Human induced pluripotent stem cell modeling of genetic risk factors for Alzheimer's disease

项目摘要 阿尔茨海默病(AD)现在是一个全球性的健康危机，仅在美国就有与AD相关的费用 预计到2050年将超过1万亿美元。由于人口日益老龄化和缺乏有效的治疗方法， 迫切需要提高我们对这种疾病的病因和发病机制的了解。 有趣的是，虽然95%的AD病例是散发性的(SAD)，因此没有已知的原因，但SAD表现为高 (~60%-80%)遗传力和全基因组关联研究(GWAS)已经确定了20个基因组基因座 影响SAD风险，强烈暗示遗传风险因素在SAD发病机制中的作用。有几条证据 提示许多GWAS风险基因座通过对基因表达的特定细胞类型的影响来影响疾病风险。 然而，在1)解释非编码基因组的功能方面的困难(其中90%的疾病- 相关的风险变量)和2)产生与生理相关的人类脑细胞类型 模拟人类大脑疾病阻碍了对这些基因座的机械性询问的进展。为了 为了解决这些问题，这项提案将把最先进的功能基因组学方法与人类 诱导多能干细胞(HiPSC)技术以确定SAD遗传风险基因座在SAD中的作用 发病机制。基因匹配的脑和HiPSC来源的表观基因组和转录图谱 神经元、星形胶质细胞、少突胶质细胞和小胶质细胞将识别活性基因、顺式调节元件和 每种细胞类型中的增强子/启动子相互作用，没有个体间变异的混杂变量， 并将有助于确定悲伤风险基因座的优先顺序和功能解剖。在三个优先基因座上，表观基因组 将进行编辑和精确的基因组编辑，以确定导致悲伤的风险变体及其目标 基因。然后，将使用等基因的HiPSC系来研究这些变异对已知的SAD- 相关的细胞表型，包括淀粉样蛋白β和tau的水平和病理，淀粉样蛋白β纤维诱导的毒性， 神经元兴奋性毒性，星形胶质细胞和小胶质细胞吞噬淀粉样蛋白β纤维，以及神经胶质依赖性 对神经元活性的影响。最后，将进行基因集浓缩分析以识别细胞 被以不偏不倚的方式打乱的可悲的因果风险变量的路径，这将通知未来 实验，并可能确定潜在的治疗靶点。总之，这项提案将从功能上剖析 SAD风险基因座潜在的机制，以确定新的SAD相关基因和途径 下游治疗开发。

Project Summary Alzheimer’s disease (AD) is now a global health crisis, with AD-associated costs in the U.S. alone expected to exceed $1 trillion by 2050. Due to the growing aging population and lack of effective treatments, there is an urgent need to improve our understanding of the causes and pathogenesis of the disease. Interestingly, while >95% of AD cases are sporadic (sAD) and thus have no known cause, sAD displays high (~60-80%) heritability, and genome-wide association studies (GWAS) have identified >20 genomic loci that affect sAD risk, strongly implicating genetic risk factors in sAD pathogenesis. Several lines of evidence suggest that many GWAS risk loci affect disease risk through cell type-specific effects on gene expression. However, difficulties in 1) interpreting the functions of the non-coding genome (in which >90% of disease- associated risk variants reside) and 2) generating physiologically-relevant human brain cell types with which to model human brain disease have impeded progress in the mechanistic interrogation of these loci. In order to address these issues, this proposal will combine state-of-the-art functional genomics approaches with human induced pluripotent stem cell (hiPSC) technology in order to determine the role of sAD genetic risk loci in sAD pathogenesis. Epigenomic and transcriptomic profiling of genetically-matched brain- and hiPSC-derived neurons, astrocytes, oligodendrocytes and microglia will identify the active genes, cis-regulatory elements, and enhancer/promoter interactions in each cell type, without the confounding variable of inter-individual variation, and will facilitate the prioritization and functional dissection of sAD risk loci. At three prioritized loci, epigenome editing and precise genome editing will be performed to determine the causal sAD risk variants and their target genes. Isogenic hiPSC lines will then be used to investigate the effects of these variants on known sAD- relevant cellular phenotypes, including amyloid β and tau levels and pathology, amyloid β fibril-induced toxicity, neuronal excitotoxicity, phagocytosis of amyloid β fibrils by astrocytes and microglia, and glial-dependent effects on neuronal viability. Finally, gene set enrichment analyses will be performed to identify cellular pathways that are disrupted sAD causal risk variants in an unbiased manner, which will inform future experiments and may identify potential therapeutic targets. In summary, this proposal will functionally dissect the mechanisms underlying sAD risk loci in order to identify novel sAD-related genes and pathways for downstream therapeutic development.