Despite significant progress over the past several decades, coronary artery disease (CAD) continues to be the most important cause of mortality in this country. Formation of a platelet thrombus is a central event in myocardial infarction (MI) and unstable angina (USA), and this process requires the binding of adhesive ligands to the alphaIIb-beta3 (gIycoprotein IIb-IIIa) receptor on platelets. Platelet membrane glycoproteins, especially alphaIIb-beta3 are highly polymorphic, and our preliminary studies demonstrate a statistically significant association between the PlA2 polymorphism on beta3 and premature ischemic coronary events, particularly in young individuals. Because of the central role played by alphaIIb-beta3 in the development of myocardial ischemic events, the hypothesis driving our studies is that the PlA2 form of the receptor induces a conformational change making it hyper-reactive for one of its adhesive ligands. We will use in vitro platelet assays, cell and molecular biology, as well as genetic linkage analyses to test our hypothesis. The major goal of this proposal is to characterize functional differences between the PlA1 and PlA2 forms of integrin beta3 as it exists in the two important receptors, alphaIIb-beta3 and alpha-v- beta3. The major thrust of our studies will be to characterize differences in binding of platelets homozygous for PlA1 and PlA2 to the major alphaIIb-beta3 ligands, fibrinogen and von Willebrand factor, under static and flow conditions. By using inhibitors which interrupt pathways mediating different aspects of platelet physiology, mechanistic information regarding the intrinsic difference between A1 and A2 platelets can be dissected. Such experiments may also provide information with respect to the future use of anti-platelet therapy in A2 patients at high risk of acute CAD. Post-receptor occupancy signaling mechanisms, particularly as relates to receptor cross- linking, for PlA1 and PlA2 platelets will be studied. To address possible effects of the A2 polymorphism on the alpha-v-beta-3 expressed in endothelial and smooth muscle cells, stable cell lines will be generated and tested for their ligand binding characteristics. Finally, affected sibling pairs from families with premature coronary heart disease will be genotyped to perform genetic analyses to test for linkage between the PlA locus and acute CAD. These studies will enhance our understanding of the role of integrin biology (especially in platelets), and help to achieve our long-term goal of implementing a more rational anti-thrombotic therapy.
