Mechanism of gamma-secretase modulators

摘要 阿尔茨海默病(AD)是一种毁灭性的神经退行性疾病，目前影响5.8 百万美国人。然而，目前还没有FDA批准的能够延迟或停止的治疗方法 疾病的发展。淀粉样β蛋白(Aβ)斑块被认为是阿尔茨海默病发病过程中不可或缺的部分。 在“淀粉样级联假说”中的作用，在该假说中，β多肽的积累启动了一系列 导致神经变性并最终导致阿尔茨海默病的病理事件。γ-分泌酶是一种天冬氨酸蛋白酶 负责各种基材的加工。它被认为是有吸引力的药物靶点，因为它 在蛋白降解的最后一步裂解淀粉样前体蛋白(APP)产生不同种类的β多肽 此外，γ分泌酶的催化亚单位APP和早老素的突变与 家族性阿尔茨海默病并显示可改变Aβ的产生。不幸的是，所有γ分泌酶抑制剂的临床试验都证明 由于对其他γ-分泌酶底物的脱靶效应，如缺口，不成功。γ的失败- 分泌酶抑制剂刺激了一系列新化合物的发现和开发，这些化合物被称为γ- 分泌酶调节剂(GSM)。最初发现的NSAID的一个子集可以选择性地降低 致病的Aβ42有利于致病性较低的Aβ38而不抑制Notch。对这些方面的改进 化合物导致了第二代GSM。这个项目将集中在两个第二代班级 GSM：源自非类固醇抗炎药和咪唑GSM的酸性GSM。酸和咪唑GSM已经被 以多个群体为特征，在增加致病Aβ物种的同时显示出减少致病A DNA物种的前景 致病性越低的物种。然而，他们在有关β的临床试验中仍显示出局限性 选择性、效力和毒性。我们的目标是明确GSM的分子机制，以便 改进它们以用于AD药物开发。本实验室用基于GSM的光探针标记γ-分泌酶 已发现酸和咪唑GSM结合在γ分泌酶的早老素亚基上的不同位置。基座 根据这一信息，我们假设酸和咪唑GSM结合到γ-的不同的变构位置- 分泌和协同作用，有效地调节Aβ的产生。在界面上使用方法 生物学和化学，本课题的目标是：1)确定GSM与γ分泌酶的结合机制 2)确定GSM联合对A-β产生、毒性和认知功能的影响。为了调查 GSM的分子机制，我们将使用化学探针结合质谱学和结构学方法 建模以确定准确的结合位点。我们还将评估Aβ中的酸和咪唑GSM对。 并使用最有效的浓度来分析毒性和认知能力 AD小鼠模型。总体而言，拟议的研究将使我们能够破译GSM的机制， 最终，开发具有最高选择性和效力的GSM。这项提案的发现将揭开新的 调节γ-分泌酶以更有效地治疗AD的可能性。

Abstract Alzheimer’s disease (AD) is a devastating neurodegenerative disorder which currently affects 5.8 million Americans. However, there are currently no FDA-approved treatments capable of delaying or stopping disease progression. Amyloid-beta (Aβ) plaques are believed to be integral to AD pathogenesis through their role in the “amyloid cascade hypothesis,” in which the accumulation of Aβ peptides initiates a chain of pathological events leading to neurodegeneration and ultimately AD. γ-secretase is an aspartyl protease responsible for processing a wide range of substrates. It is considered an attractive drug target because it cleaves amyloid precursor protein (APP) in the final step of proteolysis to generate Aβ peptides of varying lengths; moreover, mutations in APP and presenilin, the catalytic subunit of γ-secretase, have been linked to familial AD and shown to alter Aβ production. Unfortunately, all clinical trials of γ-secretase inhibitors proved unsuccessful due to off target effects on other γ-secretase substrates, such as Notch. The failures of γ- secretase inhibitors spurred the discovery and development of a new series of compounds known as γ- secretase modulators (GSMs). A subset of NSAIDs were initially discovered to selectively reduce levels of the pathogenic Aβ42 in favor of the less pathogenic Aβ38 without inhibition of Notch. Improvements on these compounds led to second generation GSMs. This project will focus on two classes of second generation GSMs: acid GSMs derived from NSAIDs and imidazole GSMs. Acid and imidazole GSMs have been characterized by multiple groups and showed promise in decreasing pathogenic Aβ species while increasing the less pathogenic species. However, they still demonstrated limitations in clinical trials regarding Aβ selectivity, potency, and toxicity. Our objective is to define the molecular mechanism of GSMs in order to improve them for AD drug development. Using GSM-based photoprobes to label γ-secretase, our laboratory has found that acid and imidazole GSMs bind to distinct sites on the presenilin subunit of γ-secretase. Based on this information, we hypothesize that acid and imidazole GSMs bind to distinct, allosteric sites of γ- secretase and work in synergy to effectively modulate Aβ production. Using approaches at the interface of biology and chemistry, this project aims to: 1) Determine the mechanism of binding of GSMs on γ-secretase and 2) Determine the effects of GSM combination on Aβ production, toxicity, and cognition. To investigate the molecular mechanisms of GSMs, we will use chemical probes combined with mass spectrometry and structural modeling to identify the precise binding sites. We will also evaluate pairs of acid and imidazole GSMs in Aβ- secreting cell lines and use the most effective concentrations to analyze toxicity and cognitive performance in an AD mouse model. Overall, the proposed studies will enable us to decode the mechanism of GSMs and, ultimately, develop GSMs with the highest selectivity and potency. Findings from this proposal will unlock new possibilities for modulation of γ-secretase to more effectively treat AD.