Investigating the Functional Impact of AD Risk Genes on Neuro-Vascular Interactions

项目总结/摘要 脑血管病变存在于阿尔茨海默病的各个阶段，并与认知功能相关。 变化有强有力的证据表明，血管功能障碍是神经病理学的重要驱动因素。我们长久以来- 长期目标是了解血管细胞中阿尔茨海默病相关风险基因的功能， 对脑血管病理学发展的贡献以及将这些信息用于 治疗发展有超过27个阿尔茨海默病相关风险（AD风险）位点，包括 非编码区和编码区的许多遗传变异以及数百个相关基因。我们的总体 一种假说是AD风险基因的子集损害血管功能，导致炎症因子的释放， 血脑屏障（BBB）损伤和灌注减少，从而导致神经变性。到 为了解决这一问题，我们组建了一个具有良好合作记录的多学科团队，包括 与ADSP和ADGP成员一起，他们带来了痴呆症血管病理学方面的专业知识，内皮细胞（EC） 信号传导和EC功能测试，阿尔茨海默病基因组学，单细胞和核转录组学， 生物信息学，基于CRISPR的基因编辑用于大规模筛选和AD小鼠模型用于深入研究 体内功能评估。值得注意的是，我们将解决与重要的基因效应相关的差异， 生物变量、性别和代谢疾病。男性和女性患阿尔茨海默氏症的遗传风险不同 疾病，性别特异性多基因风险评分可更好地预测发病、进展和病理 而不是男女混合的分数超过80%的阿尔茨海默病患者患有共病代谢疾病， 从而加重血管病变。我们已经确定了前50个AD风险SNP和600个AD相关SNP。 基因，并且这些基因将被靶向用于诱导多能干细胞（iPSC）衍生的内皮细胞（EC）筛选 分别通过引物编辑和基于CRISPR的基因抑制和激活方法。基于iPSC 在2D和3D模型中的人类EC和壁细胞的生产已经被优化和缩放，以实现高效的 基因变化影响的功能测试，包括对脑类器官中神经血管相互作用的影响。 在这些人类细胞系统中的发现将通过对基因表达的深入研究来验证 在阿尔茨海默病脑样本的大量收集中， 单核测序EC翻译组也将从小鼠阿尔茨海默病模型中获得 包括性和代谢疾病。这些不同的数据集将被统一和整合， 绘制AD风险基因的血管表型，并确定作为靶向驱动因素的关键分子途径 AD脑血管病理学这些数据将增加其他人收集的知识的广度。 进一步阐明神经元、神经胶质、小胶质、内皮和壁细胞间的相互作用， 在很大程度上促成了阿尔茨海默病病理学的复杂结构。 1

PROJECT SUMMARY/ABSTRACT Cerebrovascular pathology is present throughout stages of Alzheimer’s Disease and is correlated with cognitive changes. There is strong evidence that vascular dysfunction is a significant driver of neuropathology. Our long- term objective is to understand the function of Alzheimer’s Disease-associated risk genes in vascular cells, their contribution to the development of cerebrovascular pathology and the opportunities to use this information in therapeutic development. There are over 27 Alzheimer’s Disease-associated risk (AD-risk) loci encompassing numerous genetic variants in non-coding and coding regions and hundreds of linked genes. Our overarching hypothesis is that a subset of AD-risk genes impairs vascular function, causing release of inflammatory factors, blood brain barrier (BBB) impairment, and reduced perfusion, thus contributing to neurodegeneration. To address this, we have assembled a multi-disciplinary team with a proven track record of collaboration, including with ADSP and ADGP members, who bring expertise in vascular pathology in dementia, endothelial cell (EC) signaling and EC functional testing, Alzheimer’s Disease genomics, single cell and nuclear transcriptomics, bioinformatics, CRISPR-based gene editing for large scale screening and AD mouse models for in-depth functional assessment in vivo. Notably, we will address differences in gene effects related to the important biological variables, sex and metabolic disease. Men and women differ in their genetic risk for Alzheimer’s Disease, with sex-specific polygenic risk scores providing better prediction of onset, progression, and pathology than pooled-sex scores. Over 80% of individuals with Alzheimer’s Disease have co-morbid metabolic disease, which exacerbates vascular pathology. We have identified the top 50 AD-risk SNPs and 600 AD-associated genes, and these will be targeted for induced pluripotent stem cell (iPSC)-derived endothelial cell (EC) screens by prime editing and CRISPR-based gene inhibition and activation approaches respectively. iPSC-based production of human ECs and mural cells in 2D and 3D models has been optimized and scaled to enable efficient functional testing of the impact of gene changes, including on neuro-vascular interactions in cerebral organoids. Discoveries made in these human cell systems will be validated by an in-depth investigation of gene expression changes in individual ECs and mural cells across a large collection of Alzheimer’s Disease brain samples using single nuclear sequencing. The EC translatome will also be obtained from mouse Alzheimer’s Disease models that incorporate sex and metabolic disease. These diverse datasets will be harmonized and integrated in order to map vascular phenotypes of AD-risk genes and identify critical molecular pathways that are targetable drivers of AD cerebrovascular pathology. These data will add to the breadth of knowledge being gathered by other groups to further elucidate underlying neuronal, glial, microglial, endothelial and mural cell-cell interactions that contribute in a substantial way to the complex architecture of Alzheimer Disease pathology. 1