Epitopes and biological activities of two monoclonal antibodies to platelet integrin alpha IIb beta 3.
We have developed two monoclonal antibodies (MAbs), B6A3 and C4G1. The whole molecules of the two MAbs inhibited in vitro human platelet aggregation induced by either ADP, collagen or thrombin, and their F(ab')2 fragments inhibited ex vivo platelet aggregation induced by ADP in monkey. The concentrations necessary for complete inhibition were 5 and 1 microgram/ml for B6A3 and C4G1, respectively. The Fab fragment of C4G1 but not B6A3 inhibited platelet aggregation. B6A3 and C4G1 bound to activated platelets with dissociation constants of 0.25 and 0.82 nM, respectively. B6A3 recognized an epitope on beta 3, which was sensitive to reduction and alkylation of cystine residues, and C4G1 recognized a conformational epitope on the alpha IIb beta 3 complex, which was sensitive to EDTA. The binding of fibrinogen to activated platelets was inhibited by both MAbs. However, the binding of fibrinogen to isolated alpha IIb beta 3 was inhibited by the whole molecule of C4G1 but not B6A3, although both MAbs bound to the isolated alpha IIb beta 3. The binding of these MAbs to the isolated alpha IIb beta 3 was not inhibited by either Arg Gly Asp Ser (RGDS) or fibrinogen gamma-peptide. In addition, B6A3 but not C4G1 bound to human endothelial cells. These MAbs should contribute to the elucidation of the mechanism of platelet aggregation.
Yano
S
S
Molecular Medicine Research Laboratories, Yamanouchi Pharmaceutical Co., Ltd., Ibaraki.
Suzuki
K
K
Katoh
M
M
Sugita
Y
Y
Kaku
S
S
Kawamura
K
K
Masuho
Y
Y
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology* / Q000378:metabolism / Q000494:pharmacology*; D001792:Blood Platelets / Q000187:drug effects* / Q000378:metabolism / Q000502:physiology*; D015153:Blotting, Western; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000276:immunology*; D005340:Fibrinogen / Q000378:metabolism; D006801:Humans; D007114:Immunization; D007140:Immunoglobulin Fab Fragments / Q000494:pharmacology; D016023:Integrins / Q000276:immunology* / Q000378:metabolism; D008252:Macaca fascicularis; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D010974:Platelet Aggregation / Q000187:drug effects*; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D016024:Receptors, Cytoadhesin / Q000276:immunology; D013154:Spleen / Q000166:cytology / Q000276:immunology
D000911:Antibodies, Monoclonal; D000939:Epitopes; D007140:Immunoglobulin Fab Fragments; D016023:Integrins; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D016024:Receptors, Cytoadhesin; D005340:Fibrinogen