and loop 5 in GPVI, especially human GPVI domain 1, or loop 9, or loop 9 and loop 11 in domain 2, preferably loop 9, or loop 9 and loop 11 in domain 2 or loop 2 in domain 1, more preferably loop 9, or loop 9 and loop 11 in domain 2, further preferably loop 9 in domain 2. Herein, at least a part of each loop described above is, for example, the residues, which are different from the corresponding amino acid residues of heterogeneous GPVI such as monkey, mouse or rat GPVI. A modeling structure of GPVI such as human GPVI is presumable by the method described in EXAMPLES. A position of each loop structure is shown in FIGs. 1, 3 and 47. Among the abovementioned loops, loop 9, or loop 9 and loop 11 in domain 2 and loop 2 in domain 1, preferably loop 9, loop 9 and loop 11 in domain 2, further preferably loop 9 in domain 2 are important as a recognition region of the antibody of the present invention, and the antibody which can recognize the region is preferred. Preferred examples are an antibody listed in Tables 6 and 11, or a chimeric or humanized antibody which is recombined with human IgG, more preferably human IgG4. The antibody of the second embodiment of the present invention can be classified or can be confirmed as for its binding region by the binding property with the peptide of the eighth embodiment of the present invention and/or the polypeptide of the ninth embodiment of the present invention. That is, the present invention provides an anti-GPVI antibody which has a distinct binding property, preferably a decreased binding property with the specified substance among the peptide of the eighth embodiment of the present invention and/or the polypeptide of the ninth embodiment of the present invention. Specifically, it is an anti-GPVI antibody which has a binding property with the specified GPVI mutant significantly different from that with human GPVI and/ or other GPVI mutants, preferably a decreased binding property. Specific examples of the method of confirmation, polypeptides to be used, preferred classification and preferred antibodies are illustrated in EXAMPLES. An antibody of the present invention is not neccessarily limited regarding its antigen-binding valency and may be a monovalent antibody such as Fab or scFv. From the aspect of stability in vivo, especially in the blood, and/or binding property to GPVI or strength of action, preferred is a multivalent antibody having two or more valencies, e.g., a divalent, trivalent, tetravalent or decavalent antibody; a divalent antibody is more preferable. Accordingly, in the second embodiment of the present invention, a monovalent antibody and a polyvalent antibody having two or more valency which recognizes a specific region, especially loop 9 on GPVI, such as a divalent, trivalent, tetravalent or decavalent antibody; preferably a divalent antibody can be -provided. Herein, examples of tetravalent antibody include IgA, and examples of decavalent antibody include IgM. However, they are not limited to these examples. Further, a trivalent antibody does not phisologically exit. However, by chemically or through genetic engineering binding 8 a natural or synthetic peptide having an intrinsic trimerization property such as the domain of tenascin molecule (AA 110-139, Swissprot #P10039 (chicken) or Swissprot #P24821 (human)) to a monovalent antibody (scFv or Fab and so on), a trivalent antibody can be prepared (see, JP 2004-508828 publication). In addition, the antibody of the second embodiment of the invention is preferably an antibody which exhibits a specific function or property of the antibody of the first embodiment. The third embodiment of the present invention is an anti-GPVI antibody which comprises a novel amino acid sequence of CDR or variable region, preferably a chimeric antibody which is recombined with human IgG, especially human IgG4, more preferably a CDR grafted antibody, especially a humanized antibody. Specifically, it includes: (15) An anti-GPVI antibody, its active fragment or derivatives thereof, wherein at least three CDR of either H-chain or L-chain in the antibody, preferably six CDR of both H-chain and L-chain in the antibody comprises an amino acid sequence of CDR in the clones listed in Tables 8, 9, 12 and 13, preferably an antibody which recognizes loop 9 of GPVI as an amino acid sequence of the corresponding CDR; (16) A heavy chain of anti-GPVI antibody, its active fragment, or derivatives thereof, wherein the amino acid sequences of SEQ ID NOs: 15, 16, and 17, , the amino acid sequences of SEQ ID NOs: 18, 19, and 20, the amino acid sequences of SEQ ID NOs: 21, 22, and 23, the amino acid sequences of SEQ ID NOs: 24, 25, and 26, the amino acid sequences of SEQ ID NOs: 27, 28, and 29, the amino acid sequences of SEQ ID NOs: 30, 31, and 32, the amino acid sequences of SEQ ID NOs: 33, 34, and 35, the amino acid sequences of SEQ ID NOs: 36, 37, and 38, the amino acid sequences of SEQ ID NOs: 39, 40, and 41, the amino acid sequences of SEQ ID NOs: 42, 43, and 44, the amino acid sequences of SEQ ID NOs: 45, 46, and 47, or the amino acid sequences of SEQ ID NOs: 48, 49, and 50, or VH CDRl, VH CDR2, and VH CDR3 of any clone listed in Table 12 are comprised in VH CDRl, VH CDR2, and VH CDR3, respectively; (17) A light chain of anti-GPVI antibody, its active fragment, or derivatives thereof, wherein the amino acid sequences of SEQ ID NOs: 51, 52 and 53, the amino acid sequences of SEQ ID NOs: 54, 55 and 56, the amino acid sequences of SEQ ID NOs: 57, 58 and 59, the amino acid sequences of SEQ ID NOs: 60, 61 and 62, the amino acid sequences of SEQ ID NOs: 63, 64 and 65, the amino acid sequences of SEQ ID NOs: 66, 67 and 68, the amino acid sequences of SEQ ID NOs: 69, 70 and 71, the amino acid sequences of SEQ ID NOs: 72, 73 and 74, the amino acid sequences of SEQ ID NOs: 75, 76 and 77, the amino acid sequences of SEQ ID NOs: 78, 79 and 80, the amino acid sequences of SEQ ID NOs: 81, 82 and 83, or the amino acid sequences of SEQ ID NOs: 84, 85 and 86, or VL CDRl, VL CDR2, and VL CDR3 of any clone listed in Table 13 are comprised in VL CDRl, VL CDR2 and VL CDR3, respectively; 9 (18) An anti-GPVI antibody, its active fragment, or derivatives thereof, wherein the amino acid sequences of SEQ ID NOs: 15, 16, 17, 51, 52 and 53, the amino acid sequences of SEQ ID NOs: 18,19, 20, 54, 55 and 56, the amino acid sequences of SEQ ID NOs: 21, 22, 23, 57, 58 and 59, the amino acid sequences of SEQ ID NOs: 24, 25, 26, 60, 61 and 62, the amino acid sequences of SEQ ID NOs: 27, 28, 29, 63, 64 and 65, the amino acid sequences of SEQ ID NOs: 30, 31, 32, 66, 67 and 68, the amino acid sequences of SEQ ID NOs: 33, 34, 35, 69, 70 and 71, the amino acid sequences of SEQ ID NOs: 36, 37, 38, 72, 73 and 74, the amino acid sequences of SEQ ID NOs: 39, 40, 41, 75, 76 and 77, the amino acid sequences of SEQ ID NOs: 42, 43, 44, 78, 79 and 80, the amino acid sequences of SEQ ID NOs: 45, 46, 47, 81, 82 and 83, or the amino acid sequences of SEQ ID NOs: 48, 49, 50, 84, 85 and 86, or VH CDR1, VH CDR2, VH CDR3, VL CDR1, VL CDR2, and VL CDR3 of any clone listed in Tables 12 and 13 are comprised in VH CDR1, VH CDR2, VH CDR3, VL CDR1, VL CDR2 and VL CDR3, respectively; (19) An anti-GPVI antibody, their active fragments or derivatives thereof, wherein at least variable region of either H-chain or L-chain in the antibody, preferably variable regions of both H-chain and L-chain in the antibody comprises an amino acid sequence of variable region in the clones listed in Table 7 or Table 14, preferably an antibody which recognizes loop 9 of GPVI as an amino acid sequence of the corresponding variable regions, especially a chimeric antibody which is recombined with human IgG, preferably human IgG4. The antibody of the third embodiment of the present invention is preferably an antibody which possesses characteristics and/or specificity for recognition region of the antibody of the first and/or the second embodiment. The fourth embodiment of the present invention is a polynucleotide or nucleic acid which comprises the base sequence encoding at least three CDR of either H-chain or L-chain, preferably six CDR of both H-chain and L-chain, or variable region in the antibody of the first to the third embodiments, its active fragment or derivatives thereof. Specifically, it includes: (20) A polynucleotide which comprises the base sequence encoding H-chain and/ or L-chain of the antibody of the first to the third embodiments, its active fragment or derivatives thereof; (21) The polynucleotide of (20) which comprises the base sequence encoding the corresponding CDR in the clones listed in Tables 8 and 9, or Tables 12 and 13, preferably an antibody which recognizes loop 9 of GPVI as the base sequence encoding at least three CDR of either H-chain or L-chain in the antibody, preferably six CDR of both H-chain and L-chain in the antibody; (22) A polynucleotide which comprises the base sequence encoding the corresponding variable region in the gene of any clone, preferably an antibody which recognizes loop9 of GPVI, listed in Tables 7 and 14, as the base sequence encoding at least variable region of either H-chain or L-chain in the antibody, preferably variable regions of both H-chain and L-chain in the antibody; 10 (23) A polynucleotide comprising the base sequence having SEQ ID NO: 280 and encoding variable region of the H-chain and the base sequence having SEQ ID NO: 284 and encoding variable region of the L-chain, and a polynucleotide comprising the base sequence having SEQ ID NO: 282 and encoding variable region of the H-chain and the base sequence having SEQ ID NO: 284 and encoding variable region of the L-chain. Further, the present invention provides an anti-human GPVI antibody gene, or its heavy chain or light chain variable region gene thereof, which is derived from an antibody gene comprising a combination of specific mouse germ-line antibody gene segments. That is, (24) An anti-human GPVI antibody gene, or a heavy chain variable region gene thereof, which is derived from an antibody heavy chain gene comprising any combination of mouse germ-line antibody gene segments VH, DH and JH listed in Table 16. (25) An anti-human GPVI antibody gene, or a heavy chain variable region gene thereof, comprising a nucleotide sequence encoding CDR amino acid sequence in the aforementioned antibody heavy chain variable gene. (26) An anti-human GPVI antibody gene, or a light chain variable region gene thereof, which is derived from an antibody light chain gene comprising any combination of mouse germ-line antibody gene segments VL and JL listed in Table 16. (27) An anti-human GPVI antibody gene, or a light chain variable region gene thereof comprising a nucleotide sequence encoding CDR amino acid sequence in the aforementioned antibody light chain variable gene. Herein, among the mouse germ-line antibody gene segments listed in Table 16, a combination of segments having a high score indicated on the first line of each antibody clones, for example, a combination of VH (3:3.9), DH (DSP2.7 or DSP2.5) and JH (JH4) in the heavy chain gene of clone F1246-1-1 is preferred. Further, a gene derived from the above-mentioned antibody gene includes the antibody gene itself or a gene with mutation in one base or more as long as an antibody encoded by the gene exhibits a similar antigen-binding specificity. In addition, both of naturally occurring or artificially introduced mutations may be acceptable . At the same time as the above, the present invention provides an antibody, its active fragment or derivatives thereof, encoded by the anti-human GPVI antibody gene or its heavy chain or light chain variable region gene, which is derived from an antibody gene comprising a combination of specific mouse germ-line antibody gene segments. That is, (28) An anti-human GPVI antibody or its heavy chain variable region polypeptide encoded by the antibody gene or its heavy chain variable region gene of (24) to (25). (29) An anti-human GPVI antibody or its light chain variable region polypeptide encoded by the antibody gene or its light chain variable region gene of (26) to (27). 11 In addition, the present invention provides an anti-GPVI antibody, especially anti-human GPVI antibody, specifically, the above-mentioned antibody of the present invention, preferably the antibody recognizing loop 9 of GPVI, or its active fragment or derivatives thereof, which is polyethyleneglycolated (PEGylated). The method for PEGylating the antibody etc. may be in accordance with publicly known method (for example, Roberts M. J. et al., Advanced Drug Delivery Reviews 54 (2002) 459-476), and specifically is described in EXAMPLE 31. The fifth embodiment of the present invention is a cell which produces the antibody of the first through the third embodiment, its active fragment or derivatives thereof, or a cell which comprises the polynucleotide of the fourth embodiment. Specifically, it includes: (30) A cell which produces any antibody described in the aforementioned (1) through (19), its active fragment, or derivatives thereof, especially a transformant or a hybridoma; (31) A cell which comprises any polynucleotide described in the aforementioned (20) through (23), especially a transformant or a hybridoma. The sixth embodiment of the present invention is a method of manufacturing the antibody of the first through the third embodiment, which is characterized by using the polynucleotide of the fourth embodiment or an expression vector comprising the same, or the cell of the fifth embodiment. Specifically, it includes: (32) A method of manufacturing the antibody of the first through the third embodiment, which comprises a process, in which the cell of the aforementioned (30) or (31) is cultured, and a process, in which a monoclonal antibody produced by the cell is collected; (33) A method of manufacturing the antibody of the first through the third embodiment, which comprises a process, in which any among the polynucleotide of the aforementioned (19) through (23), an expression vector comprising the same, and the cell of the above (30) or (31) is used. The seventh embodiment of the present invention relates to a medical composition comprising the antibody of the first to the third embodiment of the present invention, its active fragment or derivatives thereof as an effective ingredient, and preferably is a medical composition for prophylaxis and/or therapy of thrombotic, embolic or arteriosclerotic disease. The antibody of the present invention hardly has side effects such as activation of platelet, platelet aggregation, thrombocytopenia and prolongation of bleeding time and so on, and is useful for prevention and/or therapy of the above-mentioned diseases. The eighth embodiment of the present invention is a peptide which constitutes the specific structure, especially a loop structure on GPVI, specifically, 12 (34) a peptide which comprises loop 2, loop 3 and loop 5, or loop 4 and loop 5 in domain 1, or loop 9, or loop 9 and loop 11 in domain 2 of GPVI, especially human GPVI, especially, a peptide which consists of any one of amino acid sequence among them. Herein, the peptide may comprise an amino acid sequence derived from heterogeneous GPVI or an amino acid sequence of a polypeptide other than GPVI such as Fc. The ninth embodiment of the present invention is a specified GPVI mutant, for example, a mutant with an amino acid substitution, a domain substitution between species or a partial sequence substitution between species such as a loop substitution, and the like. Preferred is a mutant wherein amino acids constituting one or more loop structures of GPVI shown in FIGs 1 and 3 may be substituted with other amino acids or amino acids of the corresponding loop of other species such as human, monkey, mouse and rat. Specific examples are described in Table 4 or EXAMPLES. Specifically, (35) a polypeptide which comprises the amino acid sequence of SEQ ID NOs: 137 through 151. The tenth embodiment of the present invention is a method of screening an antibody, its active fragment or derivatives thereof, which comprises the following process: a) A process for measuring the binding property with platelet membrane glycoprotein VI (GPVI), especially human GPVI; b) A process for measuring the action to activate a platelet and/ or the function to induce thrombocytopenia in vivo; and c) A process for measuring the activity to at least partially deplete GPVI on the platelet membrane by contacting with a platelet. The eleventh embodiment of the present invention is a method of estimating an epitope of an antibody or a method of identifying a recognition region of an antibody, which comprises a process for measuring the reactivity, for example, the binding property with the antibody of the peptide of the eighth embodiment or the polypeptide of the ninth embodiment. The twelfth embodiment is a method of manufacturing an antibody specific for GPVI, which is characterized by using the peptide of the eighth embodiment or the polypeptide of the ninth embodiment, specifically, (36) a method of manufacturing an antibody specific for GPVI, preferably the antibody of the first to the third embodiments of the present invention, which is characterized by using the peptide of the eighth embodiment or the polypeptide of the ninth embodiment as an antigen for immunization or as an antigen for in vitro immunization; 13 (37) a method of manufacturing an antibody specific for GPVI, preferably the antibody of the first to the third embodiments of the present invention, which is characterized by using the peptide of the eighth embodiment or the polypeptide of the ninth embodiment as an antigen for detection or identification of the antibody. That is, using a recombinant GPVI as an immunogen and/or an antigen for detection, wherein the amino acid sequence on human GPVI, such as the amino acid sequence corresponding to the loop structure, which can be recognized by the antibody of the present invention, is integrated into mouse GPVI, a novel antibody which can recognize the same recognition region can be obtained. As a more preferred method of preparing an antibody for human therapy, the method using human antibody gene-transgenic non-human animal has been disclosed (WO 2002/070648 (Tokuhyou 2005-504507) and WO 2002/043478 (Tokuhyou 2004-515230)). When using a protein, wherein a partial amino acid sequence of human GPVI is integrated into a heterogeneous GPVI, for example, mouse GPVI, the above-mentioned transgenic animal such as a mouse is immunized, it will be considered that a human antibody which reacts with the integrated amino acid sequence from human, preferably epitope, but not the amino acid sequence from mouse on GPVI can be obtained efficiently. Therefore, the human antibody obtained by such method is useful as a human antibody having the features of the antibody of the first or the second embodiment, and said method is particularly useful. The thirteenth embodiment of the present invention is a method of detecting or quantifying GPVI in the test sample using the antibody of the first to the third embodiments. By the method, GPVI on the platelet or in the body fluid, especially in the blood can be measured. Further, it can be applied to a method of diagnosing diseases, preferably a method of diagnosing diseases associated with clot formation. Furthermore, the method can be applied to monitoring for treatment relating to GPVI, particularly prediction or determination of efficacy of anti-GPVI antibody, or prognostic determination using GPVI on the platelet as an index. BRIEF DESCRIPTION OF DRAWINGS Figure 1 is the alignment of the amino acid sequence of human soluble GPVI and mouse soluble GPVI. The squares show the positions of each domain region of GPVI and the loop region deduced by the modeling (L1-L14). Figure 2 presents the results of the competitive test of an antibody from a patient with GPVI deficiency with mouse anti-human GPVI monoclonal antibody. YA-Abs-88 and YA-Abs-03 mean anti-GPVI antibodies derived from a patient with GPVI deficiency. 14 Figure 3 is the alignment of the amino acid sequence of human soluble GPVI and rat soluble GPVI. The squares show the positions of each domain region of GPVI and the loop region deduced by the modeling (L1-L14). Figure 4 shows the results of reactivity for mouse hybridoma antibody and chimeric antibody. Figure 5 shows the inhibition of binding between GPVI and collagen by the chimeric antibody and the mouse hybridoma antibody. Figure 6 presents the results of a binding property of anti-human GPVI antibody to GPVI mutant. Figure 7 shows the results of a reactivity of cF1232-37-2 with various hGPVI mouse loop substitution mutants. Figure 8 shows the activating action on human and cynomolgus platelets. Expression level of CD62P (P-selectin) on human platelet (A) and cynomolgus monkey platelet (B) was assayed by FACS and the results were shown by mean fluorescent intensity (MFI). Figure 9 shows the aggregation-inducting action on human platelet. Figure 10 shows the action of F1232-37-2 on collagen-induced human platelet aggregation. Figure 11 shows the action of F1232-37-2 on ADP-induced human platelet aggregation. Figure 12 shows the results of the test of intravenous administration of mouse anti-human GPVI monoclonal antibodies F1232-37-2 and F1199-6 into cynomolgus. Figure 13 shows the results of ex vivo test of single intravenous administration of mouse/human chimeric anti-human GPVI antibody (cF1232-37-2) in cynomolgus. Figure 14 shows the results of ex vivo test of repetitive intravenous administrations of mouse/human chimeric anti-human GPVI antibody in cynomolgus. When 0.3 mg/kg of cF1232-37-2 was administered four times to cynomolgus monkey every other day, collagen-induced platelet aggregation (A) and platelet GPVI level (B) were measured. Figure 15 shows the results of ex vivo test of subcutaneous administration of mouse/human chimeric anti-human GPVI antibody in cynomolgus. After cF1232-37-2 was subcutaneously administered to cynomolgus monkey, the blood was collected with time, and platelet aggregation (A) induced by 2 mg/mL of collagen and platelet GPVI level (B) were measured. 15 Figure 16 shows an inhibiting action of F1232-37-2 Fab on collagen-induced platelet aggregation. Figure 17 shows the results of ex vivo test of F1232-37-2 F(ab')2 in cynomolgus. Figure 18 shows the construction of a stable co-expression plasmid for both chains of cF1232-37-2. Figure 19 shows a reactivity for antigen binding of cF1232-37-2 expressed in COS cells and CHO cells. Figure 20 shows an amino acid sequence of heavy chain variable region and humanization thereof. Figure 21 shows an amino acid sequence of light chain variable region and humanization thereof. Figure 22 shows a binding specificity of humanized antibody to GPVI. Figure 23 presents a result of collagen-induced aggregation test of platelet from cynomolgus monkey administered with chimeric anti-GPVI antibody. Figure 24 presents a result of the bleeding time test in cynomolgus monkey. A: shows the collagen-induced platelet aggregation at five minutes after intravenous administration of eptifibatide and at 48 hours after administration of anti-GPVI antibody cF1232-37-2. B: shows a result of the bleeding time at five minutes after intravenous administration of eptifibatide and at 48 hours after administration of anti-GPVI antibody cF1232-37-2 compared with the bleeding time prior to administration. Figure 25 shows a result of confirmation test for platelet GPVI antigen shedding by anti-GPVI antibodies. Figure 26 presents a result of PEGylation of anti-GPVI whole antibody and Fab antibody. Lanes shows 1: F1232-37-2 whole antibody; 2: PEGylated product of F1232-37-2; 3: PEGylated purified product of F1232-37-2; 4: F1232-37-2 Fab antibody; 5: PEGylated product of F1232-37-2 Fab; and 6: PEGylated purified product of F1232-37-2 Fab, respectively. Figure 27 shows the result of GPVI antigen binding assay for PEGylated anti-GPVI antibody. Figure 28 shows a nucleotide sequence of the rat GPVI gene and an amino acid sequence encoded by the same. Figure 29 presents a result of SDS-PAGE for rGPVI-Fc fusion protein obtained in EXAMPLE 35. Lanes 1, 2 and 3 show a molecular weight marker, rGPVI-hFc fusion protein and rGPVI-mFc fusion protein, respectively. 16 Figure 30 presents a result of assay for binding ability to GPVI loop substitution mutant. Figure 31 presents a result of assay for binding ability of anti-rat GPVI antibody to rat platelet. Figure 32 presents a result of assay for aggregation ability of rat platelet to which the anti-rat GPVI antibody was administered. Figure 33 shows a result of GPVI depletion in EXAMPLE 40. Figure 34 shows a result of assay for effects of anti-GPVI antibody to collagen-induced lethal model. Figure 35 shows a result of assay for effects of anti-GPVI antibody to electric stimulus-induced arterial thrombosis model. Figure 36 shows an assessment of antigen binding for CypHer5E-labeled cF1232-37-2/CHO. Figure 37 presents a result of in vitro internalization of CypHer5E-labeled cF1232-37-2/CHO. Figure 38 shows an assessment for antigen binding of CypHer5E-labeled F1239-6-1. Figure 39 presents a result of in vitro internalization of CypHer5E-labeled F1239-6-1. Figure 40 presents a result of in vivo internalization of CypHer5E-labeled F1239-6-1. Figure 41 shows a result of assessment for platelet activation (CD62P) of cF1232-37-2S/COS. Figure 42 shows a suppressing effect on collagen-induced platelet aggregation of CF1232-37-2S/COS. Figure 43 presents a result of ex vivo test using cynomolgus monkey for cF1232-37-2S/COS. The result was shown by collagen-induced platelet aggregation. Figure 44 shows a result of assay for antigen binding activity of PEGylated F1239-6-lFab. Figure 45 indicates a bleeding time of cynomolgus monkey under high-dose administration of cF1232-37-2. Figure 46 shows a result of assay for the bleeding time of rat in EXAMPLE 50. 17 Figure 47 shows an alignment of the amino acid sequences of both human soluble GPVI and cynomolgus soluble GPVI. Box indicates respective domain regions of GPVI and the position of loop regions (L1-L14) predicted by modeling. BEST MODE FOR CARRYING OUT THE INVENTION (Constitution) The antibody of the present invention is the one which specifically recognizes GPVI, the membrane glycoprotein existing on platelet such as mammalian platelet GPVI, specifically the platelet from human, monkey, rat or mouse, especially the human platelet. Further, the GPVI which is recognized by the antibody of the present invention is not always limited to the GPVI on the platelet, and for example, GPVI on megakaryocyte can also be recognized. Herein, the GPVI which is subjected by the present invention is a mammalian GPVI. For example, the GPVI from human, monkey, rat or mouse, especially the human GPVI is exemplified. The present invention will be described in detail as follows. In addition, in the specification, sometimes, an amino acid residue is denoted as one-letter code or three-letter code. The antibody of the present invention may be a polyclonal antibody, or preferably is a monoclonal antibody. A method of preparing the monoclonal antibody is not limited to the specified method. The monoclonal antibody may be any of a monoclonal antibody produced by hybridoma, a monoclonal antibody produced by a recombinant cell, in which a gene encoding the antibody is incorporated, or a monoclonal antibody produced by the transformed cell with Epstein-Barr virus (EBV). In addition, it may be a mixture of antibodies or a polyclonal antibodies comprising at least one of the monoclonal antibodies of the present invention, or a mixture of plural monoclonal antibodies of the present invention. Further, the antibody of the present invention encompasses a bispecific antibody or a polyspecific antibody. The antibody of the present invention is an antibody which specifically binds to mammalian GPVI, specifically GPVI from human, monkey, rat or mouse, especially human GPVI. The binding of the antibody of the present invention with GPVI, especially human GPVI can be measured by publicly known methods, specifically the method described in EXAMPLES. Dissociation constant (Kd value) between GPVI, especially human GPVI and the antibody of the present invention is 4 x 10"8 M, preferably equal to or less than 10"8 M, more preferably equal to or less than 4 x 10"9 M, further preferably equal to or less than 10-9 M. A method of determining dissociation constant between human GPVI and the antibody is not limited to the specified method, and the conventional method can be used. For example, it can be measured with the protein interaction analyzer such as BIACORE3000 using GPVI-Fc immobilized on chip. Alternatively, using a platelet, especially human or monkey platelet, it can be assayed with a publicly known method such as a method using Rl-labeled antibody. Specifically, it can be illustrated in EXAMPLE 5 and EXAMPLE 52. 18 The antibody of the first embodiment of the present invention has an activity for disappearing GPVI on the platelet membrane at least partially by bringing in contact with the platelet, in particular, by bringing in contact with the platelet in vivo. The activity can be confirmed by bringing the antibody of the present invention in contact with the platelet for a given time followed by isolating the platelet, and assaying the expression level of GPVI on the surface. The expression level of GPVI can be measured by the conventional method using FACS, etc., and the specific method is illustrated in EXAMPLES. The antibody of the present invention has an activity for depleting GPVI on the platelet equal to or more than 20%, preferably equal to or more than 40%, more preferably equal to or more than 60%, further preferably equal to or more than 80% in comparison with the value prior to administration or the control value at the dosage of 3 mg/kg, preferably 1 mg/kg, more preferably 0.3 mg/kg, further preferably 0.1 mg/kg. The antibody of the present invention is an antibody which has GPVI-depleting ability without mediating shedding of platelet GPVI, in particular, shedding of GPVI from platelets by methalloprotease-mediated cleavage accompanying with platelet activation, or an antibody having a weak function of the shedding induction, preferably no significant inducing function, more preferably no substantial inducing function, which at least partially depletes GPVI on the platelet membrane by contacting with platelets. Herein, shedding can be detected by the publicly known method (Stephens G and other four members, Blood, 2005 Jan 1; 105 (1): 186-191; Gardiner EE et al, Blood 2004, 104: 3611-3617; Bergmeier W et al., Thromb Haemost. 2004; 91: 951-958), and specifically the method described in EXAMPLE 30 may be applicable. The antibody of the present invention is an antibody, its active fragment or derivatives thereof which at least partially depletes GPVI on the platelet membrane by contacting with the platelet via internalization of the platelet GPVI. These antibodies are different from an antibody which induces the shedding of GPVI from the platelet by cleavage via methalloprotease associated with platelet activation. As described below, since they themselves have a weak action for activating platelets and/or inducing thrombocytopenia in vivo, preferably little action, they are useful. Preferred examples of such antibody include an antibody which recognizes loop 9 in GPVI. The internalization of the platelet GPVI by the antibody of the present invention may be detected with a publicly known method. Preferably, by a method using a labeling substance as shown in EXAMPLES, such as fluorescent material, preferably pH-sensitive fluorescent material, specifically by a method labeling the antibody of the present invention directly or indirectly with these labeling substances, uptake of antibody-bound GPVI into platelets can be detected, or an amount of the uptake can be determined. Preferred method is shown in EXAMPLES. The antibody of the present invention per se has a low activity for activating a platelet and/or inducing thrombocytopenia in vivo, preferably no activity. The activation of platelet can be measured by known methods, and an expression level 19 of a platelet surface antigen, preferably CD62P can be used as an index. For example, a method of isolating a platelet from a living body that the antibody has been administered after a given period and measuring the expression level of CD62P by a conventional method, a method of bringing the antibody of the present invention in contact with a platelet isolated from a living body, and assaying the expression level of CD62P after a given period by a conventional method, and the like are included. The specific method is shown in EXAMPLES. The activation of platelet by the antibody of the present invention is, when the expression level of CD62P is used as an index, at the dose or the concentration that at least partially depletes GPVI on the platelet, equal to or less than five-fold, preferably equal to or less than two-fold, more preferably equal to or less than 1.5-fold, further preferably almost the same as the platelet for control. Thrombocytopenia in vivo can be confirmed by collecting blood with time after in vivo administration of the antibody of the present invention , calculating the number of platelets with a conventional method and comparing the number with a value prior to administration or the number of platelets of an individual as a control. The specific method is shown in EXAMPLES. The platelet number by the antibody of the present invention is, at the dose or the concentration that at least partially disappears GPVI on the platelet, when the value prior to administration or the control value is set as 100%, equal to or more than 50%, preferably equal to or more than 70%, more preferably equal to or more than 90%, further preferably almost the same. The antibody of the present invention has the activity that suppresses the human platelet aggregability against the collagen, that is, the activity that attenuates or deletes aggregability of the platelet responsive to collagen by contacting with the platelet in vivo. The activity can be confirmed by administering the antibody of the present invention in vivo to make contact with the platelet, followed by isolating the platelet with time, and measuring a collagen-induced platelet aggregation. Herein, the platelet aggregation can be measured by the publicly known method. For example, it can be measured by calculating an aggregation rate using a light transmission with the platelet aggregometer as an index, and in general, may be represented by the aggregation rate at the point that exhibits a maximum light transmission (hereinafter, sometimes referred to as the maximum aggregation rate). In the method described in EXAMPLE 8 later, the antibody of the present invention has an activity that at the dosage of 3 mg/kg, preferably 1 mg/kg, more preferably 0.3 mg/kg, further preferably 0.1 mg/kg, makes the collagen-induced aggregability of the platelet decrease by 20%, preferably equal to or more than 40%, more preferably equal to or more than 60%, further preferably equal to or more than 80% in comparison to the value prior to the administration or the control value. Preferably, the antibody of the present invention hardly has an effect on aggregation induced by platelet aggregation-inducing substances except for collagen, for example, ADP or thrombin. At the dosage or the concentration that affects on collagen-induced aggregability, the maximum aggregation rate is preferably equal to or more than 80% of the control, more preferably equal to or 20 more than 90% of the control, further preferably equal to or more than 95% of the control. A method of assaying suppression of a human platelet aggregation by platelet aggregation-inducing substances except for collagen can be performed by the conventional methods. In addition, the antibody of the present invention is an antibody that has a weak prolonging activity on bleeding time, preferably no significant prolonging activity, more preferably no substantial prolonging activity. The bleeding time can be assayed by the publicly known method, and specifically the method described in EXAMPLE 28 or 50 may be applicable. The antibody of the present invention substantially does not prolong the bleeding time at the therapeutic dosage or more, for example, 0.3 mg/kg, preferably 1 mg/kg, more preferably 3 mg/kg, further preferably 10 mg/kg. Specifically, the bleeding time is less or equal to 5-fold, preferably less or equal to 3-fold, further preferably less or equal to 2-fold, particularly preferably less or equal to 1.5-fold as the value prior to administration, normal value or control. Such preferred examples include an antibody that recognizes loop 9 of GPVI, especially human GPVI Since most of the antibodies to the human GPVI, including the above-mentioned human autoantibody, that have been reported until now, possess, in vitro, an activity that activates a platelet by the antibody itself, and/or an activity that induces or enhance a platelet aggregation, when they are administered in vivo, the possibility to cause a thrombocytopenia may be considered. In the form of the Fab fragment and so on, the one that does not induce a platelet aggregation is also reported, meanwhile in vivo, a possibility that Fab behaves in a similar manner to IgG by cross-linking or aggregating from any cause cannot fully be denied. Therefore, in an intact antibody molecule, but not an active fragment of the antibody, for example, IgG form, an anti-GPVI antibody that does not exhibit the above-mentioned activity, or has a low activity is preferred. Also, for behavior and stability in vivo, the antibody molecule that is the natural form, e.g. IgG, is superior. Generally, a half-life of the IgG in the blood is much longer than that of the fragment such as Fab. Thus especially, for chronic diseases such as thrombosis, especially thrombosis associated with atrial fibrillation, or clinical conditions that necessitate an antibody administration over long period, a molecular form having a long half-life in the blood, particularly IgG is desirable. The antibody of the present invention may specifically inhibit the binding of GPVI on the platelet to collagen. For example, in the method described in EXAMPLES later, the antibody of the present invention is an antibody that inhibits the binding of GPVI and collagen by 50% at the concentration preferably equal to or less than 100 mg/mL, more preferably equal to or less than 10 mg/mL, further preferably equal to or less than 1 mg/mL, especially preferably equal to or less than 0.1 mg/mL. A method of measuring the binding of collagen and GPVI is not limited to a specific method and can also be done by the other conventional method. 21 The second embodiment of the present invention is an anti-GPVI antibody that is defined by novel recognition region, binding site or epitope on GPVI, preferably a monoclonal antibody. The recognition region on GPVI by the antibody of the present invention and the like can be confirmed or deduced by publicly known methods. For example, by applying the method of the eleventh embodiment of the present invention and measuring a reactivity with the peptide of the eighth embodiment or the polypeptide of the ninth embodiment, it can be performed. The specific method is illustrated in EXAMPLES. For example, in the method described in EXAMPLE 7 or EXAMPLE 18, on the basis where the reactivity or inhibition rate is significantly varied in comparison to the control (e.g., hGPVI-Fc), for example, it is reduced to 50%, preferably 30%, more preferably less or equal to 10%; or where a value for IC50 and the like are significantly changed, for example, it is increased three-fold, ten-fold, more preferably 30-fold, further preferably 100-fold, the antibody can be confirmed or deduced. The antibody, wherein recognition region, binding site or epitope on GPVI is confirmed, is useful for detection of specific GPVI molecular species, or for analysis of relationship between the structure and the function of GPVI, solely or by combination with other antibodies. As the third embodiment of the present invention, there is an anti-GPVI antibody comprising a novel amino acid sequence of CDR or variable region. On the N-terminal end of heavy chain and light chain of antibody, variable region exists, and is designated heavy chain variable region (VH) and light chain variable region (VL), respectively. Within the variable region, complementarity determining region (CDR) is present, and assumes specificity for recognition of antigen. A region except CDR in variable region has the role that maintains the structure of CDR and is called framework region (FR). On the C-terminal end of heavy chain and light chain of antibody, constant region exists, and is designated heavy chain constant region (CH) and light chain constant region (CL), respectively. In the heavy chain variable region, three complementarity determining regions exist: the first complementarity determining region (CDR1), the second complementarity determining region (CDR2) and the third complementarity determining region (CDR3). These three complementarity determining regions in the heavy chain variable region collectively mean a heavy chain complementarity determining region. As is the case with the heavy chain, in the light chain variable region, three complementarity determining regions exist: the first complementarity determining region (CDR1), the second complementarity determining region (CDR2) and the third complementarity determining region (CDR3). These three complementarity determining regions in the light chain variable region collectively mean a light chain complementarity determining region. CDR sequence in the antibody of the present invention is not always limited. Preferred combination of the amino acid sequences as VH CDR1, VH CDR2 and VH CDR3, preferred combination of the amino acid sequences as VL CDR1, VL CDR2 and VL CDR3, and further, preferred combination of the amino acid sequences as VH CDR1, VH CDR2, VH CDR3, VL CDR1, VL CDR2 and VL CDR3, are listed in 22 Tables 8 and 9, and Tables 12 and 13. Preferably, it is an antibody that comprises any one or more, preferably three of heavy chains, more preferably all among the amino acid sequences of CDR of an antibody that recognizes loop 9 of GPVI The amino acid sequences except for CDR are not particularly limited. The antibody of the present invention includes so-called CDR-grafted antibody, wherein the amino acid sequences except for CDR are derived from another antibody, especially the antibody from other species. Among them, preferred is a humanized antibody, wherein the amino acid sequences except for CDR are derived from human. The humanized antibody may have addition, deletion, substitution and/or insertion of one to several amino acid residues in the framework (FR) region, if desired. As a method of preparing the humanized antibody, publicly known methods can be used, and the specific method is shown in EXAMPLES. The amino acid sequence of VH and VL of the antibody of the present invention is not always limited, but preferred antibody is an antibody that comprises any one or more among the amino acid sequence of SEQ ID NO: 281 as VH or the amino acid sequence of SEQ ID NO: 285 as VL, or an antibody that comprises any one or more among the amino acid sequence of SEQ ID NO: 283 as VH or the amino acid sequence of SEQ ID NO: 285 as VL. In addition, the antibody of the present invention is not always limited to that with the specific amino acid sequence, and within the range where there is virtually no influence on its activity and/or antigenecity, as for the amino acid sequence of the antibody of the present invention, for example, variable region, especially FR , addition, deletion, substitution and/or insertion of one to several amino acid residues are permissible. The antibody of the present invention is an antibody that the constant region of the antibody consists of an amino acid sequence derived from preferably human antibody, more preferably human IgG, further preferably human IgG4. The antibody of this invention is not always limited to the specific molecular species. The structure of an antibody, i.e. an immunoglobulin consists of heavy chain (H-chain) and light chain (L- chain) and is divided into five isotypes (IgG, IgA, IgM, IgD, IgE) based on the class of the heavy chain (y, a, u, 5, t). Among them, IgG and IgA are divided, based on difference of the heavy chain (e.g., in the case of the human, \1, y2, \3, y4/ ctl, a2), into subclasses (e.g., in the case of the human, IgGl, IgG2, IgG3, IgG4, IgAl, IgA2). The light chain is classified into either K or A type. The class, the subtype or the isotype of the antibody of the present invention is not limited, and may be the one that is classified into either. Preferred isotype is IgG, and further preferably, the subclass is IgG4 in the point that there is no complement fixation. The antibody of the present invention, as long as the antibody has the activity such as a binding ability to GPVI, may be the fragment, particularly the active fragment or the part of the antibody. Herein, the active fragment of the antibody means to be a fragment having at least one activity, especially antigen binding activity of the antibody. For example, Fab (fragment of antigen binding), Fab1, (Fab')2, single-chain 23 antibody (scFv), disulfide stabilizing antibody (dsFv), diabody, sc(Fv)2 (see, e.g., Orita T, Blood. 2005; 105: 562-566), nanobody (see, e.g., Cortez-Retamozo V., Cancer Research 64, 2853-2857, 2004) and the peptide comprising CDR and so on are included. Also, a derivative of an antibody can be a substance derived from the antibody, having at least one activity of the antibody, especially the antigen binding activity. Examples include an antibody bound to other substance or active fragment thereof, modified antibody that is modified by other substance or active fragment thereof, or a molecule wherein a mutation is introduced to the structure of the antibody, especially the amino acid sequence. An antibody of the present invention is not always limited regarding antigen binding valency and may be monovalent antibody such as Fab or scFv. From the perspective of stability in vivo, especially in the blood, and/or binding property to GPVI or strength of action, preferred is a polyvalent antibody having two or more valencies, e.g., a divalent, trivalent, tetravalent or decavalent antibody; a divalent antibody is more preferable. In the fourth embodiment of the present invention, a polynucleotide or nucleic acid encoding the antibody of the first embodiment to the third embodiment of the present invention is provided. The polynucleotide is not always limited as long as that encodes the amino acid sequence of the antibody of the present invention, and includes DNA and RNA. A polynucleotide encoding CDR sequence in the antibody of the present invention is not always limited. Preferred combination of the base sequences encoding the amino acid sequence as VH CDR1, VH CDR2 and VH CDR3, preferred combination of the base sequences encoding the amino acid sequence as VL CDR1, VL CDR2 and VL CDR3, and further, preferred combination of the base sequences encoding the amino acid sequence as VH CDR1, VH CDR2, VH CDR3, VL CDR1, VL CDR2 and VL CDR3, are listed in Tables 8 and 9, and Tables 12 and 13. The polynucleotide encoding the amino acid sequence of VH and VL of the antibody of the present invention is not always limited, and preferably is a polynucleotide that comprises any one of the base sequences of SEQ ID NOs: 87, 89, 91, 93, 95, 97, 99,101,103,105,107 and 109 encoding the amino acid sequence as VH or the base sequences of SEQ ID NOs: 88, 90, 92, 94, 96, 98, 100, 102, 104, 106, 108 and 110 encoding the amino acid sequence as VL, more preferably both base sequences, or a polynucleotide that comprises either the base sequences having the corresponding sequence identification number and encoding the amino acid sequence as VH or the base sequences having the corresponding sequence identification number and encoding the amino acid sequence as VL, more preferably both base sequences. The polynucleotide encoding the constant region of the antibody of the present invention comprises a base sequence derived from preferably human antibody, more preferably human IgG, further preferably human IgG4. 24 By transferring the vector comprising a nucleotide sequence encoding the antibody of the present invention, or the gene into a cell, a cell that produces the antibody of the present invention can be manufactured. A method of transfer may be according to publicly known methods, and the specific methods are illustrated in EXAMPLES. As the fifth embodiment of the present invention, a cell that produces the antibody of the present invention is provided. Examples of such cell include hybridoma, transformant, or recombinant cell, in which a gene encoding the antibody of the present invention is introduced. Hybridoma that produces the antibody specifically includes clones listed in Tables 6 and 11. In addition, the present invention provides an antibody that is produced by the abovementioned cells of the present invention. The eighth and the ninth embodiments of the present invention provide a novel peptide or polypeptide relating to GPVI. These peptides can be prepared by publicly known methods, and the specific method is shown in EXAMPLES. The peptide of the eighth embodiment and the polypeptide of the ninth embodiment can be used as an antigen for immunization, or as an antigen for detection of the anti-GPVI antibody. (Manufacturing process) The sixth embodiment of the present invention provides a method of manufacturing an antibody. The method of preparing the antibody of the present invention is not always limited, and the antibody can be prepared by the method described below. That is, by administering human GPVI, its fragment or derivatives thereof, for example, human GPVI-Fc as an antigen to animal such as mouse, collecting lymphocytes from the peripheral blood, hybridoma with mouse myeloma cell may be prepared. An antibody that is produced by the prepared hybridoma is recovered to select the antibody having a binding ability to GPVI and characteristics of the first to the third antibody. Thereby, a cell, in which the antibody is produced, can be obtained. By culturing the cell, the antibody of the present invention can be obtained. The antibody of the present invention can be prepared as the recombinant human antibody using publicly known methods (a lot of methods are developed since Nature, 312: 643, 1984, and Nature, 321: 522, 1986 were published, respectively). Firstly, from the cells that produce the antibody of the present invention, e.g. the lymphocytes, preferably, the hybridoma which produces anti-GPVI monoclonal antibody, nucleic acid encoding VH or VL, e.g. cDNA may be obtained, and the base sequence and the amino acid sequence are determined. Then, by inserting the obtained cDNA encoding VH and VL into the expression vector for animal cells comprising a gene encoding human antibody CH and/ or human antibody CL that has been prepared from the same or other human cell, respectively, human antibody-expressing vector may be constructed. By introducing the vector into the animal cell and expressing it, the antibody of the present invention can be manufactured. A method of preparing the gene to be introduced to the animal cell 25 is not limited, and may be obtained from genomic DNA or cDNA derived from hybridoma, by PCR from mRNA of hybridoma, or also by the chemical synthesis. The vector, into which to the nucleic acid encoding VH or VL of the antibody of the present invention is incorporated, is not always limited, but a vector or a vector for high expression that generally is used for expression of gene encoding protein and adapted to expression of the antibody gene is preferred. Preferred example includes a vector containing EF promoter and/or CMV enhancer, specifically pEF-BOS or the vector used in EXAMPLES. In addition, the vector that the nucleic acid encoding VH or VL is incorporated is usually prepared independently, and co-transfected into host cells. However, the nucleic acid may be incorporated into a single expression vector. The host cell, in which the expression vector is introduced, is not always limited, and the cell that generally is used for expression of gene encoding protein and adapted to expression of the antibody gene is preferred. For example, bacteria (Escherichia coli, etc.), actinomyces, yeasts, insect cell (SF9, etc.), mammalian cell (COS-1, CHO, myeloma cell, etc.) are included. To industrially produce a recombinant antibody, generally a recombinant animal cell line that stably and highly expresses the antibody, for example, a CHO cell line may be utilized. For preparation of such recombinant cell lines, cloning, gene amplification for high expression and screening, a publicly known method can be used (see, e.g., Omasa T.: J. Biosci. Bioeng., 94, 600-605, 2002; and others). Further, two kinds of promoters can be used for establishment of animal cell line for stably high expression. Herein, by using combination of different promoter activities such as high activity and low activity, preferably utilizing a promoter having a relatively weak activity as an expression promoter for selection marker, a clone having an high expression ability can be efficiently (or selectively) acquired. Examples of preferred combination of promoters and specific methods are indicated in EXAMPLES. As the constant region of the human antibody to use for preparing a recombinant human antibody, any human antibody constant region, for example, Cyl and Cy4 for the human antibody heavy chain constant region, and CK for the human antibody light chain constant region can be used. Among the antibodies of the present invention, the antibody comprising an amino acid sequence derived from human includes, as well as an antibody that is naturally occurring in the human body, a combinatorial library consisting of variable heavy chain and variable light chain, for example, a phage library for human antibody, and an antibody obtained from human antibody producing-transgenic animal, etc. The phage library for human antibody is a library, wherein an active fragment of antibody such as Fab and single-stranded antibody is made express on the phage surface by inserting a gene encoding the antibody prepared from human B cell into phage gene. The antibody of the present invention can be obtained by screening these libraries. These and other methods are well-known by persons skilled in the 26 art (Huse et al., Science 246: 1275-1281 (1989); Winter and Harris, Immunol. Today 14: 243-246 (1993); Ward et al., Nature 341: 544-546 (1989); Harlow and Lane, supra, 1988); Hilyard et al., Protein Engineering: A practical approach (IRL Press 1992); Borrabeck, Antibody Engineering, 2nd Ed. (Oxford University Press 1995)). From the library, using an binding activity to the substrate immobilized an antigen as an index, a phage expressing an active fragment of the antibody having the desired antigen binding activity can be recovered. The active fragment of the antibody can further be converted into a human antibody molecule consisting of two intact H-chains and two intact L-chains by genetic engineering techniques. The present invention includes, in addition to the antibody consisting of two heavy chains and two light chains, the active-fragment of the antibody of the present invention. For example, the active fragment of the antibody includes Fab (fragment of antigen binding), Fab', F(ab')2. The substance that the active fragment of the antibody is linked by linker and so on includes, for example, single-stranded antibody (single chain Fv: scFv) and disulfide stabilized Fv : dsFv. The peptide that contains the active fragment of the antibody includes, for example, a peptide containing CDR. These can be manufactured by the method of processing the antibody of the present invention with the suitable protease or the publicly known methods such as the recombinant DNA techniques. Fab of the present invention can be obtained by treating the anti-GPVI antibody of the present invention with pepsin, the proteolytic enzyme in case of IgM, or by processing it with the protease papain in case of IgG. Alternatively, Fab can be produced by inserting DNA encoding Fab of the antibody into prokaryotic or eukaryotic expression vector, introducing the vector into prokaryotes or eukaryotes, and expressing the same. F(ab')2 of the present invention can be obtained by treating the anti-GPVI antibody of the present invention with pepsin, the proteolytic enzyme. Alternatively, it can be prepared by linking the following Fab' with the thioether bond or the disulfide bond. Fab' of the present invention can be obtained by treating F(ab')2 that specifically reacts to GPVI with the reducing agent, dithiothreitol. As VH and VL that are comprised in scFv of the present invention, those derived from either the antibody or the human antibody, which the hybridoma of the present invention produces, can be used. The scFv of the present invention can be manufactured by obtaining cDNA encoding VH and VL of the anti-GPVI antibody of the present invention, constructing the DNA encoding scFv, inserting the DNA into prokaryotic expression vector or eukaryotic expression vector, and introducing the vector into prokaryotes or eukaryotes to express the vector. The term "dsFv" means an antibody that two polypeptides, wherein one amino acid residue in each of VH and VL is substituted with cysteine residue, bind each other via disulfide bond between said cysteine residues. The amino acid residue to be 27 substituted for the cysteine residue can be selected based on the three-dimensional structure prediction of the antibody according to the method shown by Reiter et al. [Protein Engineering, 7, 697 (1994)]. As for VH and VL that is contained in dsFv of the present invention, the ones, which are derived from the antibody of either the first or second embodiment of the present invention may be used. The dsFv of the present invention can be manufactured by obtaining cDNA encoding VH and VL of the anti-GPVI antibody of the present invention, constructing DNA encoding dsFv, inserting the DNA into prokaryotic or eukaryotic expression vector, and introducing the expression vector into prokaryotes or eukaryotes to express the same. The peptide containing CDR is constituted by including at least one region or more of the H-chain CDR or of L-chain CDR. Plural CDRs can be bound directly or through the appropriate peptide linker. The peptide that contains CDR of the present invention can be manufactured by obtaining cDNA encoding VH and VL of the anti-GPVI antibody of the present invention, constructing DNA encoding CDR, inserting the DNA into prokaryotic or eukaryotic expression vector, and introducing the expression vector into prokaryotes or eukaryotes to express the same. Alternatively, the peptide that contains CDR can be manufactured by the chemical synthesis such as the Fmoc method (the fluorenylmethyloxycarbonyl method), the tBoc method (t-butyloxycarbonyl method), or the like. The antibody of the present invention, its active fragment or derivatives thereof include, for example, an antibody that is produced by hybridoma, an antibody that is produced by the cell transformed with EBV, a recombinant antibody expressed from cDNA, or an antibody, wherein a radioisotope, protein, peptide or low molecular, etc. is chemically conjugated or through genetic engineering fused with the active fragment of the antibody. For example, an antibody bound to polyethyleneglycol etc. is highly useful in respect of stability, and is one of preferred examples. To the N-terminal or the C-terminal end of H-chain or L-chain in the anti- GPVI antibody of the present invention or an active fragment thereof, an appropriate residue or a side chain in the antibody or an active fragment of the antibody, and further a sugar chain in the antibody or an active fragment of the antibody, a radioisotope, protein, peptide or low molecular weight compound, etc. can be conjugated by a chemical method [The introduction to antibody engineering (Koutai kougaku nyuumon) (written by Osamu Kanemitsu, 1994, Chijin Shokan)]. Hybridoma means a cell that produces the monoclonal antibody having the desired antigen specificity, wherein it is obtained by fusing a lymphocyte with the myeloma cell derived from human, mouse, rat and so on, and can be prepared by publicly known methods. When preparing a monoclonal antibody, in consideration of the compatibility with myeloma cell used for the cell fusion, selection is preferably performed. As for the myeloma cell, publicly known various cells are usable. These include SKO-007 from 28 human, SHM-D33, which is a human-mouse heterozygous myeloma, P3, P3U1, SP2/0, NS-1 derived from mouse, and YB2/0 and Y3-Agl through Ag3 from rat. In the case of human antibody, a method of preparing hybridomas utilizing activation of lymphocytes by in vitro immunization and a method of preparing hybridomas using an animal, in which human antibody gene is recombined, in particular, a transgenic mouse such as KM mouse are included (WO 2002/070648 (Tokuhyou 2005-504507) and WO 2002/043478 (Tokuhyou 2004-515230)). When using a heterogeneous GPVI, for example, a protein, wherein a partial amino acid sequence of human GPVI is integrated into mouse GPVI, the above-mentioned transgenic animal such as a mouse is immunized, it will be considered that a human antibody that reacts with the integrated amino acid sequence from human, preferably epitope, but not the amino acid sequence from mouse on GPVI can be obtained efficiently. Therefore, the human antibody obtained by such method is useful as a human antibody having the features of the antibody of the first or the second embodiment, and said method is particularly useful. A cell used for preparation of hybridomas is not always limited. In the case of preparing human antibody, among plural cells for preparation of hybridomas, at least one is preferably a cell derived from human. As the cell derived from human, lymphocytes from peripheral blood, lymph nodes or spleen can be used, and especially, human lymphocytes, in which production of autoantibody is confirmed, is preferred. Activation of lymphocyte can be according to the publicly known method. For example, preferred are a method of preparing hybridoma with the myeloma cell derived from the human B cell or the mouse myeloma cell by collecting B cell from peripheral blood or spleen of the human and stimulating an antigen with in vitro immunization, a method of fusing with the mouse myeloma cell by transforming with EBV, and a method of fusing by stimulating with mitogen such as PWM and activating B cell to polyclonal antibody (Immunological experiment procedures (Men-eki jikken sousa-hou) I and II, edited by Shunsuke Migita et al., Nankoudo). The antigen used for immunizing an animal or for stimulation of the cell is not always limited. The animal, from which the protein as an antigen is originated, can be appropriately selected for any purpose of the antibody. The protein as an antigen may be naturally occurring product, genetically engineered product, chemically synthesized product, or fusion protein with other protein or peptide, and the like. For example, the platelet, the membrane of the platelet, purified GPVI, recombinant GPVI, and GPVI-Fc, preferably GPVI-Fc can be used. In addition, the peptide of the eighth embodiment of the present invention and the polypeptide of the ninth embodiment can appropriately be used as an antigen for immunization to prepare an anti-GPVI antibody. Fusion of the activated lymphocytes with myeloma cells can be performed using the publicly known methods such as the method by Milstein et al. (Methods in Enzymol., volume 73, pages 3). The methods include, for example, the method using polyethylene glycol (PEG) as a fusing agent (Introduction to the monoclonal 29 antibody experiment procedure (Tan-kuron koutai jikken sousahou nyuumon), written by Tamie Ando and Takeshi Chiba, Kodansha) or the electrofusion method. The mixing ratio of the immunocyte and the myeloma cell is not limited as long as it is the ratio that the cells can be fused. Preferably, 1/10 to equal amount of the myeloma cells to the activated lymphocytes may be used. In the cell fusion using PEG (average molecular weight: 1,000-4,000), PEG concentration is not always limited, but 50% is preferable. In addition, as the fusion efficiency accelerator, auxiliary substance such as dimethylsulfoxide (DMSO) may be added. The fusion is started by adding pre-warmed PEG solution at 37°C to the mixed cells, and is terminated by adding medium after the reaction for 1-5 minutes. The hybridoma, which was formed by this fusion is cultured for one (1) to ten (10) days in selection medium such as the medium containing hypoxanthine, thymidine and aminopterin (HAT medium) to isolate unfused cells. The obtained hybridoma is further selected based on the antibody to be produced. The selected hybridoma is isolated to a single clone by the publicly known limiting dilution. Thereby, a monoclonal antibody-producing hybridoma is established. For a method of detecting the activity of the antibody that is produced by the hybridoma, the publicly known method can be used. Herein, the activity of the antibody is detected in the following two steps: the binding ability to GPVI antigen as the first step and the activity of inhibiting the binding of GPVI and collagen as the second step. Examples of the detection method for the first step include ELISA, Western blotting, radioimmunoassay and the like. As the detection method for the second step, ELISA (inhibiting the binding), protein interaction analysis (BIACORE and so on), and a platelet aggregation suppression assay are given. In addition, the peptide of the eighth embodiment of the present invention and the polypeptide of the ninth embodiment can be used as an antigen for detecting the anti-GPVI antibody. The specific method is illustrated in EXAMPLES. The established hybridoma can be cultured by publicly known method, and from the culture supernatant, a monoclonal antibody can be obtained. The antibody can be purified using the publicly known purification means such as the salting-out method, gel filtration, ion exchange chromatography or affinity chromatography. The concentration of the antibody can be measured by the publicly known quantification method of protein, e.g. the measurement of absorbance at 280 nm absorbance. For a method of confirming the antigen binding property of the anti-GPVI antibody of the present invention or a method of detecting GPVI in the biological sample using the anti-GPVI antibody of the present invention, fluorescence antibody technique, enzyme linked immunosorbent assay (ELISA), radioimmunoassay (RIA), immunohistochemical method such as immunohistological staining and immunocytological staining (ABC method, the CSA method and so on), the 30 Western blotting method, immuno-precipitation method, enzyme-linked immunoassay as described above, sandwich ELISA method [Tan-kuron koutai jikken manual (The Monoclonal Antibody Experiment Manual) (Kodansha Scientific, 1987), Zoku seikagaku jikken kouza (The Continued Biochemical Experiment Course 5) Menneki-seikagaku kenkyuuhou (Immunobiochemical Research Method) (TOKYO KAGAKU DOZIN, 1986)] can be used. (Uses) The antibody of the present invention is an antibody that specifically binds to human GPVI. The antibody of the present invention, the active fragment of the antibody, the modified antibody that binds to the chemicals or the composition that comprises these mixtures have a variety of uses including prevention, diagnosis and treatment of the human diseases, and detection of human GPVI in test sample, cells, tissues and the like. Uses: Medicaments Since the antibody of the present invention has a high specificity for binding to GPVI, and solely has little activity that enhances or induces human platelet activation and/or thrombocytopenia, in particular, it is useful for prevention and/or treatment of human diseases, for example, diseases caused by activation or aggregation of platelet, or vascular endothelial disorders or arteriosclerotic reaction. In addition, it can be used for prevention and/or treatment of diseases caused by thrombus or embolus such as thrombosis, embolism and the like. Examples of these diseases include venous thrombosis as well as arterial thrombosis, or cerebral infarction caused by atrial fibrillation. Specific examples of human diseases or clinical conditions that are able to prevent and treat by the antibody of the present invention include myocardial infarction, vascular endothelial hypertrophy, restenosis of blood vessel, angina pectoris or myocardial infarction at the time of thrombolytic therapy, percutaneous transluminal coronary angioplasty (PTCA), stent operation, bypass surgery or artificial blood vessel operation, or thereafter; atrial fibrillation or atrial flutter, and thrombosis, embolism or cerebral infarction caused by these diseases; thromboangiitis obliterans; acute arterial occlusion; arteriosclerosis obliterans or deep venous thrombosis etc.; cerebral infarction (atheromatous thrombotic infarction, lacunar infarction, cardiogenic infarction); transient cerebral ischemic attack; cerebrovascular spasm after subarachnoid bleeding; pulmonary thrombosis; pulmonary embolism; vascular purpura; idiopathic thrombocytopenic purpura; thrombotic thrombocytopenic purpura; disseminated intravascular coagulation; prevention of blood coagulation at the time of extracorporeal circulation; systemic lupus erythematosus; multiple arteritis; anti-phospholipid antibody syndrome; purpura nephritis; endothelial cell injury associated with diabetes mellitis; diabetic nephritis; diabetic retinopathy; nephritic embolism; complications associated with transplantation (veno-occlusive disease of liver, graft-versus-host disease); and so on. 31 The antibody of the present invention can be administered to the diseases to be objective for the aforementioned prevention and/ treatment solely or in combination with other pharmacologically active ingredient. Examples of such pharmacologically active ingredient include publicly known thrombolytic agents such as tissue plasminogen activator(t-PA) and derivatives thereof (including mutant or so-called second generation); urokinase, streptokinase or publicly known platelet inhibitor (e.g. aspirin, ticlopidine, clopidogrel, thromboxane antagonist, thromboxane synthesis inhibitor, GPIIb/GPIIIa antagonist); publicly known anticoagulant (e.g. warfarin, heparin, low molecular weight heparin, pentasaccharide, thrombin inhibitor, FXa inhibitor, FVIIa inhibitor); and the like. Herein, the term "combination" includes the case where combined formulation comprising both the antibody of the present invention and the pharmacologically active ingredient is administered and the case where the antibody of the present invention and the pharmacologically active ingredient are administered as an independent formulation at the same time or time difference, and for the dosage form, no object as long as both exist simultaneously in the blood of a patient. A medicament comprising the antibody of the present invention and pharmaceutically acceptable composition as an effective ingredient can be prepared as tablet, injectable solution, powdered drug, suppository and the like using carrier for formulation, excipient and other additives used generally, and administered to human and other animals. When applied to the human, route for administration includes oral, intraveneous (bolus, continuous drip, intermittent drip), subcutaneous, intramuscular, intraarticular, transdermal, and transnasal administration. In general, oral administration or intraveneous administration may be used. Clinical dose of the antibody of the present invention to the human may appropriately be determined in consideration of conditions, body weight, ages, sex, etc. of the patient to be administered. In general, to adult, by intraveneous administration, dose of 1-10000 mg, preferably 10-1000 mg per day may be used, and the amount can be administered at one time or within several times. Since dosage varies depending on various conditions, there is a case where the amount less than the above-mentioned range is effectively administered. Herein, the antibody of the present invention includes various antibodies having different mechanisms meanwhile sharing recognition of GPVI in common. For example, since in the antibody that directly inhibits the binding of GPVI to collagen, or that suppresses activation and/or aggregation of platelet by cleaving GPVI, a relatively immediate effect can be expected, there have possibilities that the antibody is useful at least in the acute period of the disease (for example, in the time of myocardial infarction or PTCA implementation, or right before or after the events). In such a case, preferably, to make the antibody of the present invention bind to most of GPVI on the surface of platelet in the blood, relatively massive antibodies can be administered, e.g. by a single or divided intravenous injection or an intravenous drip. Also, in the antibody, in which GPVI is incorporated 32 internally, a continuous effect, but not an immediate effect can be expected considering a life-time of human platelet in the blood (about 9-10 days) and a half-life period of human antibody in the blood (in the case of IgG, several weeks). Thus, for example, there have a possibility that it is useful in chronic stage of diseases (several days to several months after development of myocardial infarction or PTC A implementation). In such a case, the antibody, whose amount is necessitated for depleting GPVI on the platelet surface to the extent of inhibiting a reactivity of platelet in the blood against collagen partially, preferably fully, can be administered at intervals of relatively long duration, e.g. from several days to several weeks per cycle, for example, by single dose or divisional intravenous injection, or the intravenous drip infusion. Therefore, in the preferred embodiment, the antibody of the present invention may possess these effects in parallel. In addition, a treatment, wherein multiple anti-GPVI antibodies that respective effects can be expected are combined, may be performed. A composition for parenteral administration generally includes a solution of the immunoglobulin dissolved into the acceptable carrier, preferably aqueous carrier or its mixture. Various aqueous carriers such as water, buffer solution, a phosphate buffered saline (PBS), 0.4% of saline, 0.3% of glycine, human albumin solution and the like can be used. These solutions are aseptic, and generally, microparticle material does not exist in these solutions. These compositions can be sterilized by the conventional and well-known method of sterilization. To approximate physiological conditions, the compositions may be included, on demand, pharmacologically acceptable auxiliary substance such as pH adjusting and buffering agent, toxicity regulating agent, and the like, specifically sodium acetate, sodium chloride, potassium chloride, calcium chloride and sodium lactate. The concentration of the antibody in these formulations can vary extensively, i.e. change from less than about 0.005% by weight (usually, at least about 1% by weight) to the large quantity, i.e. 15% or 20% by weight, and may be selected according to the selected and specific mode of administration, mainly based on the volume of the solution, the viscosity and the like. An actual method of preparing a parenteral administration composition is publicly known or obvious for persons skilled in the art, and further is described in detail in Remington's Pharmaceutical Sciences (15th Edition, Mack Publishing Company, Easton, Pennsylvania, 1980), wherein the reference is incorporated by reference in its entirety. A composition suitable for wash (lavage) or the other route is selected according to the intended specific use. Some pharmaceutical compositions can include an anti-GPVI antibody and the other treatment agent which is regularly used in the disease. In any cases, the bolus administration and the continuous administration can be applied. In addition, an effective amount for prevention or treatment is arbitrarily determined depending on object diseases, clinical condition and the condition of the patient and the like. 33 The antibody of the present invention may be frozen or lyophilized for storage, and prior to use, can be reconstituted in the suitable carrier. This technique is known to be effective in the conventional immunoglobulin. Further the publicly known techniques for lyophilization and reconstitution can be used. It is recognized for persons skilled in the art that lyophilization and reconstitution bring about activity loss of the antibody at various degrees (e.g. in the conventional immunoglobulin, IgM antibody, larger activity loss than that of IgG antibody tends to occur), and that use level may have to be regulated for compensation of loss. Use: Detection of GPVI A method of detecting GPVI in test sample using the antibody of the present invention or an active fragment thereof comprises the process, in which the test sample is brought in contact with the antibody of the present invention or an active fragment thereof, and the process, in which GPVI in the test sample bound to the antibody of the present invention or an active fragment thereof is detected. The process for quantifying GPVI in the test sample may further be comprised. Using the method of detecting GPVI in test sample, diseases can be diagnosed. In particular, it is possible to use for diagnosing human diseases such as thrombotic, embolic or arteriosclerosis diseases. Examples of the method of detecting GPVI in the test sample using the antibody of the present invention includes, but is limited to, the sandwich ELISA system, the inhibition-ELISA system, fluorescence antibody method, immunohistochemical staining method, radioisotope-labeled antibody method, Western blotting method, immunoprecipitation method and so on. As a target test sample, a biological sample is used, but is not limited. Examples of the sample include body fluid, tissues or cells from animal, particularly human, bacterial cells, and extracts thereof, culture supernatant, smears and sections, but preferred is platelet or plasma or serum. In addition, measurement of GPVI on the platelet can be applied to monitoring of therapy associated with GPVI, especially prediction or determination of the effect of anti-GPVI antibody, or prognostic determination using disappearance of GPVI on the platelet as an index. EXAMPLES By the following examples, the present invention will further be described in detail. However, the present invention should not be understood by the limitation of these examples. 34 EXAMPLE 1 Preparation of GPVI extracellular region-Fc fusion protein A. Preparation of human GPVI extracellular region-mouse Fc fusion protein (hGPVI-mFc) (1) Construction of the expression plasmid for hGPVI-mFc fusion protein Using mouse genomic DNA as a template, gene regions encoding respective domains of mouse immunoglobulin (mIgG2a) heavy chain constant region were amplified. That is, with the following primer pair, PCR reaction was performed. As a result, CHI domain, hinge region, CH2 domain and CH3 domain were amplified with mIgG2a-a (SEQ ID NO: 152) and mIgG2a-c (SEQ ID NO: 154), mIgG2a-b (SEQ ID NO: 153) and mIgG2a-e (SEQ ID NO: 156), mIgG2a-d (SEQ ID NO: 155) and mIgG2a-g (SEQ ID NO: 158), and mIgG2a-f (SEQ ID NO: 157) and mIgG2a-h (SEQ ID NO: 159), respectively. Then these four amplified products were mixed, and PCR reaction using primers mIgG2a-a and mIgG2a-h was performed to obtain an amplified product, wherein respective domains were ligated (DNA fragment encoding heavy chain constant region (Cy2a)). After the amplified product was cloned into pT7-BlueT vector, DNA fragment encoding mouse Fc region was excised with restriction enzymes Bam HI and Kpn I to obtain fragment A. On the other hand, from pCAGGS-GPVI-Fc plasmid, DNA fragment encoding an extracellular domain of human GPVI was excised with restriction enzymes Xba I and Bgl II to obtain fragment B. These fragments were ligated to the downstream of EF promoter in the expression vector pEF2cew, which was prepared by cleavage with Xba I and Kpn I, so that fragment A + fragment B was made. As a result, a plasmid (pTK-2249) that expresses hGPVI-mFc (SEQ ID NO: 222) was constructed. (2) Expression and purification of hGPVI-mFc fusion protein Cos-1 cells were cultured with Dulbecco's MEM medium supplemented with 10% fetal bovine serum. Transfection was performed by mixing an appropriate amount of pTK-2249 with transfection reagent (FuGENE6, Roche Diagnostics), dropping the above mixture into serum-free Dulbecco's MEM medium, and replacing this with culture fluid. Under condition of 5% CO2, the cells were cultured at 37°C for three days. The culture supernatant was subjected to the purification through Protein A column (Prosep-A, MILLIPORE), and the purified preparation was used as an antigen for preparation of anti-GPVI antibody. B. Construction and expression of expression plasmid pTK-2233 for human GPVI extracellular region-human Fc fusion protein Fragment J that is obtained by cleaving the plasmid pCAGGS-GPVI-Fc harboring a gene encoding human GPVI-hFc with restriction enzymes Xba I and Eco T22I, and fragment K that is obtained by cleaving it with restriction enzymes Eco T22I and Bgl II was prepared respectively. By ligating the two fragments to the downstream of EF-la promoter on the expression vector pEF2cew to make fragments J+K, a 35 hGPVI-hFc expressing plasmid pTK-2233 was constructed. In addition, the expression and purification of hGPVI-hFc was performed in a similar manner of the case for hGPVI-mFc. C. Construction of expression plasmid pTK-2440 for mouse GPVI extracellular region-human Fc fusion protein (mGPVI-hFc) Using mouse genomic DNA as a template and primer pairs listed, PCR reaction was performed. As a result, the PCR amplified product 'hi' was obtained with mGPVI-h (SEQ ID NO: 162) and mGPVI-i (SEQ ID NO: 163); the PCR amplified product 'jk' with mGPVI-j (SEQ ID NO: 164) and mGPVI-k (SEQ ID NO: 165); the PCR amplified product 'lm' with mGPVI-1 (SEQ ID NO: 166) and mGPVI-m (SEQ ID NO: 167); the PCR amplified product 'no' with mGPVI-n (SEQ ID NO: 168) and mGPVI-o (SEQ ID NO: 169); and the PCR amplified product 'pc' with mGPVI-p (SEQ ID NO: 170) and mGPVI-c (SEQ ID NO: 160). Using a mixture of these amplified products as a template and a mixture of primers mGPVI-e (SEQ ID NO: 161), mGPVI-q (SEQ ID NO: 171), mGPVI-r (SEQ ID NO: 172), mGPVI-s (SEQ ID NO: 173) and mGPVI-c, PCR reaction was performed to obtain an amplified product, wherein each fragment is ligated. This amplified product was cloned into pT7-BlueT vector and designated pTK-2437. Since in the gene region including mouse GPVI extracellular domain in pTK-2437, there have Nhe I restriction enzyme recognition site at 5' side and Bam HI restriction enzyme recognition site at 3' side, DNA fragment was prepared by cleavage with these enzymes. Then the fragment was inserted into pTK-2233, which is prepared by cleaving with restriction enzymes Xba I and Bam HI, to construct a mouse mGPVI-Fc expressing plasmid, pTK-2440. D. Construction of expression plasmid for cynomolgus monkey GPVI extracellular region-human Fc fusion protein (1) Construction of expression plasmid for cynomolgus monkey D1D2-human D3 chimeric GPVI-human Fc fusion protein Based on the information of human GPVI gene that is a known sequence, an appropriate primer pairs were designed and prepared. Then, using these primers and cynomolgus monkey genomic DNA as a template, PCR was performed to determine a part of gene sequence encoding cynomolgus monkey GPVI. Then, based on the obtained sequence, novel primer pairs for cynomolgus monkey were designed and prepared. Using each primer pairs and cynomolgus monkey genomic DNA as a template, PCR reaction was performed again. As a result, the amplified product 'ab' was obtained with macGPVI-a (SEQ ID NO: 174) and macGPVI-b (SEQ ID NO: 175); the amplified product 'dc' with hGPVI-d (SEQ ID NO: 180) and macGPVI-c (SEQ ID NO: 176); the amplified product 'dh' with macGPVI-d (SEQ ID NO: 177) and hGPVI-h (SEQ ID NO: 182) and the amplified product 'gg' with hGPVI-g (SEQ ID NO: 181) and macGPVI-g (SEQ ID NO: 178). MeanwhUe, using pTK-2233 as a template and macGPVI-h (SEQ ID NO: 179) and IgGl-i (SEQ ID NO: 183), PCR reaction was performed to obtain the amplified product 'hi'. Using a mixture of the above-mentioned five amplified products as a template, PCR 36 reaction with macGPVI-a and IgGl-i was performed once again. The amplified product obtained by this procedure includes a chimeric GPVI gene, in which cynomolgus monkey GPVI Dl and D2 and human D3 are fused. After cleavage with restriction enzymes Nhe I and Bam HI, the amplified product was inserted into pTK-2233 prepared by cleavage with restriction enzymes Xba I and Bam HI to construct an expression plasmid, pTK-2462 for cynomolgus monkey D1D2-human D3 chimeric GPVI-human Fc fusion protein (GPVI-FFH-hFc; SEQ ID NO: 223). EXAMPLE 2 Preparation of anti-GPVI antibody A. Preparation of rabbit polyclonal antibody To prepare a polyclonal antibody to human GPVI, rabbit was immunized. That is, 20 mg of hGPVI-mFc prepared in EXAMPLE 1A was diluted with 500 ml of saline mixed with 500 ml of Freund complete adjuvant (DIFCO). And the mixture was subcutaneously administered to the dorsal of female New Zealand white rabbit (KITAYAMA LABES) whose weight was 2.1-2.2 kg. Two weeks later, once again, 20 mg of hGPVI-mFc was diluted with 500 ml of saline and mixed with 500 ml of Freund incomplete adjuvant (DIFCO). Then, the mixture was subcutaneously administered to the dorsal. One week after the administration, the blood was collected from the ear vein, and according to the conventional method, anti-serum was prepared to purify an antibody. That is, to the anti-serum ammonium sulfate was added to make the final concentrations of saturated solution 33%. After stirring at 4°C for one hour, the precipitate was isolated by centrifugation. Next, the precipitate was dissolved in the Dulbecco's phosphate buffered saline (hereinafter, referred to as D-PBS) and dialyzed against D-PBS for overnight. After filtration of the dialysate, it was applied to Protein A column (Prosep A, Millipore) to get a purified antibody by eluting a bound IgG fraction with 0.1-M Glycine-hydrochloride buffer solution (pH3.0). The obtained eluate fraction was immediately neutralized in 1 M Tris-HCl buffer (pH7.0) and dialyzed against D-PBS. After dialysis, protein concentration was calculated from the absorbance at 280 nm (The absorbance coefficient: 0.714 mg/mL). Hereinafter, the obtained antibody is denoted as the anti-GPVI polyclonal antibody. B. Preparation of an anti-human GPVI monoclonal antibody By mixing 20 mg of GPVI-mFc and Freund complete adjuvant (DIFCO) at equal volume, an antigen for immunization was prepared. The antigen was administered twice to female ddY mouse (8-week age, SLC), and 3 days later, lymphocyte was isolated from the lymph node. The obtained lymphocyte was mixed with P3 x 63-Ag. 8. Ul (ATTC), and according to "The Introduction to Monoclonal Antibody Experimental Procedure" written by Tamie Andoh and Takeshi Chiba (Kodansha, p83), using polyethylene glycol (PEG1500, Sigma) a cell fusion was performed. Hybridoma was selected by HAT medium and one week later, the screening of the hybridoma that produces a desired antibody was performed by two ways. That is, a method of using a binding activity to hGPVI-hFc immobilized to a plate as an index, and a method of using an inhibiting activity for binding of collagen with GPVI as an index were utilized. 37 (1) Screening of hybridoma using the binding activity as an index The hGPVI-hFc prepared by EXAMPLE 1B was diluted with D-PBS to 2 mg/mL and added to immunoplate (Maxisorp J NUNC) at 50 mL/well. After incubation at 37°C for one hour, the well was washed five times with ion-exchanged water, and D-PBS (pH7.4) containing 2% StabilGuard (Surmodics) was added to each well in 100 mL for blocking. Next, culture supernatant was added to each well and incubated at 37°C for one hour. After incubation, the well was washed three times with the saline containing 0.05% Tween20. Peroxidase-labeled anti-mouse immunoglobulin antibody (DAKO, P260) was 1000-fold diluted with D-PBS containing 10% rabbit serum and added to each well at 50mL. After incubation at 37°C for one hour, washing in a similar manner was performed five times and TMB solution (BioFix) was added to each well. After ten minutes at room temperature, the reaction was terminated with 0.5 M sulfuric acid solution. Subsequently, absorbance at 450 nm was measured with the plate spectrophotometer (Multiscan JX, Dainippon Pharmaceutical). As a result, the cell whose culture supernatant reacted to hGPVI-hFc was selected and a cloning with limiting dilution method ("The Introduction to Monoclonal Antibody Experimental Procedure" written by Tamie Andoh and Takeshi Chiba (Kodansha, p97)) was performed. Eight days later, screening was done in a similar manner and the antibody which reacts to hGPVI-hFc was selected. (2) Screening of hybridoma using an inhibiting activity for binding of collagen with GPVI as an index Collagen (Horn) was diluted with D-PBS to 10 mg/mL and added to immunoplate (Maxisorp, NUNC) at 50 ml/well. After incubation at 4°C for overnight, the well was washed five times with ion-exchanged water and blocked with D-PBS containing 5% BSA. Next, culture supernatant was added to each well at 25 mL/well. Further, hGPVI-hFc prepared to 2 mg/mL with D-PBS was added to the well at 25 mL/well. After incubation at 37°C for one hour, the well was washed three times with the saline containing 0.05% Tween20. Peroxidase-labeled anti-human IgG antibody (BioMeda) was 1000-fold diluted with D-PBS containing 10% goat serums and added to each well at 50 mL. After incubation at 37°C for one hour, washing in a similar manner was performed five times and TMB solution (BioFix) was added to each well. After ten minutes at room temperature, the reaction was terminated with 0.5 M sulfuric acid solution. Subsequently, absorbance at 450 nm was measured with the plate spectrophotometer (Multiscan JX, Dainippon Pharmaceutical). The well, wherein the absorbance decreased by equal to or more than 50% in comparison with the well without the antibody, was selected as a hybridoma that produces an anti-GPVI antibody. As a result, by performing the multiple cell fusion (the F number shows 1 batch), hybridoma having an inhibiting activity for the binding of collagen with GPVI was selected. (3) Preparation of antibody produced by hybridoma 38 Hybridoma that produces an anti-GPVI antibody was cultured in 10% FCS/RPMI-1640 medium (Sigma). After replacement of the medium with Hybridoma-SFM medium (Invitrogen), hybridoma was further cultured to produce the antibody. From the culture supernatant, the antibody was purified using Protein A column (Prosep-rA, Millipore). That is, the obtained culture supernatant was subjected to Protein A column (Prosep-A, Millipore) pre-equilibrated with D-PBS. After washing non-adsorbed protein with D-PBS, adsorbed fraction was eluted with 25 mM Glycine-hydrochloride buffer (pH3.0). Thereafter, the fraction was dialyzed against saline. Concentration of the obtained antibody was calculated from absorbance at 280 nm using an absorption coefficient (E1%: 1.4). Hereinafter, the obtained antibody is referred to as the anti-GPVI monoclonal antibody. EXAMPLE 3 Group classification of the anti-GPVI monoclonal antibody In order to classify the antibody obtained in EXAMPLE 2 by the binding property to GPVI, i.e. the difference of the binding region, using F1199, F1201, F1202, F1210 and F1211 antibodies, respectively, competitive assay was performed. Firstly, according to the method described by Nakane et al. (J. Histochem. Cytochem., 22,1084,1974), each peroxidase-labeled anti-GPVI monoclonal antibody was prepared. Then, using the peroxidase-labeled antibody, competitive assay for respective purified antibodies was carried out to classify them. That is, hGPVI-hFc was diluted with D-PBS to 2 mg/mL, and the diluted solution was added to immunoplate (Maxisorp, NUNC) at 50 mL/well. After reaction at 37°C for one hour, the well was washed five times with ion-exchanged water, and D-PBS containing 2% StabilGuard was added to each well for blocking. Then, 25 mL each of the above-mentioned labeled antibody and the purified antibody were added to the well and incubated at 37°C for one hour. After washing five times with saline containing 0.05% Tween20, color development was done by TMB solution (BioFix). After reacting at room temperature for 10 minutes, the reaction was terminated with 0.5 M sulfuric acid solution. With plate spectrophotometer (Multiscan JX, Dainippon Pharmaceutical Co. Ltd.), absorbance at 450 nm was measured. The absorbance in the absence of a purified antibody was made 100%, and the inhibiting activity to each labelled antibody was calculated to classify each purified antibody. As a result, the one which competes with labeled antibodies (i), (ii), (v) and (viii) was designated group a. The one which competes with labelled antibodies (i), (v) and (vii) was designated group b. The one which competes with labelled antibodies (i), (ii), (vii) and (viii) was designated group c. Also, the one which competes with labeled antibodies (iii) and (iv) was designated group d. The one which competes with labeled antibodies (iii), (iv) and (vi) was designated group e. The one which competes with labeled antibodies (iii) and (vi) was designated group f. Also, the one whose regularity cannot be seen was designated groups g and h. Since the groups g and h did not compete with each other, they formed 39 another group. Therefore, antibodies are classified into eight groups in their recognition site of the surface of GPVI. EXAMPLE 4 Group classification of the antibodies used in the monkey ex vivo test Among the prepared anti-human GPVI monoclonal antibodies, antibodies that bind to monkey GPVI and are applicable for ex vivo test were classified according to the method for the group classification performed in EXAMPLE 3. That is, using each labeled antibody prepared in EXAMPLE 3, in a similar method to that of EXAMPLE 3, the competitive assay was done to classify each antibody. As shown in Table 1, just like EXAMPLE 3, the antibodies were classified into seven groups of the antibody that recognizes at least seven regions. Table 1 Group a F1232-10-1 Group b F1232-16-1 Group c F1232-39-3 F1232-21-1 F1232-13-3 Group d F1232-7-1 F1232-19-1 F1232-9-1F1232-37-2 F1232-11-1 F1201-18 Group e F1232-17-1 F1232-18-3 Group f F1232-8-3 F1232-38-1 F1232-14-2 F1232-45-1 F1232-15-1 F1232-29-2 F1232-24-1 Group g F1201-20 F1232-27-1 Group h F1232-43-3 EXAMPLE 5 Determination of dissociation constant for the anti-GPVI monoclonal antibody Dissociation constant for the anti-GPVI monoclonal antibodies prepared in EXAMPLE 2 was measured using protein interaction analyzer BIACORE3000 (BIACORE). That is, GPVI substitution mutant (hGPVIHHH-hFc and FFH-hFc) that is prepared in EXAMPLE 1 was coupled to sensor chip, CM5 chip (BIACORE), according to the manual. Then each antibody was diluted with HBS-EP buffer solution (BIACORE) to prepare a series of diluted solution from 1.25 to 40 nM, and analyzed with BIACORE3000. Every each antibody bound chip, the chip was regenerated with glycine buffer solution at pH1.5. The obtained result was analyzed by the evaluation software (BIACORE) with Bivalent analyte, to calculate dissociation constant. The result was shown in Table 2. It was shown that each anti-GPVI monoclonal antibody has a sufficient affinity to GPVI-HHH-hFc. 40 Table 2 Classification Antibody Dissociation Constant (K D) M GPVI GPVI Group a F1232-10-2 8.33x10-1° 1.09x10-9 Group c F1232-21-1 1.39X10-9 3.56x10-9 Group d F1232-7-1 F1232-19-1 F1232-37-2 F1201-18 3.47x10-8 1.75x10-9 4.03x10-10 1.16x10-9 3.43X10-7 2.35x10-9 1.00x10-9 6.19x10-10 Group e F1232-17-1 F1232-18-3 5.22x10-10 1.65x10-9 1.77x10-10 3.65x10-7 Group f F1232-14-2 F1232-24-1 1.56x10-8 2.5x10-10 2.14x10-8 7.1x10-10 Group g F1201-20 1.50x10-8 7.55x10-9 Group h F1232-43-3 1.36x10-9 1.69x10-9 EXAMPLE 6 Preparation of sandwich EIA system using anti-GPVI polyclonal antibody In order to prepare sandwich EIA system, the anti-GPVI polyclonal antibody obtained in EXAMPLE 2 was labeled with peroxidase just like EXAMPLE 3. Then a plate coated with anti-GPVI polyclonal antibody was prepared. That is, the antibody was diluted with D-PBS to 10 mg/mL and 50 mL of the antibody was added to each well of immunoplate (Maxisorp, NUNC) to incubate at 45°C for 30 minutes. Then, the well was washed five times with ion-exchanged water and to each well, 100 mL of D-PBS containing 2% StabilGuard (Surmodics) was added for blocking. As a standard sample, the purified GPVI-hFc was diluted with 0.1% BSA/D-PBS to 0.75 ng/ml, 1.5 ng/ml, 3.1 ng/ml, 6.25 ng/ml, 12.5 ng/ml, 25 ng/ml and 50 ng/ml. As a blank, D-PBS containing 0.1% BSA was used. At first, a blocking agent on the plate was discarded and 50 mL of the prepared standard sample and the blank was dispensed to each well for incubation at 25°C for overnight. The plate was washed three times with saline containing 0.05% Tween20. Subsequently, 50 mL of the peroxidase-labeled anti-GPVI polyclonal antibody that was diluted with D-PBS containing 10% rabbit serum and 0.1% Tween20 to 10 mg/ml was added to the well to incubate at 37°C. Similarly, after washing five times, TMB solution (BioFix) was added to each well to incubate at room temperature for 20 minutes, and the reaction was terminated with 0.5 M sulfuric acid solution. With the plate spectrophotometer (Multiscan JX, Dainippon Pharmaceutical), an absorbance at 450 nm was measured to prepare standard curve. 41 EXAMPLE 7 Analysis for recognition region of the GPVI antibody By replacing a part of the amino acid sequence in human GPVI with the corresponding amino acid sequence of mouse GPVI and examining the change of the antibody reactivity, the recognition region of each anti-GPVI antibody was refined. That is, since an extracellular region of GPVI, the platelet membrane protein is composed of three domains, i.e. immunoglobulin-like regions 1 and 2 (sometimes referred to as domain 1 or Dl, and domain 2 or D2) and mucin-like domain (sometimes referred to as domain 3 or D3) (FIG 1), substitution mutants for each domain were prepared and a binding assay of anti-GPVI antibody for each substitution mutants was performed. In addition, with the purpose of further refinement of the anti-GPVI antibody recognition region, among the proteins registered to the Protein Databank (PDB), based on the information of human NK cell activating receptor Nkp46, Ig-like transcript 2 (ILT2), whose homology of the amino acid sequence to GPVI immunoglobulin-like region is relatively high, a modeling for human GPVI was conducted to deduce the regions in which amino acid mutations can be introduced (FIG 1). Then, expression plasmids for human GPVI mutants having amino acid substitutions in those loop regions with mouse GPVI were constructed, and the binding assay of the anti-GPVI antibodies to each mutants was performed. As a result, each domain and loop region of GPVI, which each anti-GPVI monoclonal antibody recognizes could be confirmed. In addition, in the specification, the domain or the amino acid substitution mutant of GPVI is sometimes merely referred to as GPVI substitution mutants. Table 3 Amino acid sequence of each loop in domain 1 and domain 2 of soluble human GPVI Loop No. Sequence No. Amino acid sequence Loop 1 1 GPLPKP Loop 2 2 PSSLVPLEKP Loop 3 3 PPGVDL Loop 4 4 SSSRYQDQ Loop 5 5 PAMKRSLAGR Loop 6 6 QNGSLWSLPSDQ Loop 7 7 VFAKPS Loop 8 8 AQPGPAVSSGGD Loop 9 9 TRYGFDQ Loop 10 10 KEGDPA Loop 11 11 ERWYR Loop 12 12 ITVTAAHS Loop 13 13 FSSRDPYL Loot? 14 14 ELVVTG 42 (1) Construction of expression plasmids for human-mouse GPVI domain substitution mutants Using hGPVI-hFc (hereinafter referred to as GPVI-HHH-hFc; SEQ ID NO: 135) expressing plasmid (pTK2233) and mGPVI-hFc (hereinafter referred to as GPVI-MMM-hFc; SEQ ID NO: 136) expression plasmid (pTK2440), which are prepared in EXAMPLE 1, as a template, an expressing plasmid for GPVI substitution mutant, wherein the domains of human GPVI and mouse GPVI are exchanged, was constructed. That is, to construct an expression plasmid for GPVI substitution mutant comprising mouse GPVI domain 1 and human domains 2 and 3 (hereinafter referred to as GPVI-MHH-hFc; SEQ ID NO: 137), using pTK2440 as a template and sense primer 1 that is designed at the position upstream mouse GPVI sequence (Table 4; SEQ ID NO: 184) and anti-sense primer 3, wherein 15mer of the N-terminus of domain 2 in human GPVI is ligated to 15mer of the C-terminus of domain 1 in mouse GPVI (Table 4; SEQ ID NO: 186), PCR was performed to amplify DNA fragment of domain 1 in mouse GPVI (417 bp). Next, using pTK2233 as a template and anti-sense primer 2 that is designed at the position on hFc sequence (Table 4; SEQ ID NO: 185) and sense primer 4, wherein 15mer of the N-terminus of domain 2 in human GPVI is ligated to 15mer of the C-terminus of domain 1 in mouse GPVI (Table 4; SEQ ID NO: 187), PCR was performed to amplify DNA fragment (571 bp) comprising human GPVI domain 2, domain 3 and the N-terminus of hFc. Then, by performing PCR using the amplified two DNA fragments, sense primer 1, anti-sense primer 2, DNA fragment (973 bp), wherein the domain 1 of mouse GPVI, the domain 2 and domain 3 of human GPVI, and the N-terminal sequence of hFc, was amplified. After cutting this amplified DNA fragment with restriction enzymes Xbal and BamHI, the fragment was inserted into Xbal-BamHI site of pTK2233 to construct GPVI-MHH-hFc expressing plasmid. With respect to the expression plasmids for GPVI substitution mutants comprising sequences of human domain 1, mouse domain 2 and human domain 3 in GPVI (hereinafter represented by GPVI-HMH-hFc; SEQ ID NO: 138) and GPVI substitution mutant comprising sequences of mouse domain 1 and domain 2, and human domain 3 in GPVI (hereinafter represented by GPVI-MMH-hFc; SEQ ID NO: 139), in a similar manner, using sense and anti-sense primers (Table 4; SEQ ID NOs: 191 and 190, respectively) that are prepared by ligating 15mer each of human and mouse GPVI sequences at the joint position of the domain in GPVI desired to replace, sense primer 1 and anti-sense primer 2, needed DNA fragment was amplified. After cleaving with restriction enzymes Xbal and BamHI, by inserting the fragment into the Xbal-BamHI site of pTK2233, the expression plasmid was constructed. In addition, the amino acid sequences of five GPVI mutants were shown in SEQ ID NOs: 141-151. 43 (2) Construction of expression plasmids for GPVI substitution mutants, wherein a loop region of human GPVI is replaced with the corresponding amino acid sequence derived from mouse Expression plasmids for GPVI substitution mutants, wherein a single loop region to be a recognition region of an anti-GPVI monoclonal antibody is replaced with the corresponding amino acid sequence of mouse GPVI, was constructed as follows. First, in the loop region L2 of human GPVI, the base sequence was replaced so that the amino acid sequence from human was substituted for that from mouse. Further, sense primer 10, wherein 11mer of human GPVI base sequence was ligated upstream from the replaced base (Table 4) and anti-sense primer 9, wherein 13mer of human GPVI base sequence was ligated downstream from the replaced base (Table 4) were prepared. Next, by performing PCR using pTK2233 as a template, and sense primer 1 and anti-sense primer 9, DNA fragment (215 bp) corresponding to the N-terminus of the human GPVI, wherein the loop region L2 is substituted for the sequence from mouse, was amplified. In a similar manner, using pTK2233 as a template, and sense primer 10 and anti-sense primer 2, PCR was performed to amplify DNA fragment (773 bp), wherein the N-terminal sequence of hFc is connected to the C-terminal region of human GPVI, in which the loop region L2 is substituted for the sequence from mouse. Then, by performing PCR using the amplified two DNA fragments, sense primer 1, and anti-sense primer 2, DNA fragment (958 bp), wherein the N-terminal sequence of hFc is connected to the C-terminal region of human GPVI, in which the loop region L2 is substituted for the sequence from mouse, was amplified. After cutting this amplified DNA fragment with restriction enzymes Xbal and BamHI, the fragment was inserted into Xbal-BamHI site of pTK2233 to construct an expressing plasmid for GPVI substitution mutant, wherein the region L2 of human GPVI is replaced with the amino acid sequence of mouse GPVI (hereinafter represented by hGPVI-mL2-hFc; SEQ ID NO: 46). Expression plasmid for GPVI substitution mutants, wherein other loop regions are replaced, such as hGPVI-mL3-hFc, hGPVI-mL4-hFc, hGPVI-mL5-hFc, hGPVI-mL6-hFc, hGPVI-mL7-hFc, hGPVI-mL8-hFc, hGPVI-mL9-hFc, hGPVI-mL10-hFc, hGPVI-mLll-hFc, hGPVI-mL13-hFc, hGPVI-mL14-hFc, were constructed in a similar manner. The primer sequences that are used for construction of each GPVI substitution mutant-expressing plasmid were shown in Table 4. In addition, the amino acid sequences of the GPVI substitution mutants prepared, was shown by SEQ ID NOs: 47-57. 44 45 Table 4 Primer sets and PCR-amplified regions for the construction of expression plasmid of each domain-substituted or loop-substituted GPVI mutant 3) Preparation of GPVI substitution mutants The GPVI substitution mutant-expressing plasmid constructed in EXAMPLE 7 (1) and (2), pTK2233 and pTK2440 were introduced into the COS-1 cell and expressed in a similar manner shown in EXAMPLE 1. From culture supernatants obtained, the desired GPVI substitution mutants were purified by the Protein A column (Prosep-A, Millipore). Purity of GPVI substitution mutants obtained was confirmed by SDS-PAGE under both reducing and non-reducing conditions, and silver staining. (4) Binding activity with human-mouse GPVI domain substitution mutant By replacing the domain of human GPVI with the corresponding domain of mouse GPVI, the recognition region of the antibody was refined. That is, the binding activity of GPVI-HHH-hFc, GPVI-MHH-hFc, GPVI-HMH-hFc, GPVI-MMH-hFc and GPVI-MMM-hFc with each anti-GPVI monoclonal antibody was assayed. First, rabbit anti-human IgG antibody (DAKO) was immobilized to the plate (Maxisorp, Nunc) at 5 mg/mL, and the plate was blocked with 2% StabiliGuard (SurModics). Next, after removing blocking solution, the GPVI substitution mutant diluted with 0.1% BSA/PBS was added to react at 37°C for one hour. After washing with 0.9% saline containing 0.05% Tween20, blocking with 1% human serum (Cosmo Bio) diluted with 0.1% BSA/PBS was done at 37°C for one hour. Once again, after washing with 0.9% saline containing 0.05% Tween20, the peroxidase-labeled antibody prepared in EXAMPLE 3 was diluted to 0.5 mg/mL with 0.1% BSA/PBS, and added to the plate to incubate at 37°C for one hour. Finally, after washing with 0.9% saline containing 0.05% Tween20, the reaction mixture was developed using H2O2/TMB solution. The reaction was terminated with 0.5 M sulfuric acid, and an absorbance at 450 nm of wavelength was measured. As a result, in F1232-10-2, F1232-21-1, F1201-20, F1232-43-3 and F1232-14-2 antibodies, the binding activity to GPVI-MHH-hFc, GPVI-MMH-hFc and GPVI-MMM-hFc, wherein domain 1 was substituted for mouse GPVI, remarkably reduced. From the fact, it became clear that the recognition region of these antibodies existed in the domain 1 of GPVI. On the other hand, in F1232-7-1, F1232-19-1, F1232-37-2, F1232-17-1, F1232-18-3 and F1232-24-1 antibodies, the binding activity to GPVI-HMH-hFc, GPVI-MMH-hFc, GPVI-MMM-hFc, wherein domain 2 was substituted for mouse GPVI, declined remarkably. It was supposed that the recognition region of these antibodies existed in the domain 2 of GPVI. The antigen recognition region of each anti-GPVI antibody was shown in Table 5. 46 (5) Binding activity with GPVI substitution mutant, wherein the loop region in human GPVI was replaced with the corresponding amino acid sequence derived from mouse The binding activity of GPVI-HHH-hFc, hGPVI-mL2-hFc, hGPVI-mL3-hFc, hGPVI-mL4-hFc, hGPVI-mL5-hFc, hGPVI-mL6-hFc, hGPVI-mL7-hFc, hGPVI-mL8-hFc, hGPVI-mL9-hFc, hGPVI-mL10-hFc, hGPVI-mLll-hFc, hGPVI-mL13-hFc and hGPVI-mL14-hFc with each anti-GPVI monoclonal antibody was measured. By comparing the binding activity to GVI substitution mutant, wherein the loop region in human GPVI was replaced with the loop region from mouse, with that to human GPVI, the recognition region of the antibody was refined to the loop region replaced. In a similar manner as described in EXAMPLE 7 (4), peroxidase-labeled antibody prepared in EXAMPLE 3 was diluted to a measurable concentration with 0.1% BSA/PBS in conformity with the affinity of each antibody to GPVI-HHH-hFc, and used for the measurement. As a result, in F1232-10-2, F1232-21-1 and F1232-43-3 antibodies, the binding activity to hGPVI-mL2-hFc remarkably decreased. In F1232-14-2 antibody, the activity to hGPVI-mL3-hFc and hGPVI-mL5-hFc declined. In F1201-20 antibody, the activity to hGPVI-mL4-hFc and hGPVI-mL5-hFc declined. Also, in F1232-7-1, F1232-37-2, F1232-17-1, F1232-18-3 and F1232-24-1 antibodies, the activity to hGPVI-mL9-hFc and in F1232-19-1 antibody, the activity to hGPVI-mL9-hFc and hGPVI-mLll-hFc reduced, respectively. The antigen recognition region of each anti-GPVI antibody was shown in Table 5. Table 5 • D maens a case of ressuts that the antibody binding with a loop-substituted hGFVi is 30% or less of binding with hGFVi-hFc. 47 Table 6: Results of ex vivo test Thronbocytopenia : - ; 20%>, ± ;20%D 40D ,+;40%D 60%, ++; 60% Platelet activation : -; 2-fold, ± ;2-foldD 5-fold, +; 5-foldD 10-fold, ++; 10-fold Depletion of platelet membrane protein (FACS): -; 30%>(lmg/kg administration), ± ; 30%D 70% (lmg/kg administration), +; 70%(lmg/kg administration), ++; 70%(lmg/kg administration), +; 70% 48 t—» EXAMPLE 8 Evaluation of anti-GPVI monoclonal antibody by ex vivo experiment using cynomolgus monkey Each anti-GPVI monoclonal antibody prepared in EXAMPLE 2 was intravenously administered to male cynomolgus monkey (about 6 kg) at intervals of 24 hours in the dose of 0.3 mg/kg and 1 mg/kg. Prior to administration, 24 hours and 48 hours after the first administration, blood was collected, and number of platelet, expression level of CD62P protein (marker for platelet activation), expression level of platelet membrane protein (GPIIb/IIIa; CD41a and GPVI; CD42a), expression level of platelet GPVI and platelet aggregability (the reactivity to collagen and ADP) were assayed. A. Measurement of number of platelet Number of platelet of citrated blood was counted using Sysmex F-820. In the monkeys, to which clones F1232-18-3, F1232-10-2, F1232-14-2, F1232-43-3, F1201-20 and F1232-37-2 were administered respectively, no decrease in platelet number was observed (the decreasing rate was less than 20%; in Table 6, represented by symbol "-"). Further, in the monkeys, to which F1201-18 and F1232-17-1 were administered, downward trend of platelet number was observed (the decreasing rate was 20% to 40%; in Table 6, represented by symbol "±"). Meanwhile, in the monkeys, to which F1232-7-1, F1232-24-1 and F1232-21-1 were administered, apparent decreasing of platelet counts was observed (the decreasing rate was 40% to 60%; in Table 6, represented by symbol "+"), especially the decreasing was remarkable in the case of F1232-19-1 (the decreasing rate was equal to or more than 60%; in Table 6, represented by symbol"++"). B. Detection of CD62P protein on platelet Citrated blood was centrifuged at 100 x g at 25°C for 20 minutes to prepare platelet rich plasma (PRP). After PRP was diluted with PBS containing 0.5% heat-inactivated FBS and 2.5 mM EDTA (hereinafter referred to as FACS buffer) to make platelet number of PRP 1 x 108 cells/mL, using anti-human CD62P-PE (BD Biosciences Pharmingen), expression of CD62P protein on the platelet from monkey was analyzed by FACS. That is, anti-human CD62P-PE was added to PRP and the mixture was stood at room temperature for 30 minutes. After that, the platelet was washed with FACS buffer, and fluorescent intensity of the platelet was measured by flow cytometer CYTOMICS FC500 (BECKMAN COLETER). As a result, in any PRP prepared from the monkeys, to which anti-GPVI monoclonal antibody was administered, no increasing of expression of CD62P was observed in comparison with the level before administration (induction of expression was less than twofold; in Table 6, represented by symbol "-"). 49 C. Detection of CD41a and CD42a proteins on platelet A measurement of the expression level of CD41a and CD42a proteins on monkey platelet was performed by FACS analysis using anti-human CD41a-FITC (BD Biosciences Pharmingen) and anti-human CD42a-PE (BD Biosciences Pharmingen) respectively after preparation of PRP in a similar manner to the case of CD62P. As a result, in the PRP prepared from the monkey, to which F1232-21-1 and F1232-19-1 have been administered, a slight decrease in CD41a and CD42a proteins was observed (the depletion rate after administration of 1 mg/kg was 30% to 70%; in Table 6, represented by symbol "±"). However, in the PRP prepared from the monkey, to which other anti-GPVI monoclonal antibodies have been administered, no effect on the expression of CD41a and CD42a proteins was observed (the depletion rate after administration of 1 mg/kg was less than 30%; in Table 6, represented by symbol"-"). D. Detection of GPVI protein on platelet Confirmation of GPVI protein on monkey platelet was performed by FACS analysis using anti-GPVI polyclonal antibody labeled with fluorescent dye Af488 after preparation of PRP in a similar manner to the case of CD62P. As a result, in the PRP prepared from the monkey, to which F1201-18, F1201-20 and F1232-37-2 had been administered, depletion of GPVI protein was observed (the depletion rate after administration of 1 mg/kg was equal to or more than 70%; in Table 6, represented by symbol "++"). Particularly, in F1232-7-1, F1232-18-3, F1232-10-2, F1232-21-1, F1232-43-3, F1232-17-1 and F1232-19-1, depletion was remarkable (the depletion rate after administration of 0.3 mg/kg was equal to or more than 70%; in Table 6, represented by symbol "++"). Meanwhile, in the PRP prepared from the monkey, to which F1232-24-1 and F1232-14-2 have been administered, partial disappearance was observed (the depletion rate after administration of 1 mg/kg was 30% to 70%; in Table 6, represented by symbol "±"). E. Assay for platelet aggregability The platelet aggregation response to collagen or ADP was measured using platelet aggregation analyzer (PA-200 Aggregation Analyzer, Kowa). Firstly, after dilution of PRP with saline to make platelet number 3 x 108 cells/ mL, CaCk solution at the final concentration of 1 mM was added, and the mixture was incubated at 37°C for 3 minutes. Further, collagen solution at the final concentration of 2 mg/ml or ADP solution at the final concentration of 10 mM was added for incubation at 37°C for 12 minutes. The platelet aggregation rate was calculated by measuring light transmission with PA-200 Aggregation Analyzer (Kowa). As a result, in the PRP prepared from the monkey, to which F1232-7-1 and F1201-20 have been administered, decrease in collagen-responsive platelet aggregability was observed (the rate of loss after administration of 1 mg/kg was equal to or more than 70%; in Table 6, represented by symbol "+"). Particularly, in F1232-18-3, F1232-10-2, F1232-21-1, F1201-18, F1232-43-3, F1232-37-2, F1232-17-1 and F1232-19-1, the decrease was remarkable (the decreasing rate after administration of 0.3 mg/kg was equal to or more than 70%; in Table 6, represented by symbol "++"). Further, in the PRP prepared from the monkey, to which Fl 232-24-1 has been administered, partial 50

Documents:

1899-MUMNP-2007-ABSTRACT(GRANTED)-(21-3-2012).pdf

1899-mumnp-2007-abstract.doc

1899-mumnp-2007-abstract.pdf

1899-MUMNP-2007-CANCELLED PAGES(24-2-2012).pdf

1899-MUMNP-2007-CANCELLED PAGES(27-5-2011).pdf

1899-MUMNP-2007-CLAIMS(AMENDED)-(24-2-2012).pdf

1899-MUMNP-2007-CLAIMS(AMENDED)-(27-5-2011).pdf

1899-MUMNP-2007-CLAIMS(GRANTED)-(21-3-2012).pdf

1899-MUMNP-2007-CLAIMS(MARKED COPY)-(24-2-2012).pdf

1899-MUMNP-2007-CLAIMS(MARKED COPY)-(27-5-2011).pdf

1899-mumnp-2007-claims.doc

1899-mumnp-2007-claims.pdf

1899-mumnp-2007-correspondence(26-3-2008).pdf

1899-MUMNP-2007-CORRESPONDENCE(27-5-2011).pdf

1899-MUMNP-2007-CORRESPONDENCE(4-3-2009).pdf

1899-MUMNP-2007-CORRESPONDENCE(IPO)-(21-3-2012).pdf

1899-mumnp-2007-correspondence-others.pdf

1899-mumnp-2007-correspondence-received.pdf

1899-mumnp-2007-description (complete).pdf

1899-MUMNP-2007-DRAWING(GRANTED)-(21-3-2012).pdf

1899-mumnp-2007-drawings.pdf

1899-MUMNP-2007-EP DOCUMENT(24-2-2012).pdf

1899-mumnp-2007-form 1(13-11-2007).pdf

1899-MUMNP-2007-FORM 1(24-2-2012).pdf

1899-MUMNP-2007-FORM 1(27-5-2011).pdf

1899-mumnp-2007-form 1(4-2-2008).pdf

1899-mumnp-2007-form 13(4-2-2008).pdf

1899-MUMNP-2007-FORM 18(4-3-2009).pdf

1899-MUMNP-2007-FORM 2(GRANTED)-(21-3-2012).pdf

1899-MUMNP-2007-FORM 2(TITLE PAGE)-(24-2-2012).pdf

1899-MUMNP-2007-FORM 2(TITLE PAGE)-(27-5-2011).pdf

1899-MUMNP-2007-FORM 2(TITLE PAGE)-(GRANTED)-(21-3-2012).pdf

1899-MUMNP-2007-FORM 26(24-2-2012).pdf

1899-mumnp-2007-form 3(13-11-2007).pdf

1899-mumnp-2007-form 5(13-11-2007).pdf

1899-mumnp-2007-form-1.pdf

1899-mumnp-2007-form-2-1.doc

1899-mumnp-2007-form-2.doc

1899-mumnp-2007-form-2.pdf

1899-mumnp-2007-form-26.pdf

1899-mumnp-2007-form-3.pdf

1899-mumnp-2007-form-5.pdf

1899-mumnp-2007-form-pct-ib-304.pdf

1899-mumnp-2007-general power of attorney(13-11-2007).pdf

1899-MUMNP-2007-GENERAL POWER OF ATTORNEY(27-5-2011).pdf

1899-mumnp-2007-pct-search report.pdf

1899-MUMNP-2007-PETITION UNDER RULE 137(27-5-2011).pdf

1899-MUMNP-2007-REPLY TO EXAMINATION REPORT(24-2-2012).pdf

1899-MUMNP-2007-REPLY TO EXAMINATION REPORT(27-5-2011).pdf

1899-MUMNP-2007-SEQUENCE LISTING(27-5-2011).pdf

1899-MUMNP-2007-SEQUENCE LISTING(GRANTED)-(21-3-2012).pdf

abstract1.jpg