Received for publication March 3, 2006.
Revised July 7, 2006.
Accepted for publication July 11, 2006.

Prevention of platelet glycoprotein IIb/IIIa activation by a novel tyrosine kinase inhibitor 3,4-Methylenedioxy--nitrostyrene

Abstract

Binding fibrinogen to activated glycoprotein (GP) IIb/IIIa is the final common pathway of platelet aggregation, and has become a successful target for antiplatelet therapy. In the present study, we found that a small chemical compound, 3,4-methylenedioxy--nitrostyrene (MNS), exhibited potent and broad-spectrum inhibitory effects on human platelet aggregation caused by various stimulators. Moreover, addition of MNS to human platelets that had been aggregated by ADP caused a rapid disaggregation. We demonstrated that the anti-aggregatory activity of MNS is due to inhibition of GPIIb/IIIa activation by measuring the binding amount of PAC-1 in platelets. On the other hand, MNS is not a direct antagonist of GPIIb/IIIa, since MNS did not affect fibrinogen binding to fixed ADP-stimulated platelets. Investigating how MNS inhibits GPIIb/IIIa activation, we found that MNS potently inhibited the activity of tyrosine kinases (Src and Syk), prevented protein tyrosine phosphorylation, and cytoskeletal association of GPIIb/IIIa and talin, but had no direct effects on protein kinase C, Ca²⁺ mobilization, Ca²⁺-dependent enzymes (myosin light chain kinase and calpain) and arachidonic acid metabolism, and did not affect the cellular levels of cyclic nucleotides. Therefore, MNS represents a new class of tyrosine kinase inhibitor that potently prevents GPIIb/IIIa activation and platelet aggregation without directly affecting other signaling pathways required for platelet activation. Because MNS inhibits GPIIb/IIIa functions in a manner different from GPIIb/IIIa antagonists, this feature may provide a new strategy for treatment of platelet-dependent thrombosis.

Key words: Src and other nonreceptor tyrosine kinases, Platelets