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Small molecule mimetics of Humanin that normalize neuronal p-Akt as novel therapeutics for AD

护脑素小分子模拟物可使神经元 p-Akt 正常化，作为 AD 的新型疗法

基本信息

批准号：
10810521
负责人：
Varghese John
金额：
$ 27.48万
依托单位：
UNIVERSITY OF CALIFORNIA LOS ANGELES
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-04-01 至 2024-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10810521
关键词：
Acceleration Administrative Supplement Age Aging Alzheimer&apos s Disease Alzheimer&apos s disease model Alzheimer&apos s disease patient Alzheimer&apos s disease therapeutic Amino Acids Amyloid beta-Protein Animals Area Under Curve Autopsy Award Binding Proteins Biological Biological Assay Brain Cardiovascular Diseases Cells Characteristics Code Data Diabetes Mellitus Disease Dose Drug Kinetics Elderly Enhancers Enzyme-Linked Immunosorbent Assay Euthanasia Evaluation Future Gender Grant Half-Life Hour Human IL6ST gene Impaired cognition In Vitro Left Libraries Link Methods Microsomes Mitochondria Mus N-Methylaspartate Neuroblastoma Neurons Noise Occipital lobe Oral Peptides Permeability Pharmaceutical Preparations Pharmacodynamics Plasma Plasma Proteins Play Property Reporting Research Role Route Sampling Siblings Signal Transduction Testing Time Tissues Toxic effect Validation Work analog brain tissue candidate selection dosage evaluation/testing experimental study genome wide association study high throughput screening humanin in vivo induced pluripotent stem cell mimetics mitochondrial genome mouse model neuroblastoma cell neuroprotection novel therapeutics parent grant pharmacokinetics and pharmacodynamics scale up screening small molecule subcutaneous therapeutic candidate

项目摘要

ABSTRACT (Supplement) Humanin (HN), a 24-amino acid mitochondrial-derived peptide from the occipital lobe of postmortem sporadic AD patient brain tissue [1] is a neuroprotective peptide that could protect neurons from Aβ-related and NMDA- induced toxicity [2]. A decrease in the endogenous HN plasma level with age has been reported [3], and decreased HN levels has been linked to cognitive decline during aging. The identification of a SNP (rs2854128) using mitochondrial GWAS in the HN-coding region of the mitochondrial genome from a large sample of older adults further supports the role of HN and its circulating levels with accelerated cognitive decline [4]. The availability of high throughput screening (HTS) formattable assays, allow for screening to identify small molecules that can enhance HN expression and its levels in AD models. In the parent grant we completed screening for small molecules that act as humanin (HN) mimetics and have identified validated hits that increase both the p- gp130 and p-Akt levels in neuronal cells. Further analoging, testing and evaluation of these mimetics are ongoing following the path outlined in the grant. The availability of new assays to assess levels of HN peptide itself in vitro and in tissue has presented the opportunity to also identify small molecules that enhance HN levels. Our preliminary data using this assay shows that we can detect HN levels in SH-SY5Y cells. Hence for the Supplement research we propose is to conduct screening for HN enhancers using the human neuroblastoma SH-SY5Y cells that express and secrete HN [5]. The work we propose in the Supplement is directed toward formatting of the available HN ELISA assay for HTS and screening of a subset of the UCLA compound library to identify compounds that change HN levels in cells. Hits from the screening would be validated and then undergo secondary testing to confirm HN enhancement in SH-SY5Y and iPSC-derived human neurons, as well as in murine neuroblastoma N2a. Validated hits would also be assessed for increasing neuronal p-Akt. Thus, in the supplement we propose an Expansion of Aim1 of the parent grant to include identification of HN enhancers that along with our currently identified HN mimetics would undergo further testing and evaluation as outlined in the parent grant. Hits from the screening would then undergo secondary testing to confirm HN enhancement in SH-SY5Y and iPSC-derived human neurons, as well as in murine neuroblastoma N2a cells. Validated hits would also be assessed using our p-Akt AlphaLISA to confirm biological activity in neurons. Validated HN enhancers would undergo prioritization as part of Expansion of Aim3 of the parent grant by evaluation in in vitro ADME-T assays. Prioritized analogs would undergo pharmacokinetic (PK) and pharmacodynamics (PD) studies to identify brain permeable HN enhancers. Such an HN enhancers identified from the supplement grant would have the potential to be developed as novel therapeutic candidates for Alzheimer's disease (AD), and could in the future also be evaluated in other disorders such as diabetes and cardiovascular disease.

摘要（补充）护脑素 (HN)，一种来自死后散发性枕叶的 24 个氨基酸线粒体衍生肽 AD 患者脑组织 [1] 是一种神经保护肽，可以保护神经元免受 Aβ 相关和 NMDA- 的影响诱导毒性[2]。据报道，内源性 HN 血浆水平随着年龄的增长而下降 [3]，并且 HN 水平下降与衰老过程中认知能力下降有关。 SNP 的鉴定 (rs2854128) 在来自老年人的大量样本的线粒体基因组的 HN 编码区中使用线粒体 GWAS 成年人进一步支持 HN 及其循环水平加速认知衰退的作用 [4]。这高通量筛选 (HTS) 格式分析的可用性，允许筛选以识别小分子可以增强 AD 模型中 HN 的表达及其水平。在家长补助金中，我们完成了筛选充当护脑素 (HN) 模拟物的小分子，并已鉴定出可增加 p- 的经过验证的命中神经元细胞中的 gp130 和 p-Akt 水平。对这些模拟物的进一步模拟、测试和评估正在进行中遵循拨款中概述的路径。评估 HN 肽本身水平的新测定方法的可用性体外和组织内也提供了鉴定增强 HN 水平的小分子的机会。我们的使用该测定的初步数据表明我们可以检测 SH-SY5Y 细胞中的 HN 水平。因此对于我们建议的补充研究是利用人神经母细胞瘤筛选 HN 增强剂表达和分泌 HN 的 SH-SY5Y 细胞 [5]。我们在补编中提出的工作旨在对 HTS 可用的 HN ELISA 测定进行格式化，并筛选 UCLA 化合物库的子集，以识别改变细胞中 HN 水平的化合物。筛选中的命中将得到验证，然后进行二次测试以确认 SH-SY5Y 和 iPSC 衍生的人类神经元以及小鼠神经母细胞瘤 N2a。还将评估经验证的命中是否增加神经元 p-Akt。因此，在补充，我们建议扩大母基金的目标1，以包括 HN 增强剂的鉴定与我们目前确定的 HN 模拟物一起将接受进一步的测试和评估，如家长补助金。筛选后的命中将进行二次测试，以确认 HN 在 SH-SY5Y 和 iPSC 衍生的人类神经元，以及鼠神经母细胞瘤 N2a 细胞。经验证的点击将还可以使用我们的 p-Akt AlphaLISA 进行评估，以确认神经元的生物活性。经验证的 HN 增强剂将通过体外 ADME-T 评估作为母基金 Aim3 扩展的一部分进行优先排序化验。优先的类似物将接受药代动力学 (PK) 和药效学 (PD) 研究，以确定脑渗透性 HN 增强剂。从补充补助金中确定的此类 HN 增强剂将具有有潜力被开发为阿尔茨海默病（AD）的新型治疗候选药物，并且在未来可能还可对糖尿病和心血管疾病等其他疾病进行评估。