Screening for enhancers of sAPPalpha

 描述（由申请人提供）：阿尔茨海默病（AD）目前在美国折磨着超过540万人，估计每年的费用超过2000亿美元。目前批准的药物只能提供短期症状缓解，但不能改变疾病进展。作为AD脑的标志的神经炎斑块是由源自淀粉样前体蛋白（APP）的淀粉样β肽（Aβ）的生成增加和/或清除减少引起的。APP可以通过两种途径加工。在一种情况下，它经历连续的β-分泌酶和γ-分泌酶裂解以产生可溶性淀粉样前体蛋白β（sAPPβ）和Aβ;在另一种情况下，它被α-分泌酶裂解以产生可溶性淀粉样前体蛋白α（sAPPα），这是一种可以预防AD的促认知肽。本项目旨在推进我们先前成功鉴定sAPPα-增强子托烷司琼（F03）的研究，目的是鉴定新的sAPPα-增强子作为新的化学实体（NCE），构成一类新型AD治疗药物。我们坚实的初步数据基础表明，该项目的具体目标很有可能实现。我们利用我们的迭代、分层化学遗传学筛选方法，从小型临床化合物库中鉴定F03（一种用于术后恶心和呕吐的药物），取得了早期成功。在AD小鼠模型中，F03在体外和体内均显示出持续但适度的sAPPα增加，并在口服治疗4周后显著改善AD模型的记忆。F03目前正在进行轻度认知障碍（MCI）的临床试验， AD作为一种重新利用的药物。在这里，我们建议利用我们的高通量筛选（HTS）试验、迭代筛选流程和大型UCLA化合物库来鉴定sAPPα增强作用大于F03的化合物。在初级HTS筛选中，将识别增加sAPPα的“命中”。在二次筛选中，使用第二个体外模型验证命中。然后通过AD模型脑的离体器官型培养和通过平行人工膜渗透性分析（PAMPA）和体内药代动力学（PK）分析确定的脑渗透率进一步验证优先命中。最后，我们将使用CEREP Bioprint图谱、药物亲和力响应靶点稳定性（DARTS）、使用相似性集合方法的计算机靶点ID以及新的凝胶增强靶点识别方法（GET）确定命中诱导sAPPα增强的机制。虽然其他新的方法来确定治疗AD和MCI正在开发中，其临床结果仍然不确定。通过鉴定sAPPα增强作用大于F03的化合物，我们可能会发现有前途的NCE用于进一步的临床开发。也许最重要的是，我们对这些增强剂的机制研究可能进一步阐明sAPPα在AD病因学中的作用，为治疗开发提供新的方向。

 DESCRIPTION (provided by applicant): Alzheimer's disease (AD) currently afflicts more than 5.4 million people in the US at an estimated cost greater than $200 billion per year. Currently approved drugs offer only short-term symptomatic relief but do not alter disease progression. The neuritic plaques that are a hallmark of AD brain result from increased generation and/or decreased clearance of the amyloid beta peptide (Aβ) which originates from amyloid precursor protein (APP). APP may be processed by two pathways. In one, it undergoes sequential β-secretase and γ-secretase cleavages to generate soluble amyloid precursor protein beta (sAPPβ) and Aβ; in the other, it is cleaved by an α-secretase to generate soluble amyloid precursor protein alpha (sAPPα), a pro-cognitive peptide that may prevent AD. This project proposes to advance our previous studies that successfully identified the sAPPα-enhancer tropisetron (F03) with the goal of identifying new sAPPα-enhancers as new chemical entities (NCEs) constituting a novel class of therapeutics for AD. Our strong foundation of preliminary data indicates a high probability that the specific aims of this project can be achieved. We have had early success utilizing our iterative, hierarchical chemical-genetics screening approach in the identification of F03 - a drug used for post- operative nausea and vomiting - from a small clinical compound library. F03 was shown to consistently but modestly increase sAPPα in vitro and in vivo in a mouse model of AD, and to significantly improve memory in the AD model after four weeks of oral treatment. F03 is now in clinical trials for mild cognitive impairment (MCI) due to AD as a repurposed drug. Here, we propose utilizing our high-throughput screening (HTS) assay, iterative screening flowscheme, and the large UCLA compound library to identify compounds with sAPPα-enhancing effects greater than F03. In the primary HTS screen, "hits" that increase sAPPα will be identified. In the secondary screen, hits are validated using a second in vitro model. Prioritized hits will then be further validated by ex vivo organotypic cultue of AD model brain and brain penetrance determined by parallel artificial membrane permeability analysis (PAMPA) and in vivo pharmacokinetic (PK) analysis. Finally, we will determine the mechanisms by which hits induce sAPPα enhancement using the CEREP Bioprint profile, drug affinity responsive target stability (DARTS), in silico target ID using the similarity ensemble approach, and by a new gel-enhanced target identification approach (GET). While other new approaches to identify therapeutics for AD and MCI are under development, their clinical outcomes remain uncertain. By identifying compounds with sAPPα enhancing effects greater than F03, we may find promising NCEs for further clinical development. Perhaps most importantly, our mechanistic studies of such enhancers may further elucidate the role of sAPPα in the etiology of AD, providing new direction for therapeutic development.