Novel Antithrombotic Diadenosine Tetraphosphate Analogs

描述（由申请人提供）：I期结果是我们发现adp诱导的人血小板聚集的低纳摩尔水平抑制剂，具有独特的作用机制，值得进一步开发为一流的抗血栓药物。研究了一种新型、高效、高产的方法，制备了一系列碱修饰和磷酸修饰的四磷酸二腺苷（Ap4A）衍生物。所提出的修饰显示出明确的结构-活性趋势，并导致合成高效的人血小板聚集抑制剂，这些抑制剂在血浆中也很稳定。我们首次在血小板嘌呤素受体（P2X1、P2Y1和P2Y12）水平上研究了该类蛋白的作用机制，发现Ap4A及其类似物对P2Y1和P2Y12均有拮抗作用。此外，单一配体对两种血小板ADP依赖性血小板受体的前所未有的同时抑制似乎具有高度协同作用——大多数活性化合物抑制ADP诱导的血小板聚集的IC50比抑制P2Y1和P2Y12的相应IC50低一到两个数量级。因此，这些化合物代表了一类新的抗血小板药物，并最终具有新的作用机制的抗血栓药物。具体目标是：1、通过合成和研究另外6种Ap4A类似物来扩展和调整抑制人类血小板聚集的结构：活性关系；2、利用组织和重组受体模型，通过研究和量化3-6种最活跃的化合物与非血小板P2受体相互作用的能力，评估新一类Ap4A衍生物的选择性/特异性；优化和扩大该类合成和纯化的化学工艺；4、确定大鼠静脉给药后3种候选先导化合物的基本药代动力学参数并鉴定代谢物；5、研究这些候选先导化合物在留置大鼠体内静脉输注后血小板聚集特性；6、在已建立的Folts犬血栓形成模型中证实所选先导化合物的抗血栓活性。在这些研究之后，将指定一个单一的先导临床前化合物（加上备用）。血小板在止血及其病理生理中起着重要作用。不希望的血小板激活是许多常见病理的结果，例如高血压和动脉硬化，并导致血小板过度聚集和闭塞性血栓的产生。随后的缺血性事件，如心肌梗死和中风，是发达国家死亡的主要原因，抗血小板药物一直是药物开发的主要焦点。阿司匹林和氯吡罗格（Plavix， 2005年销售额59亿美元）是当今最受欢迎的药物。由于患者对氯吡格雷和正在临床开发的新的单受体靶向药物的反应存在巨大的差异，并且由于接受氯吡格雷的患者出血和发病率增加，因此需要开发快速、直接作用、快速清除血小板adp受体拮抗剂。这项研究的总体目标是确定一种新的抗血栓化合物用于人类，可以授权给一家大型制药公司进行进一步的临床前和临床开发，或者在获得适当的合作伙伴关系或资金后，我们可以将其开发成候选药物。

DESCRIPTION (provided by applicant): Phase I resulted in our discovery of low nanomolar level inhibitors of ADP-induced human platelet aggregation, with a unique mechanism of action, that warrant further development as first-in-class antithrombotic drugs. We developed a novel, efficient, high yield method to prepare a series of base and phosphate modified derivatives of diadenosine tetraphosphate (Ap4A). The proposed modifications revealed clear structure-activity trends, and led to the synthesis of highly potent inhibitors of human platelet aggregation that are also stable in plasma. We investigated for the first time the mechanism of action of this class at the level of the three platelet purinoreceptors (P2X1,P2Y1 and P2Y12), and showed that Ap4A and its analogs antagonize both P2Y1 and P2Y12. Moreover this unprecedented simultaneous inhibition of both platelet ADP dependent platelet receptors by a single ligand appears to be highly synergistic - IC50'sfor inhibition of ADP-induced platelet aggregation for the most active compounds is one to two order of magnitude lower than the corresponding IC50'sof inhibition of P2Y1 and P2Y12. Thus these compounds represent a new class of antiplatelet, and, ultimately, antithrombotic drugs with a novel mechanism of action. The specific aims are to 1, expand and tune the structure:activity relationship for inhibiting human platelet aggregation by synthesis and study of six additional Ap4A analogs, 2, evaluate the selectivity/specificity of the new class of Ap4A derivatives, by studying and quantifying the ability of 3-6 most active compounds to interact with non-platelet P2 receptors, utilizing tissue and recombinant receptor models, 3, optimize and scale up the chemical process of synthesis and purification of the class; 4, determine the basic pharmacokinetic parameters and identify metabolites of the 3 lead compounds candidates after IV administration in rats, and 5, study the aggregation properties of platelets after IV infusion of these candidates in catheterized rats, and 6, confirm the antithrombotic activity of the selected lead compound in the well-established Folts' canine thrombosis model. A single lead preclinical compound (plus backup) will be designated following these studies. Platelets play critical roles in hemostasis and its pathophysyology. Undesired platelet activation is a result of many common pathologies, e.g. hypertension and arteriosclerosis, and leads to excessive platelet aggregation and the generation of occlusive thrombi (thrombosis). The ischemic events that follow, such as myocardial infarction and stroke, are leading causes of death in the developed world, and antiplatelet drugs have been a major focus of drug development. Aspirin and clopidrogel (Plavix(R), $5.9B sales in 2005) are the most popular of the class today. Because of the vast interpatient variability in response to clopidogrel and newer single-receptor targeted drugs under clinical development, and because of the increased bleeding and morbidity in patients receiving clopidogrel, there is a need for the development of fast, direct acting, and rapidly cleared platelet ADP-receptor antagonists. The overall goal of this research is to identify a novel lead antithrombotic compound for human use, which can be licensed to a major pharmaceutical company for further pre-clinical and clinical development, or alternatively can be developed into a drug candidate by us after securing appropriate partnership or funding. PUBLIC HEALTH RELEVANCE: This project will result in an effective antithrombotic drug that will be used to treat arterial thrombosis in the acute setting. The candidate drug will directly and reversibly inhibit two important receptors involved in platelet aggregation, and will not have the drawbacks of slow and variable action of current drugs such as clopidogrel. The new drug will complement related drugs under development for arterial thrombosis.