Characterization of a Novel Function-Blocking Antibody Targeted Against the Platelet P2Y1Receptor

Objective— Platelet hyperactivity is associated with vascular disease and contributes to the genesis of thrombotic disorders. ADP plays an important role in platelet activation and activates platelets through 2 G-protein–coupled receptors, the Gq-coupled P2Y 1 receptor (P2Y 1 R), and the Gi-coupled P2Y 12 receptor. Although the involvement of the P2Y 1 R in thrombogenesis is well established, there are no antagonists that are currently available for clinical use. Approach and Results— Our goal is to determine whether a novel antibody targeting the ligand-binding domain, ie, second extracellular loop (EL2) of the P2Y 1 R (EL2Ab) could inhibit platelet function and protect against thrombogenesis. Our results revealed that the EL2Ab does indeed inhibit ADP-induced platelet aggregation, in a dose-dependent manner. Furthermore, EL2Ab was found to inhibit integrin GPIIb-IIIa activation, dense and α granule secretion, and phosphatidylserine exposure. These inhibitory effects translated into protection against thrombus formation, as evident by a prolonged time for occlusion in a FeCl 3 -induced thrombosis model, but this was accompanied by a prolonged tail bleeding time. We also observed a dose-dependent displacement of the radiolabeled P2Y 1 R antagonist [ 3 H]MRS2500 from its ligand-binding site by EL2Ab. Conclusions— Collectively, our findings demonstrate that EL2Ab binds to and exhibits P2Y 1 R-dependent function-blocking activity in the context of platelets. These results add further evidence for a role of the P2Y 1 R in thrombosis and validate the concept that targeting it is a relevant alternative or complement to current antiplatelet strategies.