Assay Development and Discovery of LRRK2 Inhibitors for Parkinson Disease

描述(申请人提供)：我们申请的总体目标是将关于LRRK2的最新研究成果转化为PD新疗法的药物发现流水线。LRRK2是PD发病机制中的关键角色。富含亮氨酸的重复蛋白激酶2(LRRK2)突变是迄今为止在家族性和散发性帕金森病中发现的最常见的原因。由于突变的LRRK2蛋白在体外导致LRRK2的过度激活和对神经元的细胞毒性，针对LRRK2的治疗药物可能对帕金森病非常有效。我们还认为，LRRK2代表着一个难得的治疗开发机会，因为它包含两个可药物的酶模块(激酶域和GTPase域)，可能为药物作用提供多个部位。我们建议采用两种互补的方法来开发针对LRRK2的帕金森病药物。具体目标1是利用重组全长LRRK2高通量筛选现有药物和额外的用于激酶抑制剂和GTP结合阻滞剂的化学库，以通过不同的作用机制阻止LRRK2激活。具体目的2是利用新开发的线虫帕金森病模型进行体内LRRK2抑制剂的检测。最近的研究表明，线虫可以作为一种强大的整体动物模型，用于早期药物筛选和验证。我们最近建立了表达突变LRRK2的转基因线虫动物，这些突变LRRK2表现出与帕金森病相关的神经退行性变和行为障碍的容易量化的表型。LRRK2连接PD的线虫模型将被用于从体外化学筛选中验证和表征有前景的LRRK2抑制化合物。由此得到的候选LRRK2抑制剂可能会进一步优化，以获得更好的疗效和类药物活性。一个由两名主要研究人员和几名具有LRRK2病理生物学、线虫生物学和药物发现专业知识的合作者组成的多学科团队已经建立。因此，已经产生了初步数据，以提供证据表明我们拟议的调查是基于健全的科学原则。我们相信，我们提出的研究可能有助于发现针对LRRK2的潜在候选药物，并在哺乳动物模型中进一步验证后，可能会导致治疗和预防帕金森病的新疗法。 公共卫生意义：帕金森病是一种毁灭性的致命脑部疾病，影响着美国的许多人。目前帕金森病的治疗选择非常有限，而且还没有治愈这种疾病的方法。由于到目前为止，LRRK2突变是家族性和散发性帕金森病最常见的原因，基于LRRK2的药物可能会对所有帕金森病患者产生非常广泛的影响。我们提出的研究的意义将是发现治疗和预防帕金森病的新药物线索的可能性。

DESCRIPTION (provided by applicant): The overall objective of our application is to translate latest research findings about LRRK2, a key player in PD pathogenesis, into the drug discovery pipeline for novel PD therapeutics. Mutations in leucine-rich repeat kinase 2 (LRRK2) are the most frequent cause thus far identified in both familial and sporadic PD. Since mutant LRRK2 proteins cause hyper-activation of LRRK2 in vitro and cytotoxicity in neurons, therapeutics targeting LRRK2 is likely to be highly effective for PD. We also reason that LRRK2 represents a rare opportunity for therapeutic development since it contains two druggable enzymatic modules (kinase domain and GTPase domain), potentially providing multiple sites for drug action. We propose to employ two complementary approaches to PD drug development targeting LRRK2. Specific Aim 1 is to use recombinant full-length LRRK2 for high throughput screening of existing drugs and additional chemical libraries for kinase inhibitors and for GTP binding blockers that will prevent LRRK2 activation through distinct mechanisms of action. Specific Aim 2 is to utilize a newly developed C. elegans model of PD for in vivo assays of LRRK2 inhibitors. Recent studies have demonstrated that C. elegans can serve as a powerful whole animal model for early-stage drug screening and validation. We have recently established transgenic C. elegans animals expressing mutant LRRK2 that manifest easily quantifiable phenotype of neurodegeneration and behavior impairment relevant to PD. This C. elegans model of LRRK2 linked PD will be used to validate and characterize the promising LRRK2 inhibitory compounds from the in vitro chemical screens. The resulting candidates of LRRK2 inhibitors may be further optimized for better efficacy and drug-like activities. A multi-disciplinary team consisting of two principal investigators and several collaborators with expertise in LRRK2 pathobiology, C. elegans biology, and drug discovery has been established. As a result, preliminary data have been generated to provide evidence that our proposed investigation is based on sound scientific principles. We believe that our proposed studies will likely facilitate the discovery of potential drug candidates targeting LRRK2, and upon further validation in mammalian models may lead to novel therapeutics for the treatment and prevention of PD. PUBLIC HEALTH RELEVANCE: Parkinson disease is a devastating and fatal brain disorder affecting many people in US. The current therapeutic options for PD are very limited and no cure exists for the disease. Because LRRK2 mutations are thus far the most frequent cause for both familial and sporadic PD, the LRRK2 based drugs will likely have a very broad impact in benefiting all PD patients. The significance of our proposed studies will be the possibility of discovering novel drug leads for the treatment and prevention of PD.