Identification and Characterization of ALS/FTD Associated Variants Using Whole Genome Sequencing

项目概要/摘要 肌萎缩侧索硬化症（Amyotrophic lateral sclerosis，ALS）是一种成年发病的神经退行性疾病， 运动神经元。它最终是致命的，典型的患者生存期为2至5年。症状 额颞叶痴呆（FTD）可以在高达50%的ALS患者中检测到，并且通常认为 ALS和FTD代表疾病的连续性（ALS/FTD）。虽然绝大多数的病因 ALS/FTD病例在很大程度上是未知的，罕见的遗传因素可能在疾病易感性中起主要作用。 直到最近，遗传学研究一直利用源自外显子组测序的数据，这是由于外显子组测序的高昂成本。 全基因组测序（WGS）将分析限制在基因组的编码区。非政府组织内的变体 编码遗传元件已越来越多地显示出在细胞调节中至关重要， 人类疾病此类非编码遗传元件包括miRNA、lncRNA和转录因子的增强子/沉默子。 基因表达。然而，全基因组测序成本的大幅降低现在允许 在全基因组范围内筛选病例：对照组的罕见变异。此外，识别 结构变体，如拷贝数变体、插入、缺失和大重排， 通过使用WGS进行这些元素与ALS/FTD的相关性研究来克服。通过 通过我们自己的努力和合作者的贡献，我们已经从5000多个国家收集了原始的WGS数据。 ALS/FTD病例。此外，预计该队列的规模将翻一番，在2015年至2017年期间超过10，000例ALS/FTD病例。 未来三年。在这里，我们建议识别和表征与ALS/FTD相关的新的遗传变异。 本提案的具体目的是：（1）发现与以下疾病相关的候选遗传变异体： ALS/FTD。我们的ALS/FTD队列的原始WGS将与来自 NHLBI的TOPMed项目代表多达120，000个控件。在QC之后，将变体（单核苷酸、插入缺失和插入缺失） 其他结构变体），并进行大量的病例对照分析，以确定新的 与ALS/FTD相关的候选变体。(2)新型变体的验证/表征 与ALS/FTD相关。最佳候选遗传变异将在独立复制中复制 通过Project MinE联合体，对另外约10，000例ALS/FTD病例进行队列研究。经确认的要素将 通过几种算法/数据库进行评估，以预测其潜在的功能影响，并根据 根据其预测的功能影响，以确定将进行功能分析的功能。（三） 与ALS/FTD相关的新变体的功能分析。与ALS/FTD相关的变异体将 通过几种功能在同基因iPSC衍生的运动神经元（iMNs）和小鼠模型中进行评估 分析。这些包括通过纵向成像的存活/轴突生长、轴突运输、核质 运输、DNA损伤和整体基因表达/剪接模式。这些研究将进一步促进我们的 了解该变体对ALS/FTD发病机制的影响。

Project Summary/Abstract Amyotrophic lateral sclerosis (ALS) is an adult-onset neurodegenerative disease caused by the selective loss of motor neurons. It is ultimately lethal with a typical patient survival of two to five years. Symptoms of frontotemporal dementia (FTD) can be detected in up to 50% of ALS patients and it is generally accepted the ALS and FTD represent a continuum of disease (ALS/FTD). Although the etiology of a vast majority of ALS/FTD cases is largely unknown, rare genetic factors likely play a major role in susceptibility to the disease. Until recently, genetic studies have utilized data derived from exome sequencing due to the prohibitive cost of whole genome sequencing (WGS) limiting analysis to coding regions of the genome. Variants within non- coding genetic elements have been increasingly shown to be of vital importance in cellular regulation and human disease. Such non-coding genetic elements include miRNAs, lncRNAs, and enhancers/silencers of gene expression. However, dramatic reductions in the cost of whole genome sequencing now allow for the screening of case:control cohorts for rare variants at a genome-wide scale. Furthermore, the identification of structural variants, such as copy number variants, insertions, deletions and large rearrangements are overcome through the use of WGS allowing for association studies for these elements with ALS/FTD. Through our own efforts and the contributions of collaborators, we have amassed raw WGS data from over 5,000 ALS/FTD cases. Further, this cohort is expected to double in size to over 10,000 ALS/FTD cases within the next 3 years. Here, we propose to identify and characterize novel genetic variants associated with ALS/FTD. The Specific Aims of this proposal are: (1) Discovery of Candidate Genetic Variants Associated with ALS/FTD. Raw WGS from our ALS/FTD cohort will be combined with raw WGS data generated from the NHLBI’s TOPMed project representing up to 120,000 controls. After QC, variants (single nucleotide, indels and other structural variants) will be identified and subject to numerous case:control analyses to identify novel candidate variants associated with ALS/FTD. (2) Validation / Characterization of Novel Variants Associated with ALS/FTD. Top candidate genetic variants will be replicated in an independent replication cohort of an additional ~10,000 ALS/FTD cases through the Project MinE consortium. Validated elements will be evaluated by several algorithms/databases to predict their potential functional impact and prioritized based on their predicted functional impact to determine those that will be subject to functional analyses. (3) Functional Analysis of Novel Variants Associated with ALS/FTD. Variants associated with ALS/FTD will be evaluated in isogenic iPSC-derived motor neurons (iMNs) and murine models through several functional analyses. These include survival/axon outgrowth by longitudinal imaging, axonal transport, nuclearcytoplasmic transport, DNA damage, and global gene expression/splicing patterns. These studies will further our understanding of the impact of the variant towards ALS/FTD pathogenesis.