Evaluating Human Pluripotent Stem Cell-Derived Neural Rosette Arrays as a Neural Tube Defect Risk Screening Platform

评估人类多能干细胞衍生的神经花环阵列作为神经管缺陷风险筛查平台

• 批准号: 10218408
• 负责人: Randolph S Ashton
• 金额: $ 23.31万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-04-01 至 2023-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10218408
• 关键词: Affect; Anencephaly and spina bifida X linked; Biological Assay; Biomedical Engineering; Birth; Cell Line; Cells; Chemical Exposure; Chemicals; Clinical; Complex; Congenital Abnormality; Consensus; Cryopreserved Cell; DNA Sequence Alteration; Defect; Derivation procedure; Development; Diet; Dietary Supplementation; Drug Screening; Embryo; Environmental Risk Factor; Epidemiology; Etiology; Evaluation; FDA approved; Failure; Fibrinogen; Folic Acid; Future; Genes; Genetic; Genetic Carriers; Genetic Enhancement; Genetic Models; Genetic Predisposition to Disease; Genotype; Government; Grain; Human; In Vitro; Incontinence; Investigation; Lead; Libraries; Location; Methodology; Modeling; Mutation; Neural Tube Closure; Neural Tube Defects; Neural tube; Neuraxis; Neuroepithelial Cells; Neuroepithelial Tissue; Pathology; Patients; Perinatal mortality demographics; Phenotype; Postdoctoral Fellow; Pregnant Women; Process; Research; Risk; Rodent; Sales; Screening procedure; Sensitivity and Specificity; Signal Pathway; Spinal Dysraphism; Standardization; Structure; Testing; Tissues; To specify; Validation; Work; analog; carrier testing; cell bank; clinical translation; clinically relevant; experimental study; fortification; genome wide association study; high reward; high risk; human pluripotent stem cell; in vivo; induced pluripotent stem cell; insight; motor impairment; mutant; new technology; novel; precision medicine; prevent; relating to nervous system; screening; small molecule libraries; stem cell derived tissues; tool

Project Summary Neural tube defects (NTDs) are the second leading cause of congenital malformations affecting ~3000 births per year in the US alone and with significantly higher rates elsewhere. NTDs occur within the first month post- conception and are caused by failed closure of the neural tube, which is the developmental anlage of the entire central nervous system (CNS). The most prevalent NTDs are anencephaly and spina bifida. These defects are embryonic lethal or result in lifelong motor impairment with possible incontinence, respectively. While folic acid dietary supplementation has been clinically proven to reduce NTD risk in expecting mothers, the persistence of NTD occurrences despite proper diet suggest that multifactorial environmental and genetic factors are the primary etiological origins of remaining NTD cases. Furthermore, while rodent studies provide significant insights into NTD etiology and pathology, they do not provide the requisite throughput to efficiently screen environmental factors, i.e. chemical exposures, for associated NTD risks. Moreover, incongruence between rodent genetic models and clinical observations from human epidemiological and genome-wide association studies (GWAS) has limited development of precision medicine approaches for predicting and minimizing a patients’ NTD risk. Here, we propose to evaluate whether Neural Rosette Arrays (NRAs) can serve as a quantitative, high-throughput, human pluripotent stem cell (hPSC)-derived screening platform for assessing a chemical’s or genetic mutation’s NTD risk. While NRAs are not an exact recapitulation of in vivo primary neurulation, the bioengineered assay is the first to spatially and temporally standardize neural tube analog formation, i.e. singular neural rosette tissues, in a microarray format. Rosette tissues within NRAs possess the requisite cell phenotypes, tissue cytoarchitecture, and are derived using morphogenetic signaling pathways endogenous to the in vivo neural tube formation process. Thus, we hypothesize that the NRA platform will be an effective and efficient screening platform to detect NTD risk. Aim 1 test this hypothesis using a small chemical library screen to evaluate the NRA assay’s sensitivity and specificity. Aim 2 test this hypothesis using screens of hPSC lines genetically modified with NTD-associated genetic mutations and compared to the wild- type, isogenic control. If this high risk/high reward proposal is successful, then future research efforts would use this novel technology for systematic investigation of NTD multifactorial etiologies and clinical translation via forward screens of EPA/FDA-approved compound libraries and reverse screens using NTD patient induced pluripotent stem cell lines. In this manner, the NRA platform could have a transformative effect on NTD research, regulatory efforts to prevent commercial sale of NTD-causing agents, and future development of precision medicine approaches to minimize NTD risk in genetically predisposed patients.

项目摘要 神经管缺陷（NTD）是影响约3000名新生儿的先天性畸形的第二大原因 每年仅在美国，其他地方的比率要高得多。NTD发生在术后第一个月内- 怀孕和失败的神经管关闭造成的，这是整个发育的原基 中枢神经系统（CNS）。最常见的神经管畸形是无脑畸形和脊柱裂。这些缺陷 胚胎致死或导致终身运动障碍，可能失禁。而叶酸 膳食补充剂已被临床证明可以降低孕妇NTD的风险， 尽管饮食合理，但NTD的发生表明，多因素的环境和遗传因素是NTD发生的原因。 其余NTD病例的主要病因。此外，虽然啮齿动物研究提供了重要的 由于缺乏对NTD病因学和病理学的深入了解，它们不能提供有效筛选所需的通量 相关NTD风险的环境因素，即化学品暴露。此外， 啮齿动物遗传模型和人类流行病学和全基因组关联的临床观察 研究（GWAS）限制了精确医学方法的发展，用于预测和最小化 患者的NTD风险。在这里，我们建议评估神经玫瑰花阵列（NRA）是否可以作为一个 用于评估人多能干细胞（hPSC）的定量、高通量、人多能干细胞（hPSC）衍生筛选平台 化学物质或基因突变的NTD风险。虽然NRA不是体内原发性高血压的确切重演， 神经形成，生物工程测定是第一个空间和时间标准化神经管类似物 形成，即单个神经玫瑰花结组织，以微阵列形式。NRA内的玫瑰花结组织具有 必需的细胞表型，组织细胞结构，并使用形态发生信号通路衍生 内源性的体内神经管形成过程。因此，我们假设NRA平台将 一个有效和高效的筛查平台，以检测新台币风险。目标1使用小型测试来测试此假设 化学文库筛选以评估NRA测定的灵敏度和特异性。目标2：使用以下方法检验此假设 筛选用NTD相关的遗传突变遗传修饰的hPSC系，并与野生型进行比较。 型，同基因控制。如果这种高风险/高回报的提议成功，那么未来的研究工作将 使用这种新技术进行NTD多因素病因学和临床转化的系统研究， EPA/FDA批准的化合物文库的正向筛选和使用NTD患者诱导的反向筛选 多能干细胞系。通过这种方式，NRA平台可以对NTD产生变革性影响 研究、防止NTD致病因子商业销售的监管努力以及 精准医学方法，以最大限度地降低遗传易感患者的NTD风险。