RGD-containing peptides inhibit adhesion of 293 cells transfected with GpIIb/IIIa to fibrinogen: comparison to inhibition of platelet aggregation.
Cyclic RGD-containing peptides caused a dose-dependent inhibition of binding of human embryonic kidney cells transfected with recombinant GpIIb/IIIa (r293 clone B) to human fibrinogen coated on to non-tissue culture plates. The inhibitory activity, IC50, of a panel of seventeen RGD-containing peptides ranged from 0.12 to 89.2 microM. These IC50 values correlated with those determined by the inhibition of platelet aggregation (r = 0.99). Even though there was a correlation, there were differences between the platelet aggregation and the bioadhesion assay. The binding of r293 clone B to fibrinogen was not increased by ADP suggesting that GpIIb/IIIa expressed on the surface of r293 clone B cells may be in the 'activated' form. Moreover, preincubation of r293 clone B cells with a monoclonal antibody (mAb) specific for GpIIIa (4B12) resulted in a dose-dependent decrease of binding to fibrinogen while a mAb specific for GPIIb (2D2) had no effect. Neither of these mAbs inhibited platelet aggregation. The binding of r293 clone B cells to fibrinogen required Ca2+ or Mg2+. This cell-based bioadhesion method can provide a tool for screening potential GpIIb/IIIa antagonists and investigating the interaction of GpIIb/IIIa and fibrinogen not possible with platelet aggregation.
Department of Medicinal and Analytical Chemistry, Genentech Inc., South San Francisco, CA 94080.
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Fei
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Lowe
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Bodary
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Bunting
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McLean
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Napier
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Chen
D000244:Adenosine Diphosphate / Q000494:pharmacology; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D001681:Biological Assay; D002118:Calcium; D002448:Cell Adhesion / Q000187:drug effects*; D002999:Clone Cells; D004492:Edetic Acid / Q000494:pharmacology; D004533:Egtazic Acid / Q000494:pharmacology; D005340:Fibrinogen; D006801:Humans; D008274:Magnesium; D008954:Models, Biological; D009842:Oligopeptides / Q000494:pharmacology*; D010974:Platelet Aggregation / Q000187:drug effects; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology*; D010980:Platelet Membrane Glycoproteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology*; D011994:Recombinant Proteins / Q000187:drug effects; D014162:Transfection
D000911:Antibodies, Monoclonal; D009842:Oligopeptides; D010975:Platelet Aggregation Inhibitors; D010980:Platelet Membrane Glycoproteins; D011994:Recombinant Proteins; D004533:Egtazic Acid; D000244:Adenosine Diphosphate; C047981:arginyl-glycyl-aspartic acid; D005340:Fibrinogen; D004492:Edetic Acid; D008274:Magnesium; D002118:Calcium