Genes and pathways underlying brain overgrowth and focal cortical malformations

 描述（由申请人提供）：巨头症（或MEG）是一种以全身性脑过度生长为特征的发育性脑疾病。它发生在越来越多的发育和代谢障碍中，并与严重的儿童神经系统并发症有关，包括癫痫、智力残疾、自闭症、脑积水和基亚里畸形。MEG显示与更严重但节段性脑部疾病（包括半侧巨脑畸形（HMEG）和局灶性皮质发育不良（FCD））有显著重叠，这些疾病共同构成儿童难治性癫痫的最常见原因。最近已经在MEG、HMEG和FCD中鉴定了关键PI 3 K-AKT-mTOR信号传导网络（包括PIK 3CA、PIK 3R 2、AKT 3和CCND 2）内的关键基因中的激活突变，表明这些表型构成了具有共享的分子病因学和神经病理学特征的单一广谱发育性脑障碍。 该提案的目标是询问受影响的人源性细胞和组织中这些疾病的分子基础，以确定这些疾病中的突变谱、嵌合体水平、组织分布和途径失调。首先，我建议对从癫痫手术中获得的HMEG和FCD人脑样本中受影响的神经元进行单细胞测序，以测试已知和候选基因。其次，我将使用高通量蛋白质组学在受影响的人脑组织中检测PI 3 K-AKT通路蛋白，这些脑组织中的关键PI 3 K-AKT通路基因发生突变，以识别通路失调并定位特定的子通路和下游靶点。第三，迄今为止发现的PI 3 K AKT基因解释了约75%的患有两种最常见的MEG综合征的儿童，约25%的患有HMEG-FCD的儿童，没有罕见的MEG综合征。我建议进行研究，以确定其他MEG-HMEG-FCD致病基因使用全外显子组和全基因组测序的多个组织从受影响的个人。 我的职业目标是成为一名医生科学家，致力于将尖端的基因组学和蛋白质组学技术整合到人类脑组织中，包括单细胞测序，以及准确和定量的表型分析，以进一步了解人类发育性脑疾病的分子和生物学基础。我提出了一个五年的研究计划，该计划将在威廉B博士的指导下进行教学和研究培训。综合脑研究中心（CIBR）的多宾斯。我的咨询委员会和合作者网络，加上西雅图儿童研究所和华盛顿大学的资源，将为我成功完成这项提案并过渡到独立资助的医生科学家提供必要的环境。

 DESCRIPTION (provided by applicant): Megalocephaly (or MEG) is a developmental brain disorder characterized by generalized brain overgrowth. It occurs in a growing number of developmental and metabolic disorders and is associated with severe childhood neurological complications including epilepsy, intellectual disability, autism, and hydrocephalus and Chiari malformations. MEG shows significant overlap with more severe but segmental brain disorders including hemimegalencephaly (HMEG) and focal cortical dysplasia (FCD) that collectively constitute the most common cause of intractable epilepsy in children. Activating mutations in key genes within the critical PI3K- AKT-mTOR signaling network (including PIK3CA, PIK3R2, AKT3 and CCND2) have been recently identified in MEG, HMEG and FCD, suggesting that these phenotypes constitute a single broad spectrum of developmental brain disorders with shared molecular etiologies and neuropathological features. The goal of this proposal is to interrogate the molecular basis of these disorders in affected human-derived cells and tissues to define the mutational spectrum, levels of mosaicism, tissue distribution and pathway dysregulation in these disorders. First, I propose to perform single cell sequencing of affected neurons from HMEG and FCD human brain samples obtained from epilepsy surgery to test for known and candidate genes. Second, I will globally assay PI3K-AKT pathway proteins in affected human brain tissues with mutations in key PI3K-AKT pathway genes using high throughput proteomics to identify pathway dysregulation and localize specific sub-pathways and downstream targets. Third, the PI3K AKT genes discovered so far explain ~75% of children with the two most common MEG syndromes, ~25% with HMEG-FCD and none with rare MEG syndromes. I propose studies to identify additional MEG-HMEG-FCD causative genes using whole exome and whole genome sequencing on multiple tissues from affected individuals. My career goal is to become a physician-scientist devoted to integrating cutting-edge genomic and proteomic techniques on human-derived brain tissues, including single cell sequencing, with accurate and quantitative phenotyping to further our understanding of the molecular and biologic basis of human developmental brain disorders. I propose a five-year research program that will incorporate didactic and research training under the mentorship of Dr. William B. Dobyns at the Center for Integrative Brain Research (CIBR). My advisory committee and network of collaborators, combined with the resources at the Seattle Children's Research Institute and University of Washington, will provide the environment necessary for my successful completion of this proposal and transition to an independently funded physician-scientist.