The platelet was discovered in 1842 by Donne [C. R. Acad. Sci.(paris), 14, 336-368, 1842], and has long been regarded as a component of blood necessary for hemostasis. It is now known that the platelet plays not only a main role in the hemostatic system, but also multifunctional roles, for example, a clinically notable generation of arteriosclerosis, circulatory diseases including thrombotic diseases, metastasis of cancer, inflammation, rejection after grafting, participation in immune reaction, or the like.
At present, revascularization by pharmacological or physical methods is carried out to treat thrombotic diseases and ischemic diseases. However, it has been recently found that the activation, adhesion, and/or aggregation of the platelets is promoted by a collapse of blood vessel tissue including an endothelial cell after revascularization, or a collapse of a fibrinolysis-coagulation balance caused by a medicament per se, which becomes a clinical problem. For example, it is known that, after revascularization by a thrombolytic therapy using t-PA or the like, the fibrinolytic activity and/or coagulative activity are activated, and then the systemic fibrinolysis-coagulation balance collapses. Clinically, this causes re-occlusion, and becomes a critical problem therapeutically (J. Am. Coll. Cardiol. 12, 616-623, 1988).
In addition, the PTCA (Percutaneous transluminal coronary angioplasty) therapy has quickly become widely used, and has achieved good results in the treatment of diseases based on aortostenosis or coronary stenosis such as angina, myocardial infarction, or the like. However, the therapy injures blood vessel tissue including an endothelial cell, and acute coronary obstruction, and restenosis, which is observed in approximately 30% of cases, become a problem.
The platelet plays an important role in these various thrombotic disorders (such as re-occlusion or the like) after the revascularization therapy. Therefore, an antiplatelet agent is desired as an agent for treating or preventing these thrombotic disorders.
In this connection, adenosine 5′-diphosphate (ADP) is known as an important factor which induces or promotes the activation, adhesion, and aggregation of the platelets. ADP is released from platelets activated by collagen, thrombin, or the like, or from hemocytes, vascular endothelial cells, or organs injured by revascularization or the like. It is considered that ADP activates the platelets via a G protein-coupled ADP receptor P2T located in the platelet membrane (Biochem. J., 336, 513-523, 1998).
It has been suggested that a platelet ADP receptor P2TPLC which is coupled to Gq, one of the G proteins, and increases an intracellular Ca2+ concentration via phospholipase C (PLC), and a platelet ADP receptor P2TAC which is coupled to Gi, one of the G proteins, and suppresses an activity of adenylate cyclase (AC) are present as platelet ADP receptors. At present, the platelet ADP receptor P2TPLC has been identified as the receptor known as platelet ADP receptor P2Y1, but the entity of platelet ADP receptor P2TAC is not identified (Kunapuli, S. P. et al., Trends Pharmacol. Sci., 19, 391-394, 1998).
It is considered that Ticlopidine or Clopidogrel used as an antiplatelet agent functions by inhibiting the ADP receptor P2TAC via its metabolite in a body (Savi, P. J., Pharmaclo. Exp. Ther., 269, 772-777, 1994). Further, ARL67085, which is synthesized as a derivative of adenosine triphosphate (ATP), which is an ADP receptor antagonist in a body, exhibits an activity of suppressing a platelet aggregation by the antagonist activity against the platelet ADP receptor P2TAC, and the effectiveness thereof is proven by using a thrombosis model (Mills, D. C., Thromb. Hemost., 76, 835-856, 1996; and Humphries, R. G., Trends Pharmacol. Sci., 16, 179-181, 1995). Further, a derivative of Ap4A [P1,P4-di(adenosine-5′)tetraphosphate] exhibits an activity suppressing the platelet aggregation by ADP, by the antagonist activity against the platelet ADP receptor P2TAC, and the effectiveness thereof is proven by using a thrombosis model (Kim, B. K., Proc. Natl. Acad. Sci. USA, 89, 2370-2373, 1992).
From the above information, an antagonist against the platelet ADP receptor P2TAC is desired as a strong antiplatelet agent. However, Ticlopidine or Clopidogrel exhibits a weak antiplatelet activity, and has problems such as a strong side effect or the like. Further, ARL67085 or derivatives thereof (ATP analogues), derivatives of Ap4A, or the like, which is studied as the ADP receptor antagonist, is a derivative of nucleotide, and then an oral bioavailability is not sufficient, and further problems arise such as a weak activity of suppressing the platelet aggregation. Therefore, an ADP receptor antagonist having a strong oral bioavailability is intensely desired (CAPRIE STEERING COMMITTEE, Lancet, 348, 1329-1339, 1996).
However, the ADP receptor P2TAC protein has not been identified as yet. Therefore, it is difficult to construct a convenient system for screening such a compound, and farther, the development of the ADP receptor P2TAC antagonist has not progressed.
In this connection, with regard to a DNA encoding a polypeptide consisting of the same amino acid sequence as that of a human ADP receptor P2TAC protein, which may be used in the present invention, and an amino acid sequence deduced from the DNA, there are several reports (WO00/22131, WO00/31258, WO00/28028, and WO98/50549 pamphlets). However, ligands are not elucidated in the reports, and no reports disclose that the protein is an ADP receptor located in the platelet.