Assay Development and Discovery of LRRK2 Inhibitors for Parkinson Disease

描述（由申请人提供）：本申请的总体目标是将PD发病机制中的关键角色LRRK2的最新研究成果转化为新型PD治疗药物的药物发现管道。富亮氨酸重复激酶2 （LRRK2）突变是迄今发现的家族性和散发性PD最常见的原因。由于突变的LRRK2蛋白在体外引起LRRK2的超激活和神经元的细胞毒性，因此针对LRRK2的治疗可能对帕金森病非常有效。我们还认为LRRK2代表了治疗发展的难得机会，因为它包含两个可药物的酶模块（激酶结构域和GTPase结构域），可能提供多个药物作用位点。我们建议采用两种互补的方法来开发针对LRRK2的PD药物。具体目标1是使用重组全长LRRK2进行高通量筛选现有药物和激酶抑制剂和GTP结合阻断剂的其他化学文库，这些化学文库将通过不同的作用机制阻止LRRK2的激活。具体目的2是利用新开发的秀丽隐杆线虫PD模型进行LRRK2抑制剂的体内检测。最近的研究表明秀丽隐杆线虫可以作为早期药物筛选和验证的强大的全动物模型。我们最近建立了表达突变体LRRK2的转基因秀丽隐杆线虫动物，这些动物表现出与PD相关的易于量化的神经变性和行为障碍表型。该秀丽隐杆线虫LRRK2关联PD模型将用于从体外化学筛选中验证和表征有前途的LRRK2抑制化合物。由此产生的候选LRRK2抑制剂可能会进一步优化，以获得更好的疗效和药物样活性。一个由两名主要研究人员和几名具有LRRK2病理生物学、秀丽隐杆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，以证明我们拟议的调查是基于可靠的科学原则。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，并且在哺乳动物模型中进一步验证可能会导致治疗和预防PD的新疗法。

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.