Genomics-Guided Characterization of iPS Cells from Common Mental Illnesses

描述（由申请人提供）：双相情感障碍和精神分裂症是患者、家庭和社会的重大负担。尽管一致的证据表明高遗传性（高达80%），病因仍然知之甚少。2008年，包括我们在内的几个研究小组观察到，拷贝数变异，特别是包含许多基因的大缺失，在一定程度上影响了疾病的易感性。几个研究小组还确定了精神分裂症中更普遍的罕见CNVs过量，最近，常见的单核苷酸变异已被证明有助于多基因成分，这也是疾病风险的一部分。最后，还报告了几个统计上令人信服的特定风险位点。随着遗传景观被证明是一个高度复杂的，现有的方法，模型在单个或少量的基因在整个发展的变化是不适合捕捉所有的遗传因素在一个人中相互作用。因此，该项目的总体目标是开发基于人类诱导多能干细胞（iPS）的遗传神经精神疾病模型。在R21阶段，来自双相情感障碍和精神分裂症患者沿着健康对照的预先存在的公开可用的成纤维细胞将用于试验人iPS细胞模型方法。一旦生成，将采用将这些iPS细胞分化为神经谱系的方法来表征受试者之间和受试者内的潜在差异。将使用一组神经营养因子和Wnt/GSK-3信号传导的小分子探针来表征先前涉及精神疾病的特定信号传导途径，以开发基于显微镜的自动成像和途径选择性报告基因测定。这些研究完成后，我们将建立一个系统开发精神疾病干细胞库（MISCL）的框架，该库将在第二个R33阶段扩展，以包括从我们正在进行的双相情感障碍和精神分裂症全基因组关联研究（GWAS）中获得的患者样本。R33阶段的驱动问题将是使用来自临床样本的iPS细胞，这些细胞反映了个体的整个基因组，以确定以下因素对神经发育的影响：1）染色体1q21.1，22 q11或15q13.1上的大的多基因缺失和2）多个多基因的小影响。类似地，我们将使用来自患有双相情感障碍相关的锚蛋白-G（ANK 3）单倍型的个体的iPS细胞来确定对基本离子通道生理学、轴突起始段的分子组织和神经元极性的影响。 公共卫生相关性：该项目的目的是使用新开发的方法对人体细胞进行重编程（例如，成纤维细胞）来产生诱导多能干细胞（iPS），其可用作双相情感障碍和精神分裂症的遗传准确模型。为此，我们将与马萨诸塞州总医院的哈佛干细胞研究所（HSCI）iPS核心设施密切合作，该设施在人类iPS细胞技术方面拥有丰富的经验，并已成功地从成人成纤维细胞中获得iPS系。将实施诱导iPS细胞分化为神经和神经胶质细胞的方法以及使用成像和读板仪测定法对细胞进行表型分型的新方法。

DESCRIPTION (provided by applicant): Bipolar disorder and schizophrenia are significant burdens to patients, families and society. Despite consistent evidence for high heritability (up to 80%), the etiology remains poorly understood. In 2008, several groups including ours observed that copy number variants, and in particular, large deletions each containing many genes accounted for some fraction of disease susceptibility. Several groups also identified a more general excess of rare CNVs in schizophrenia and more recently, common single nucleotide variation has been shown to contribute to a polygenic component that also accounts for a fraction of disease risk. Finally, several statistically compelling specific risk loci have also been reported. With what is proving to be a highly complex genetic landscape, existing approaches that model changes in single or small numbers of genes across development are not well suited to capture all the genetic factors interacting in an individual. Thus, the overall goal of this project is to develop human induced pluripotent stem (iPS) cell-based models of neuropsychiatric disorders that are genetically based. In the R21 Phase, pre-existing publicly available fibroblasts from bipolar disorder and schizophrenia patients along with healthy controls will be used to pilot human iPS cell model methods. Once generated, methods for the differentiation of these iPS cells into neural lineages will be employed to characterize potential between-subject and within-subject differences. Specific signaling pathways previously implicated in psychiatric disease will be characterized using a panel of neurotrophic factors and small-molecule probes of Wnt/GSK-3 signaling to develop automated microscopy- based imaging and pathway-selective reporter gene assays. Upon completion of these studies we will have established a framework for systematic development of a Mental Illness Stem Cell Library (MISCL) that will be expanded in the second R33 Phase to include samples obtained from patients from our ongoing genome-wide association studies (GWAS) of bipolar disorder and schizophrenia. The driving questions in the R33 phase will be to use iPS cells derived from clinical samples that reflect the entire genome of an individual to determine the effects on neurodevelopment of: 1) large multigenic deletions on chromosome 1q21.1, 22q11 or 15q13.1 and 2) multiple polygenes of small effect. Similarly, we will use iPS cells from individuals with bipolar disorder associated ankyrin-G (ANK3) haplotypes to determine the effect on basic ion channel physiology, molecular organization of the axon initial segment, and neuronal polarity. PUBLIC HEALTH RELEVANCE: The purpose of this Project is to use newly developed methods for reprogramming human somatic cells (e.g., fibroblasts) to create induced pluripotent stem (iPS) cells that can be used as genetically accurate models of bipolar disorder and schizophrenia. To do so we will work closely with the Harvard Stem Cell Institute (HSCI) iPS Core facility at Massachusetts General Hospital, which has extensive experience with human iPS cell technologies and have successfully derived iPS lines from adult fibroblasts. Methods for inducing the differentiation of iPS cells into neural and glial cells will be implemented as well as novel methods for phenotyping cells using imaging and plate reader assays.