Identification and molecular characterization of somatic mutations in MCD

MCD 体细胞突变的鉴定和分子特征

• 批准号: 10125649
• 负责人: Peter B Crino
• 金额: $ 37.39万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-09-30 至 2021-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10125649
• 关键词: Identification; molecular; characterization; somatic; mutations

PROJECT SUMMARY Genetic mutations causing disease may be inherited, newly acquired in parental gametes and present in the zygote, or acquired as somatic events at some point in development after fertilization (post-zygotic). The burden and localization of any acquired mutation depends on when the mutation arises. As is the case for inherited genetic variation, there is accumulating evidence that somatic mutations can lead to severe tissue- specific disease. Malformations of cortical development (MCD) represent a group of disorders characterized by a range of morphological and structural abnormalities of the cerebral cortex reflecting errors in embryonic cortical development. MCD are associated with intellectual disability, as well as refractory epilepsy, and may require the surgical removal of the affected tissue. Inherited gene mutations involved in neuronal development explain only a fraction of MCD cases. These observations have led to the hypothesis that some MCD result from somatic mutations occurring in neuroglial progenitor cells give rise to abnormal cortical development. In fact, we and others have shown that in hemimegalencephaly, a severe hemispheric MCD subtype, ~30% of patients have somatic mutations in the PI3K-AKT-mTOR signaling pathway, detectable only by directly studying abnormal cortical tissue. Thus, in this project we will evaluate the hypothesis that somatic mutations disrupt embryonic cortical development and are responsible for a substantial fraction of MCD. We, with a network of collaborators, have access to a large number of resected epilepsy surgical tissue specimens. Using high-coverage next-generation sequencing of protein-coding regions in DNA extracted from abnormal brain tissue and unaffected tissue (leukocytes) from patients with three forms of MCD, we will identify somatic mutations within each patient with MCD (Aim 1). Somatic mutations confirmed to be selectively present in affected tissue will be assessed for their likelihood of being responsible for the cortical abnormality using comparisons of patterns of somatic mutations between cases and controls, phenotypic comparisons between individuals with somatic candidates in the same gene, patterns of mutations in pathologically abnormal and normal cell populations, and evaluations of the effects of the candidate mutation on cerebral cortical development in vivo (Aim 2). Finally, initial characterization of the effects and presentation of the disease- causing mutation in the cortex will be studied using in vitro assays to evaluate the effects of the mutation on key cortical developmental processes, and mutation lineage tracing to identify cell populations carrying the mutation (Aim 3). Our project brings together clinical, genetic, and neurobiological expertise and builds on the activities of the Epilepsy Phenome/Genome Project (EPGP), Epi4K, and multiple NINDS-funded initiatives in somatic genetics. These studies will: (i) provide the first detailed assessment of the role of somatic mutations across MCD subtypes, (ii) identify novel genes/pathways underlying cortical development, and (iii) establish and test in vitro and in vivo models to understand the role of genes in cortical development.

项目总结 致病的基因突变可能是遗传的，新获得的亲本配子，并存在于 受精卵，或在受精后(受精卵后)发育的某个时间点上获得的体细胞事件。这个 任何获得性突变的负担和定位取决于突变发生的时间。就像情况一样 遗传遗传变异，越来越多的证据表明，体细胞突变可以导致严重的组织- 特定的疾病。皮质发育畸形(MCD)代表一组疾病，其特征是 反映胚胎发育错误的一系列大脑皮层形态和结构异常 皮质发育。MCD与智力残疾以及难治性癫痫有关，并可能 需要手术切除受影响的组织。与神经元发育有关的遗传性基因突变 只解释一小部分MCD病例。这些观察结果导致了一种假设，即某些MCD结果 发生在神经胶质前体细胞中的体细胞突变会导致皮质发育异常。在……里面 事实上，我们和其他人已经表明，在半巨脑，一种严重的半球MCD亚型，~30% 患者在PI3K-AKT-mTOR信号通路中存在体细胞突变，只有通过直接 研究异常的皮质组织。因此，在这个项目中，我们将评估体细胞突变的假设 干扰胚胎皮质发育，并对相当大一部分MCD负责。我们，带着一个 网络合作者，有机会接触到大量癫痫切除手术组织标本。vbl.使用 异常脑DNA蛋白质编码区的高覆盖率下一代测序 组织和未受影响的组织(白细胞)来自三种形式的MCD患者，我们将鉴定体细胞 每名MCD患者体内的突变(目标1)。体细胞突变被证实选择性地存在于 受影响的组织将被评估其对皮质异常负有责任的可能性 病例和对照的体细胞突变模式比较，表型比较 具有相同基因的体细胞候选的个体，病理异常和 正常细胞群体，以及候选突变对大脑皮层影响的评估 体内发育(目标2)。最后，疾病的影响和表现的初步特征-- 将使用体外试验来研究在大脑皮层引起突变，以评估突变对大脑的影响 关键的皮质发育过程和突变谱系追踪以确定携带 突变(目标3)。我们的项目汇集了临床、遗传学和神经生物学的专业知识，并建立在 癫痫表型/基因组计划(EPGP)的活动，Epi4K，以及NINDS资助的多项倡议 体细胞遗传学。这些研究将：(I)首次详细评估体细胞突变的作用 在MCD亚型中，(Ii)识别支持皮质发育的新基因/途径，以及(Iii)建立 并测试体外和体内模型，以了解基因在皮质发育中的作用。