DEVELOPING A TRANSLATABLE PLATFORM FOR REVERSIBLE PLATELET INHIBITION

原理和总体目标：我们的总体目标是开发一种快速可逆的抗血小板药物，抑制血栓形成的高度血栓形成设置，如在冠状动脉血运重建过程中遇到的。血小板在动脉粥样硬化斑块和其他血栓形成表面上的不受调节的积聚是血栓形成的绝对先决条件，血栓形成反过来代表了血栓形成的直接原因。 心肌梗死、设备故障、缺血性中风和心血管死亡的原因（1-3）。 抗血小板药物是血小板生物学基础知识迅速扩展的直接产物。现有的抗血小板治疗都不能阻止血栓形成的起始步骤-血小板粘附。因此，开发特异性工程化以抑制在血小板粘附中起作用的生物学相关靶标的新药代表了解决未满足的临床需求的重要一步。糖蛋白（GP）Ib-IX-V是属于血小板粘附受体的一种。 富含亮氨酸的重复蛋白质家族。其主要功能是在高剪切应力血流下引发血小板聚集，导致血栓形成（4）。在GP Ib-1X-V粘附到血管性血友病因子（VWF）后，血小板被活化，经历细胞骨架形状变化并分泌血小板激动剂，其募集额外的血小板到发展中的血栓”。沿着GP IIb/IIIa，GP Ib-IX-V是唯一的血小板受体 其在止血和血栓形成中具有非多余作用，止血缺陷不如GP IIb/IIIa缺陷严重（4-7）。因此，GP IIb-IX-V-VWF相互作用的抑制剂可能具有成为有效的抗血栓形成药物的潜力，其具有比GP IIb/IIIa抑制剂更有利的安全性。 抗血栓治疗用于管理血栓性疾病患者的总体有效性，特别是那些需要侵入性操作或手术的患者，在很大程度上受到这些治疗伴随的出血风险的限制。虽然责任机制界定不清，但同时 止血和血管修复受损被认为是可能的原因。我们先前已经证明，蛋白结合寡核苷酸（适体）、抑制血小板的单链核酸、它们的功能配体和/或具有高特异性的凝血蛋白可以容易地用互补寡核苷酸解毒剂逆转（8-10）。与该提议特别相关的是，我们已经开发了针对VWF的适体，其潜在地抑制小鼠中的血栓形成，以及解毒剂寡核苷酸，其可以快速逆转适体的抗血小板活性（11）（Sulfier实验室未发表的结果）。这种适体-解毒剂策略 潜在地提供了广泛的应用，随着抗血栓药物的开发，其可以被主动控制，在临床指示时允许充分的药效学作用，随后快速滴定或完全逆转。

Rationale and Overall Objective: Our overall goal is to develop a rapidly reversible antiplatelet agent that inhibits thrombus formation in highly prothrombotic settings such as those encountered during coronary revascularization procedures. Unregulated accumulation of platelets on atheromatous plaques and other thrombogenic surfaces is an absolute prerequisite for thrombus formation, which in turn represents a proximate cause of myocardial infarction, equipment malfunction, ischemic stroke and cardiovascular death(1-3). Antiplatelet drugs represent a direct outgrowth from a rapidly expanding knowledge base of fundamental platelet biology. None of the available antiplatelet therapies prevent the initiating step for thrombus formation- platelet adhesion. Accordingly, developing novel drugs that are engineered specifically to inhibit biologically relevant targets operative in platelet adhesion represents an important step toward addressing an unmet clinical need. Glycoprotein (GP) Ib-IX-V is a platelet adhesion receptor belonging to a leucine-rich repeat family of proteins. Its major function is to initiate platelet aggregation at high-shear stress blood flow, leading to thrombus formation(4). Following GP Ib-lX-V adhesion to von Willebrand Factor (VWF), platelets become activated, undergo cytoskeletal shape change and secrete platelet agonists that recruit additional platelets to the developing thrombus". Along with GP llb/llla, GP Ib-IX-V is the only platelet receptor that has a non-redundant role in hemostasis and thrombosis, with a hemostatic defect less serious than that of GP llb/llla deficiency(4-7). Thus, inhibitors of GP Ib-IX-V - VWF interaction may have the potential to become effective antithrombotic drugs with a more favorable safety proflle than GP llb/llla inhibitors. The overall effectiveness of antithrombotic therapy for managing patients with thrombotic disorders, particularly those requiring either invasive procedures or surgery has been limited in large part by the attendant bleeding risk which accompanies these therapies. While the responsible mechanisms are poorly defined, concomitant impairment of hemostasis and vascular repair have been proposed as likely contributors. We have previously shown that protein-binding oligonucleotides (aptamers), single-stranded nucleic acids that inhibit platelets, their functional ligands and/or coagulation proteins with high speciflcity can be readily reversed with complementary oligonucleotide antidotes(8-10). Of particular relevance to this proposal, we have developed aptamers against VWF that potentiy inhibit thrombosis in mice and antidote oligonucleotides that can rapidly reverse the antiplatelet activity of the aptamers(11) (Sullenger lab unpublished results). This aptamer-antidote strategy potentially offers broad applications, with the development of antithrombotic drugs which could be actively controlled, allowing full pharmacodynamic effects when clinically indicated, followed by rapid titration or complete reversal.