The role of autism susceptibility genes in the 16p11.2 locus on the development and function of human stem cell-derived neural cells

16p11.2位点自闭症易感基因对人类干细胞源性神经细胞发育和功能的作用

• 批准号: 9922991
• 负责人: Michael Frederick Wells
• 金额: $ 10.23万
• 依托单位: BROAD INSTITUTE, INC.
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-05-01 至 2021-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9922991
• 关键词: 16p11.2; Affect; Animal Model; Attention deficit hyperactivity disorder; Award; Behavior; Biochemical; Bioinformatics; Biological Models; Brain; Brain Diseases; Cell Line; Cell model; Cells; Censuses; Child; Cilia; Communication; Critical Pathways; DNA; DNA sequencing; Data; Data Set; Defect; Development; Development Plans; Diagnostic; Disease; Disease model; Drug Targeting; Experimental Designs; Feedback; Foundations; Functional disorder; Future; Gene Expression; Genes; Genetic; Goals; Grant; Growth; High Prevalence; Human; Impairment; In Vitro; Institutes; Institution; Intellectual functioning disability; Intention; International; Investigation; Laboratories; Laboratory Research; Language Disorders; Lead; Link; Maintenance; Malignant Neoplasms; Measures; Mediating; Mentors; Methods; Modeling; Molecular; Mosaicism; Mus; Neurodevelopmental Disorder; Neuroglia; Neuronal Dysfunction; Neurons; Noise; Pathway interactions; Patients; Phase; Phenotype; Population; Prevalence; Principal Investigator; Process; Protocols documentation; Reporting; Research; Role; SHH gene; Sample Size; Signal Pathway; Susceptibility Gene; Syndrome; System; Techniques; Technology; Testing; Therapeutic Intervention; Tissue-Specific Gene Expression; Training; Type I Epithelial Receptor Cell; United States; Universities; Validation; Variant; Work; Writing; Zebrafish; autism spectrum disorder; base; brain cell; career; career development; cell type; cilium biogenesis; effective therapy; experimental study; fetal; flasks; genetic risk factor; human fetal brain; human fetal brain tissue; human genome sequencing; human model; human stem cells; improved; in vitro Model; in vivo; induced pluripotent stem cell; innovation; insight; meetings; microdeletion; migration; nerve stem cell; neurodevelopment; novel; progenitor; repetitive behavior; single-cell RNA sequencing; skills; social; stem cells; symposium; targeted treatment; transcriptome; transcriptome sequencing

PROJECT SUMMARY/ABSTRACT Recent reports estimate that 1 out of every 6 children in the United States meet the diagnostic criteria for neurodevelopmental disorders such as autism spectrum disorders (ASD), attention-deficit hyperactivity disorder (ADHD), and intellectual disability (ID). The prevalence of ASDs, which are characterized by persistent social impairments, language deficits, and repetitive behaviors, has increased by 120% over the past 15 years, a problem further exacerbated by the fact that the disease mechanisms underlying ASDs are largely unknown and no targeted therapeutic interventions exist. Recent progress in human genome sequencing has begun to illuminate pathways to disease through the identification of several genetic risk factors, the most common of which is the deletion of 16p11.2 locus (16p11.2del). Initial studies have nominated specific genes in the 16p11.2 locus in neuronal dysfunction, though these findings are built on mouse and zebrafish models rather than human neural cell types. This proposal aims to elucidate the disease mechanisms underlying 16p11.2del phenotypes using in vitro induced pluripotent stem cell (iPSC)-derived human brain cells. In Aim 1 (K99), human iPSC- derived neural progenitor cells and neurons generated using novel protocols will be compared to human fetal brain tissue using single-cell RNA sequencing (scRNA-seq) techniques to validate these in vitro cellular models for future studies. Aim 2 of this proposal (K99) will employ an innovative “population-in-a-dish” strategy in which stem cell lines from many different neurotypical and 16p11.2del patients will be pooled into one culture flask to interrogate phenotypic differences. Aim 3 (R00) will leverage these scRNA-seq techniques to decipher the role of 16p11.2 genes in specific pathways important for neurodevelopment. The successful completion of these aims could lead to the identification of genetic targets for therapeutic intervention, while also dramatically changing the way the field conducts in vitro modeling of human brain disorders. These experiments will provide new training for the principal investigator (PI) of this proposal in scRNA-seq and bioinformatics methods that will serve as the foundation of an independent research laboratory that will use stem cell-derived neural cells and large transcriptome datasets, combined with animal models, to elucidate the cellular and molecular mechanisms governing neurodevelopmental disorders. This work will be completed at the Broad Institute and Harvard University, where the opportunities for technical and intellectual growth are innumerable. The PI will attend regular meetings with mentors and collaborators to receive feedback on experimental design and career decisions. The PI will attend grant writing and project management courses at Harvard, while also improving his communication skills by presenting data at international scientific conferences. As a whole, this career development plan will help the PI establish his own group at a research-oriented academic institution and become a leader in the field of neurodevelopmental disease modeling.

项目总结/摘要 最近的报告估计，美国每6个儿童中就有1个符合以下诊断标准： 神经发育障碍，例如自闭症谱系障碍（ASD）、注意力缺陷多动障碍（ADHD）、 （ADHD）和智力残疾（ID）。自闭症的患病率，其特征是持续的社会 在过去的15年里，残疾、语言缺陷和重复行为增加了120%， 由于ASD潜在的疾病机制在很大程度上是未知的， 不存在有针对性的治疗干预。人类基因组测序的最新进展已经开始 通过识别几种遗传风险因素来阐明疾病的途径， 即16p11.2位点的缺失（16p11.2del）。最初的研究已经在16p11.2中指定了特定的基因。 基因位点的神经元功能障碍，虽然这些发现是建立在小鼠和斑马鱼模型，而不是人类 神经细胞类型本研究旨在阐明16p11.2del表型的致病机制 使用体外诱导多能干细胞（iPSC）衍生的人脑细胞。在Aim 1（K99）中，人iPSC- 将使用新方案产生的衍生的神经祖细胞和神经元与人胎儿神经元进行比较。 使用单细胞RNA测序（scRNA-seq）技术对脑组织进行分析，以验证这些体外细胞模型 为将来的研究。本提案的目标2（K99）将采用一种创新的“培养皿中的人口”战略， 将来自许多不同神经典型和16p11.2del患者的干细胞系合并到一个培养瓶中， 询问表型差异。Aim 3（R 00）将利用这些scRNA-seq技术来破译 16p11.2基因在神经发育中的重要作用。这些目标的圆满实现 可能导致识别治疗干预的遗传靶点，同时也会极大地改变 该领域进行人脑疾病体外建模的方式。这些实验将提供新的 对该提案的主要研究者（PI）进行scRNA-seq和生物信息学方法的培训， 作为一个独立研究实验室的基础，该实验室将使用干细胞衍生的神经细胞和大型 转录组数据集，结合动物模型，以阐明细胞和分子机制 控制神经发育障碍这项工作将在布罗德研究所和哈佛完成 大学，在那里技术和智力发展的机会是无数的。PI将出席 与导师和合作者定期会面，以获得有关实验设计和职业生涯的反馈 决策PI将参加哈佛的拨款写作和项目管理课程，同时也提高他的 在国际科学会议上展示数据的沟通技巧。总体而言，这一职业 发展计划将帮助PI在研究型学术机构建立自己的团队， 成为神经发育疾病建模领域的领导者。