A novel anti-platelet monoclonal antibody (3C7) specific for the complex of integrin alpha IIb beta3 inhibits platelet aggregation and adhesion.
Activation or ligand binding induces conformational changes in alpha IIb beta3, resulting in exposure of neoepitopes named ligand-induced binding sites. We reported here a novel monoclonal antibody developed by using Chinese hamster ovary (CHO) cells expressing an activated alpha IIb beta3 mutant (CHO alpha IIb beta3Delta717) as the immunogen. This IgG 2b kappa named 3C7 was specific for the complex of alpha IIb beta3 as demonstrated by flow cytometry, immunoprecipitation, and EDTA chelating. The binding of 3C7 to platelets increased significantly when platelets were activated by ADP/thrombin or occupied by RGDS peptides, fibrinogen, or PAC-1, suggesting that 3C7 was an anti-ligand-induced binding site antibody. The antibody failed to bind to the CHO cells expressing another alpha IIb beta3 mutant (beta3Y178A) suggesting that the Cys177-Cys184 loop of beta3 was likely the epitope for 3C7. 3C7 inhibited platelet aggregation, which was initiated by ADP or thrombin in a dose-dependent manner (IC50s of 5.6 and 0.05 microg/ml, respectively). The antibody also inhibited platelet adhesion to immobilized fibrinogen but not to fibronectin or collagen. These findings suggested that 3C7 was a potent antagonist of integrin alpha IIb beta3 and a potential anti-thrombotic agent.
Chen
Ping
P
Institute of Molecular Medicine and State Key Laboratory of Pharmaceutical Biotechnology, Nanjing University, 22 Hankou Road, Nanjing 210093, China.
Sun
Chong-Xiu
CX
Liu
Jian-Ning
JN
D000244:Adenosine Diphosphate / Q000494:pharmacology; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology* / Q000494:pharmacology; D000918:Antibody Specificity; D001792:Blood Platelets / Q000276:immunology* / Q000378:metabolism; D016466:CHO Cells; D006224:Cricetinae; D000939:Epitopes / Q000276:immunology; D005340:Fibrinogen / Q000378:metabolism; D006490:Hemostatics / Q000494:pharmacology; D051379:Mice; D008807:Mice, Inbred BALB C; D010973:Platelet Adhesiveness / Q000187:drug effects / Q000276:immunology*; D010974:Platelet Aggregation / Q000187:drug effects / Q000276:immunology*; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000276:immunology*; D013917:Thrombin / Q000494:pharmacology
D000911:Antibodies, Monoclonal; D000939:Epitopes; D006490:Hemostatics; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D000244:Adenosine Diphosphate; D005340:Fibrinogen; D013917:Thrombin