Large-scale functional phenotyping of ion channel arrhythmia genomic variants

描述（由申请人提供）：本提案是对RFA-HL-13-027：“筛选基因组命中的功能分析”（R21/R33）的回应。在过去的15年中，超过40个基因位点已被确定与孟德尔和多基因遗传的心律失常和/或心源性猝死风险增加有关。随着快速临床基因检测和下一代测序技术的发展，仅顶部基因座（LQT1-3）的突变/变异数量就超过了1000个。然而，这些变异中的绝大多数没有被功能性表型化。这给临床遗传学家和管理心脏病学家带来了相当大的困境-非特征变异/突变是否被视为心律失常的真正风险？这个决定是至关重要的，因为治疗包括终身药物治疗、行为矫正和经常植入心脏除颤器。此外，这些管理决策延伸到所有其他拥有变体的家庭成员。此外，随着快速、可获得和负担得起的全外显子组/基因组在临床领域占据一席之地，这些临床场景将很快变得越来越普遍。到目前为止，新变异的功能表型分型是以零零碎碎的方式进行的，每次只报告一个或几个变异。这通常是研究心脏生物学其他方面的研究人员的副业。我们建议对所有与疾病相关的遗传变异和遗传数据库中最常报道的遗传性心律失常基因座中出现的大多数罕见多态性进行功能表型分析。变体将在cdna中产生，用于在人类细胞系中表达。功能表达将通过高通量电生理系统进行评估。随后，那些表现出功能丧失表型的变异将通过生化分析进一步检测表面运输缺陷。我们将建立一个公开可用的数据库，其中包括正在进行的和已完成的结果，以便其他调查人员提供输入。这种公共资源的创建将提供

DESCRIPTION (provided by applicant): This proposal is in response to the RFA-HL-13-027: "Functional Assays to Screen Genomic Hits" (R21/R33). In the past 15 years over 40 genetic loci have been identified as associated with increased risk for cardiac arrhythmia and/or sudden cardiac death with Mendelian and multigenic inheritance. With the advance of rapid clinical genetic testing and Next-Generation sequencing, the number of mutations/variants has grown to greater than 1000 for the top loci alone (LQT1-3). A large majority of these variants have not been functionally phenotyped however. This poses a considerable dilemma for the clinical geneticist and managing cardiologist-are uncharacterized variants/mutations to be treated as true risks for arrhythmia? This decision is critical because therapy includes life-long medication, behavioral modification and often implantation of cardiac defibrillators. Moreover, these management decisions extend to all other family members harboring the variants. Furthermore, these clinical scenarios will soon become increasingly common as rapid, accessible and affordable whole exome/genome takes its place in the clinical arena. Till now, functional phenotyping of new variants has been performed in piecemeal fashion, with reports of one or a handful of variants at a time. This usually takes place as a side-project for investigators who are pursuing other aspects of cardiac biology. We propose to functionally phenotype all of the disease-associated genetic variants and most of the rare polymorphisms that have appeared in genetic databases for the loci most commonly reported for hereditary arrhythmias. Variants will be created in cDNAs for expression in human cell lines. Functional expression will be assessed with a high-throughput electrophysiology system. Subsequently, those variants exhibiting a loss-of-function phenotype will be further assayed for surface trafficking defects by biochemical analysis. We will create a publically available database with both ongoing and completed results that will allow other investigators to provide input. Creation of this public resource will provide clinical geneticist and managing physicians a valuable resource to fully analyze the results from genetic testing of their patients. Successful competition of this project will provide a framework and resource for investigators in the field and have immediate impact on clinical management of an increasing number of patients. Accordingly we believe that this proposal is in the spirit of both the RFA-HL- 13-027 and the mission of the NIH/NHLBI.