Origins of Brain Somatic Mosaicism in Developmental Brain Disease

发育性脑疾病中脑体细胞嵌合的起源

• 批准号: 10466904
• 负责人: JOSEPH G GLEESON
• 金额: $ 31.25万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-08-10 至 2026-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10466904
• 关键词: Adult; Algorithms; Alleles; Anatomy; Animal Model; Astrocytes; Autopsy; Bar Codes; Behavior; Blood; Brain; Brain Diseases; Cadaver; Candidate Disease Gene; Cell Lineage; Cell Size; Cell division; Cells; Chemicals; Childhood; Clinical; Complex; Cortical Dysplasia; DNA; DNA Methylation; DNA Repair; DNA Sequence Alteration; Data; Databases; Detection; Development; Developmental Process; Disease; Dysplasia; Electroencephalography; Electroporation; Embryo; Embryonic Development; Evaluation; Excision; Exposure to; FRAP1 gene; Freezing; Frequencies; Functional disorder; Funding; Genetic; Genetic Variation; Genomics; Genotype; Goals; Growth; Human; Individual; Intractable Epilepsy; Investments; Knowledge; Life; Maps; Metabolic stress; Methods; Microglia; Minority; Modeling; Mosaicism; Mus; Mutation; Neurons; Nuclear; Oligodendroglia; Operative Surgical Procedures; Organism; Outcome; Pathogenicity; Pathway interactions; Patients; Pattern; Pharmacology; Phenotype; Play; Predisposition; Recording of previous events; Resected; Role; Sampling; Shapes; Signal Transduction; Somatic Cell; Somatic Mutation; Sorting - Cell Movement; Synapses; Testing; Tissues; Variant; Work; brain cell; brain circuitry; cell type; clinically significant; cohort; daughter cell; deep sequencing; demethylation; disability; gene function; genome sequencing; genomic variation; human model; in utero; interest; loss of function; mTOR Inhibitor; migration; mouse model; mutant; neocortical; neurogenesis; neuron development; neuronal circuitry; neuropsychiatric disorder; neuropsychiatry; novel; recruit; relating to nervous system; resilience; response; success; synaptic function

Abstract Brain somatic mosaicism (BSM) refers to the accumulation of mutations within any of the billions of cells in the human brain, which can occur from embryogenesis through adulthood. The extent, impact and mechanisms of BSM on brain disease remain poorly understood. Prior work from the Brain Somatic Mosaicism Network (BSMN), on which the PI served, made critical breakthroughs in reliability of mosaicism detection, but also raised new questions, including the degree to which BSM exists in the healthy brain, and the mechanisms by which BSM mutations explain disease. Focal cortical dysplasia (FCD) is associated with substantial neuropsychiatric disability, and is the most common cause of intractable epilepsy in childhood. Neuropsychiatric features are seen in 15-59% of patients 5-7, and neuropathologically shows disrupted neurogenesis, migration, differentiation, and altered neural excitability. We and others previously identified mosaic mutations in the mTOR pathway in a minority of FCD cases, but most cases remain unsolved, and fundamental mechanisms are lacking. We hypothesize that: 1] FCD mutations are similar to neutral somatic mutations in their patterns and distributions, dictated by developmental processes, but differ in their functional effect. 2] BSM patterns, allelic fractions (AFs) and allele sharing between cells can reconstruct cellular lineages and migratory histories. 3] Study of FCD resected tissue can uncover novel causes of disease that would not be tolerated if present in every cell. 4] BSM modeling in mouse can unravel disrupted signaling networks of complex mosaic mutations. Our preliminary data shows: 1] From a post-mortem control cadaver, we validated 259 somatic variants using 300X genome sequencing, and started to use these variants as ‘barcodes’ to reconstruct lineage histories. 2] Deep sequencing from 314 FCD patient brain resections identified 12 new candidate genes, highlighting signaling and synaptic dysfunction, and a novel ‘two-hit’ disease mechanisms. 3] We established in utero mouse electroporation models to assess putative FCD variants as gain or loss of function, and to assess effects of ‘single-hit’ and ‘two-hit’ mutations. We propose three aims: 1] From control cadavers, we will reconstruct cell lineage across anatomical domains using BSM as barcodes. 2] With this lineage information, we will study the origins of BSM mutations in FCD, by recruiting new patients, performing both targeted and unbiased sequencing, and identifying novel causes. 3] We will functionally validate putative deleterious alleles in animal models for both ‘single-hit’ and ‘two-hit’ causes. The goal is to achieve a mechanistic understanding of the extent of BSM in control individuals, to reconstruct neural lineages and to identify novel mechanisms in developmental brain disease.

抽象的 脑体细胞嵌合体（BSM）是指大脑中数十亿个细胞中任何一个细胞内突变的积累。 人类大脑，可以从胚胎发生一直到成年。范围、影响和 BSM 对脑部疾病的机制仍知之甚少。脑体细胞之前的工作 PI服务的Mosaicism Network（BSMN）在马赛克可靠性方面取得重大突破 检测，但也提出了新的问题，包括 BSM 在健康大脑中存在的程度，以及 BSM 突变解释疾病的机制。 局灶性皮质发育不良 (FCD) 与严重的神经精神障碍有关，是 儿童顽固性癫痫的最常见原因。 15-59% 的人存在神经精神特征 患者 5-7，神经病理学显示神经发生、迁移、分化受到破坏，并且发生改变 神经兴奋性。我们和其他人之前发现了少数人 mTOR 通路中的嵌合突变 FCD 案件数量众多，但大多数案件仍未解决，且缺乏基本机制。 我们假设： 1] FCD 突变在模式上与中性体细胞突变相似，并且 分布由发育过程决定，但其功能效果不同。 2] BSM 模式，等位基因 细胞之间的分数（AF）和等位基因共享可以重建细胞谱系和迁移历史。 3] 对 FCD 切除组织的研究可以发现新的疾病原因，如果这些原因存在于 FCD 中，则无法耐受 每个细胞。 4] 小鼠 BSM 建模可以解开复杂嵌合突变中断的信号网络。 我们的初步数据显示： 1] 从尸检对照尸体中，我们验证了 259 例体细胞 使用 300X 基因组测序发现变体，并开始使用这些变体作为“条形码”来重建 血统历史。 2] 对 314 例 FCD 患者脑切除的深度测序确定了 12 个新候选者 基因，突出信号传导和突触功能障碍，以及一种新的“二次打击”疾病机制。 3]我们 在子宫小鼠电穿孔模型中建立，以评估假定的 FCD 变异作为功能的获得或丧失， 并评估“单次打击”和“两次打击”突变的影响。 我们提出三个目标：1]从对照尸体中，我们将重建跨解剖学的细胞谱系 使用 BSM 作为条形码的域。 2]有了这些谱系信息，我们将研究BSM的起源 FCD 突变，通过招募新患者、进行靶向和无偏测序，以及 找出新的原因。 3]我们将在动物模型中对假定的有害等位基因进行功能验证 “单击”和“双击”都有原因。目标是实现对 控制个体中 BSM 的程度，以重建神经谱系并识别新机制 发育性脑疾病。