A novel platelet inhibitor from bloodfeeding hookworms

描述(申请人提供)：血栓栓塞性疾病是发达国家发病率和死亡率的主要原因。由于血小板在血栓形成的激活和传播中起着关键作用，因此它们是新型抗血栓药物的重要靶点。从吸血钩虫中分离出一种有效的抑制人类血小板功能的物质。这种新的蛋白质是第一种天然存在的化合物，可以阻断血小板整合素GPIIb/IIIa和GPIa/IIa的结合，GPIIb/IIIa和GPIa/IIa分别是纤维蛋白原和胶原的细胞表面受体。这一阶段STTR资助申请的目标是提纯毫克量的活性重组钩虫血小板抑制物(RHPI)，适用于体内和体外测试。RHPI将通过全血小板聚集和黏附试验进行表征。此外，定量实验将测量RHPI阻断胶原和纤维蛋白原与其各自的整合素受体相互作用的能力。与各种多肽的竞争实验将描绘出HPI的结合部位，并进一步表征其作用的分子机制。最后，将产生RHPI的截断变体，以确定具有保留的抗血小板活性的最小蛋白结构域。长期目标是开发RHPI作为治疗人类血栓性疾病的药物。

DESCRIPTION (provided by applicant): Thromboembolic disease is a major cause of morbidity and mortality in the developed world. Because platelets play a critical role in the activation and propagation of thrombosis, they represent important targets for new anti-thrombotic drugs. A potent inhibitor of human platelet function has been isolated from the bloodfeeding hookworm, Ancylostoma caninum. This novel protein is the first naturally occurring compound shown to block the binding of platelet integrins GPIIb/IIIa and GPIa/IIa, the cell surface receptors for fibrinogen and collagen, respectively. The objective of this Phase I STTR grant application is to purify milligram quantities of active recombinant hookworm platelet inhibitor (rHPI) suitable for testing in vivo and in vitro. The rHPI will be characterized using whole platelet assays of aggregation and adhesion. In addition, quantitative experiments will measure the ability of rHPI to block the interaction of collagen and fibrinogen with their respective integrin receptors. Competition experiments with various peptides will delineate HPI binding sites and further characterize its molecular mechanism of action. Lastly, truncated variants of rHPI will be produced in order to determine the smallest protein domain with preserved antiplatelet activity. The long-term objective is to develop rHPI as a therapeutic agent for the treatment of thrombotic disorders in humans.