Discovery of Novel Genetic Variants Underlying Unexplained Hypertrophic Cardiomyopathy by Whole Genome Sequencing, RNA-seq and iPSC modeling

项目总结 该提案描述了Alireza HagHighi博士的职业发展五年计划，目标是成为 独立的心血管调查员。目前是医学讲师和临床分子遗传学家 布里格姆妇女医院(BWH)和哈佛医学院(HMS)， HagHighi博士的长期职业目标 是将基因组研究转化为心血管患者的临床进展。他的职业发展和 研究计划利用他在人类遗传学方面的培训以及BWH、HMS和 博德研究所将为他提供统计学、计算生物学和诱导多能性干细胞方面的新技能 细胞(IPSC)疾病模型，以便对肥大的机制提供新的见解 心肌病(HCM)。他的职业发展目标将通过科学调查和 合作，并得到导师、授课课程和会议的支持。HagHighi博士已经组装了一个 指导和科学咨询委员会，成员包括世界心脏病学和人类遗传学专家。 克里斯汀·塞德曼(主要导师)，乔恩·塞德曼(联合导师)，沙米尔·桑亚耶夫(顾问)和马克·戴利(顾问)， 他们将无数年轻的调查员培养成了成功的独立调查员。 肥厚性心肌炎是最常见的遗传性心血管疾病，也是#年猝死的主要原因。 年轻人。肌节蛋白的显性突变导致30%-60%的患者发生肥厚性心肌梗死--数据显示 能够进行机制研究，推动治疗肥厚型心肌炎的新治疗策略。这些机会是 严重局限于肌节阴性(原因不明)的肥厚型心肌梗死患者，他们对这些患者的发病机制尚不清楚 也不知道为什么会出现疾病。HagHighi博士在遗传未解释中的整个外显子组测序(WES)分析 HCM患者认为非编码变异可能在本病的发病机制中起一定作用。这项建议 寻求确定尚未解决的患者的肥厚型心肌病的遗传病因(重点是非编码变异) 通过全基因组测序(WGS)对WES进行测序。为了确定新的功能影响 已识别的非编码变体，HagHighi博士将对人类的优先候选变体进行建模和表征 诱导多能干细胞来源的同基因心肌细胞(IPSC-CMS)。使用这些方法，他 将探索三个具体目标：(1)识别已知HCM基因非编码区的罕见变异，并 WGS对不明原因肥厚性心肌病患者血液和心脏组织样本中的优先基因进行分析，(2)确定 通过RNAseq对已识别的变异体的基因表达异常和剪接效应进行研究，以及(3)评估其影响 人类ipscs中的优先非编码变体。通过提供对功能角色的基本新见解 对于可能发现HCM新原因(从而改善患者护理)的非编码变体，这些研究将 支持NHLBI目标，以提高我们对健康和疾病的分子基础的理解。此外, 成功完成拟议研究提前完成 HagHighi博士向独立诊所的过渡- 并为他申请校外资金提供了坚实的基础。

PROJECT SUMMARY This proposal describes a five-year plan for Dr. Alireza Haghighi's career development towards becoming an independent cardiovascular investigator. Currently an Instructor in Medicine and Clinical Molecular Geneticist in Brigham and Women's Hospital (BWH), and Harvard Medical School (HMS), Dr. Haghighi's long-term career goals are to translate genomic research into clinical advances for cardiovascular patients. His career development and research plan leverages his training in human genetics and the unique training environment of BWH, HMS, and the Broad Institute that will provide him with new skills in statistics, computational biology, and induced pluripotent stem cell (iPSC) disease modeling, in order to shed novel insight into the mechanisms underlying hypertrophic cardiomyopathy (HCM). His career development goals will be achieved through scientific investigation and collaborations, and supported by mentorship, didactic coursework and conferences. Dr. Haghighi has assembled a Mentoring and Scientific Advisory Committee that includes world experts in cardiology, and human genetics: Drs. Christine Seidman (primary mentor), Jon Seidman (co-mentor), Shamil Sunyaev (advisor) and Mark Daly (advisor), who have mentored countless young investigators to successful independent investigators. HCM is the most common inherited cardiovascular diseases and the leading cause of sudden death in young people. Dominant mutations in sarcomere proteins cause HCM in 30-60% of patients- data that has enabled mechanistic studies that propel new therapeutic strategies to treat HCM. These opportunities are profoundly limited in sarcomere-negative (unexplained) HCM patients in whom there is no understanding of how or why disease emerges. Dr. Haghighi's whole exome sequencing (WES) analyses in genetically unexplained HCM patients concluded that non-coding variants may play a role in pathogenesis of this disease. This proposal seeks to define genetic etiologies (with a focus on non-coding variants) for HCM in patients that remain unsolved after WES sequencing by whole genome sequencing (WGS). To determine the functional impact of newly identified non-coding variants, Dr. Haghighi will model and characterize prioritized candidate variants in human isogenic cardiomyocytes derived from induced pluripotent stem cells (iPSC-CMs). Using these approaches he will explore three specific aims that (1) Identify rare variants in non-coding regions of known HCM genes and prioritized genes in blood and heart tissue samples of unexplained HCM patients by WGS, (2) Define dysregulated gene expressions and splicing effects of identified variants by RNAseq, and (3) Assess the effects of prioritized non-coding variants in human iPSCs. By providing fundamental new insights into the functional role of non-coding variants that may uncover new causes of HCM (thereby improving patient care), these studies will support NHLBI goals to improve our understanding of the molecular basis of health and disease. In addition, successful completion of the proposed studies advance Dr. Haghighi's transition to an independent clinical- scientist and provide him with a solid foundation from which he will apply for extramural funding.