Πεπτιδικοί αναστολείς του υποδοχέα των αιμοπεταλίων GPIIb/IIIa: μελέτη σε ασθενείς με καρδιαγγειακή νόσο

Abstract

Coagulation and platelets play major roles in many aspects of coronary artery disease. Platelet membrane glycoproteins are involved in platelet adhesion and aggregation. The GPIIb/IIIa receptor is the most numerous receptor found on the platelet surface and represents the final common pathway for platelet aggregation. Once activated, the receptor becomes available to bind fibrinogen and vWF, which act to crosslink platelets and promote the generation of a platelet mass at the site of vascular injury. The understanding of the mechanism of action of GPIIb/IIIa receptor inhibitors is required for appreciating the potential of novel therapies directed against GPIIb/IIIa receptors in patients with acute coronary syndromes. The aim of this study, carried out in the context of a project supported by the Greek General Secretariat of Research and Technology, was to further elucidate the role of the platelet GPIIb/IIIa receptor in platelet aggregation and thrombus formation in unstable angina and myocardial infarction. It was focused on the evaluation of the anti-aggregatory activity of synthetic peptides, modelled from GPIIb amino acid sequence, previously found to inhibit platelet aggregation in platelet rich plasma (PRP) from normal individuals, in PRP from patients with acute coronary syndromes. The inhibitory effect of the peptides was determined on ADP-induced platelet aggregation in PRP and measured on the basis of the light transmittance changes in an aggregometer. It was thus shown that the sequence GPIIb 313-332 efficiently inhibited by 60% platelet aggregation, at peptide concentrations of 500μΜ. This result suggests that the peptide GPIIb 313-332 is reactive against the conformationally active form of the receptor. Interestingly, it was previously demonstrated that this sequence is a potent inhibitor of platelet aggregation in PRP from normal individuals. Based on this findings monoclonal antibodies (mAbs) against this peptide were generated and characterized. The inhibitory activity of the mAbs was subsequently determined using the same experimental procedure. It was thus demonstrated that 6 out of the 9 mAbs inhibited platelet aggregation in PRP from normal individuals. In order to map the epitopes of the mAbs, competition experiments using overlapping octapeptides within the 313-332 region were performed. Binding of the mAb Φ1(15) was completely inhibited by the 313-320 octapeptide, suggesting that the critical epitope of this antibody is within this segment. Both the identified peptides and the generated antibodies will be important tools for exploring the receptor-ligand interactions and the design of potent GPIIb/IIIa antagonists.