A Mechanistic Model for Paradoxical Platelet Activation by Ligand-Mimetic &agr;IIb&bgr;3 (GPIIb/IIIa) Antagonists

Objective—Integrins are attractive therapeutic targets. Inhibition of integrin &agr;IIb&bgr;3 effectively blocks platelet aggregation. However, limitations with intravenous &agr;IIb&bgr;3 antagonists and failure of oral &agr;IIb&bgr;3 antagonists prompted doubts on the current concept of ligand-mimetic integrin blockade. Methods and Results—Evaluating P-selectin expression on platelets by flow cytometry, we report a mechanism of paradoxical platelet activation by ligand-mimetic &agr;IIb&bgr;3 antagonists and define three requirements: (1) Induction of ligand-bound conformation of &agr;IIb&bgr;3, (2) receptor clustering, (3) prestimulation of platelets. Conformational change is inducible by clinically used ligand-mimetic &agr;IIb&bgr;3 antagonists, RGD-peptides, and anti-LIBS antibodies. In a mechanistic experimental model, clustering is achieved by crosslinking integrins via antibodies, and preactivation is induced by low-dose ADP. Finally, we demonstrate that platelet adhesion on collagen represents an in vivo correlate of platelet prestimulation and receptor clustering, in which the presence of ligand-mimetic &agr;IIb&bgr;3 antagonists results in platelet activation as detected by P-selectin, CD63, and CD40L expression as well as by measuring Ca2+-signaling. Blockade of the ADP receptor P2Y12 by AR-C69931MX and clopidogrel inhibits &agr;IIb&bgr;3 antagonist-induced platelet activation. Conclusion—These findings can explain limitations of ligand-mimetic anti-&agr;IIb&bgr;3 therapy. They describe potential benefits of concomitant ADP receptor blockade and support a shift in drug development from ligand-mimetic toward allosteric or activation-specific integrin antagonists.

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