ApoE4-targeted therapeutics that normalize SirT1

项目/摘要摘要 我们的研究首次将阿尔茨海默病的主要风险因素--载脂蛋白E4--与主要 长寿决定因素，sirtuins；并确定针对这一新联系的第一种候选疗法。 阿尔茨海默病(AD)目前在美国困扰着540多万人，据估计 每年超过2000亿美元的社会。目前批准的治疗AD的药物仅提供短期治疗 症状缓解，但不改变病情进展。阿尔茨海默病(AD)的主要危险因素 是载脂蛋白E(ApoE4)的epsilon-4(ε4)等位基因，它存在于大约三分之二的AD患者中。这个 ApoE4等位基因(染色体19q13)增加了散发性和晚发性AD(LOAD)的风险。 尽管十多年来人们知道载脂蛋白E4等位基因在某种程度上对疾病过程有贡献，但 载脂蛋白E4相关AD风险的确切分子机制仍不清楚。我们的研究揭示了 载脂蛋白E4介导的毒性的新机制和揭示了一个关键的介体−SirT1− 受ApoE4与ApoE3的影响。有趣的是，虽然ApoE3和ApoE4都绑定到APP，但只有ApoE4关联 具有纳米分子亲和力(Kd~80 nm)，且只有载脂蛋白E4显著：(A)降低sAPPα与Aβ的比率；(B) 降低SirT1的表达，导致SirT1水平和神经保护率的显著降低 SIRT1与神经毒性SIRT2结合；(C)触发tau和APP的磷酸化；(D)诱导细胞程序性死亡。 在我们对临床文库的初始筛选中，我们已经确定了一个有希望的热门(A03)，它是一个重新定位的候选者， 具有高度的大脑渗透性，并逆转了SirT1水平的下降。作为这项提议的一部分，我们计划 完成A03的临床前测试，并开发A03的新化学实体(NCE)类似物，以便进一步 发展。此外，通过筛选和“点击到领先”的优化，我们计划发现新的领先优势 参加进一步测试的候选人。该提案的最终目标是为非普洛斯提供1-2名候选人 毒性测试。我们的数据支持神经元连通性--受临界值比率影响的假说 包括SAPα：Aβ、SIRT2：SIRT1、APP：P-APP和tau：P-tau-在内的介体被ApoE4以编程方式改变。 与临床核心和Gylys实验室的合作在这个项目中很重要，因为它将提供 ApoE基因分型患者血浆和脑脊液标本中SirT1水平的初步分析这样的数据 将对未来ApoE4靶向候选药物的临床测试和开发非常有用 SIRT1可作为MCI/AD的潜在血浆标志物。此外，将SirT1与其他生物标志物进行比较 受sirtuin/NFkB信号影响的血浆/CSF是计划的，这将有助于进一步阐明其作用 ApoE4在AD中的表达。这个项目的总体主要目标是确定有效的、口服有效的、脑渗透的 SIRT1-增强适合进一步临床前IND开发的候选药物作为第一个ApoE4- 针对AD的靶向SirT1疗法，并开发必要的工具以确定目标参与度 和药效。

PROJECT / SUMMARY ABSTRACT Our studies link for the first time the major risk factor for Alzheimer's disease - ApoE4 - with major longevity determinants, the Sirtuins; and identify the first candidate therapeutics that target this new link. Alzheimer's disease (AD) currently afflicts more than 5.4 million people in the US at an estimated cost to society of greater than $200 billion per year. The currently approved drugs for AD provide only short-term symptomatic relief but do not alter disease progression. The dominant risk factor for Alzheimer's disease (AD) is the epsilon-4 (ε4) allele of apolipoprotein E (ApoE4), which is present in about two-thirds of AD patients. The ApoE4 allele (chromosomal locus 19q13) confers increased risk for sporadic and late-onset AD (LOAD). Despite over a decade of knowledge that the ApoE4 allele is somehow contributory to the disease process, the precise molecular mechanisms underlying ApoE4-associated AD risk remain unclear. Our studies shed light on a novel mechanism for ApoE4-mediated toxicity and revealed a key mediator − SirT1 − that is differentially affected by ApoE4 vs. ApoE3. Interestingly, while both ApoE3 and ApoE4 bind to APP, only ApoE4 associates with nanomolar affinity (Kd ~80nM), and only ApoE4 significantly: (a) reduces the ratio of sAPPα to Aβ; (b) reduces SirT1 expression, resulting in a marked reduction of SirT1 levels and in the ratio of neuroprotective SirT1 to neurotoxic SirT2; (c) triggers tau and APP phosphorylation; and (d) induces programmed cell death. In our initial screen of a clinical library we have identified a promising hit (A03) that is a repurposing candidate, is highly brain permeable, and reverses the reduction of SirT1 levels. As part of this proposal we plan to complete the preclinical testing of A03 and develop new chemical entity (NCE) analogs of A03 for further development. In addition, through screening and “hit-to-lead” optimization we plan to discover new lead candidates for further testing. The eventual goal of the proposal is to provide 1-2 candidates for non-GLP toxicity testing. Our data support the hypothesis that neuronal connectivity - influenced by the ratios of critical mediators including sAPPα:Aβ, SirT2:SirT1, APP:p-APP, and tau:p-tau - is programmatically altered by ApoE4. The collaboration with the Clinical Core and the Gylys lab is important in this project, as it would provide preliminary analysis of plasma and CSF samples from ApoE genotyped patients for levels of SirT1. Such data would be extremely useful for future development of ApoE4-targeted drug candidates to clinical testing and SirT1 as a potential plasma biomarker in MCI/AD. In addition, comparing SirT1 with other biomarkers in plasma/CSF that are affected by the sirtuin/NFkB signaling is planned and would help further elucidate the role of ApoE4 in AD. The overall primary objective of this project is to identify potent, orally active, brain permeable SirT1-enhancing lead candidates that are suitable for further preclinical IND development as the first ApoE4- targeted SirT1 therapeutics for AD and to development the tools necessary to ascertain target engagement and efficacy.