Targeting the genotype to phenotype link in HCM as a therapeutic strategy

室性心律失常和心源性猝死（SCD）是肥厚性心肌病的常见并发症， 心肌病（HCM），尤其是年轻人。肌节蛋白基因的致病性变体 引起大约一半的遗传性肥厚型心肌病和大约三分之一的散发性肥厚型心肌病。然而，人们对 肌节蛋白变异如何导致心律失常和心脏性猝死。有一个主要的未满足的 需要更好地了解疾病机制，以预测有SCD风险的患者， 设计基于机制的疗法。 为了解决这一空白，我们将应用高通量功能基因组学来鉴定单个蛋白质 成分的促凋亡信号传导，并建立其功能，在体外研究iPSC- 衍生的心肌细胞（iPSC-CM）和HCM突变小鼠中的体内研究。我们将开始 在iPSC衍生的细胞中产生细胞内促凋亡信号传导的功能基因组学探针， 在MYBPC 3、MYH 7和TNNT 2中携带HCM致病变体的心肌细胞。探测器将在 基于筛选合成的miRNAs（syn-miRs）， 蛋白质，因此成为复杂生物学的理想探针。探针选择性分析 对于基因变异体，将表明存在共同和/或不同的信号传导机制。同时， 我们将从肌切除术样本的分析中识别和描述候选通路， MYBPC 3突变型HCM患者。一旦我们获得了途径信息和探针， 这两种方法，我们将全面确定蛋白质介导的高通量 在MYBPC 3突变体iPSC-CM中的功能评估。基于iPSC-CM中的效果， 对于MYBPC 3的选择性和作为药物靶点的潜力，我们将优先考虑最有希望的候选物 在携带Mybpc 3突变的HCM转基因小鼠中通过AAV 9敲低进行体内评价的靶点 与iPSC-CM中的同源。我们希望调节候选人的功能 介质将抑制Mybpc 3突变小鼠中的心律失常和心动过速。 总而言之，这些研究将增加我们对以下因素引起的致瘤信号的理解： HCM突变和促进改善预后和机制为基础的发展 治疗家族性HCM患者。它也将增加我们对基本的理解， 心肌细胞生物学可能是其他心脏疾病的基础。具体目标是：1）确定 如果离散的信号传导机制导致“高”倾向性HCM基因的遗传易感性， 变体，和2）全面定义通过功能决定电重构的蛋白质 筛选MYBPC 3突变体iPSC-CM，并测试它们作为Mybpc 3治疗靶点的功效。 HCM的突变小鼠模型。

Ventricular arrhythmia and sudden cardiac death (SCD) is a prevalent complication of hypertrophic cardiomyopathy (HCM) especially in young adults. Pathogenic variants of sarcomeric protein genes cause about half of inherited HCM and about a third of sporadic HCM. Little is known, however, about how sarcomeric protein variants lead to arrhythmia and sudden cardiac death. There is a major unmet need for a better understanding of disease mechanisms in order to predict patients at risk for SCD and design mechanism-based therapeutics. To address this gap, we will apply high throughput functional genomics to identify individual protein components of arrhythmogenic signaling, and establish their function using in vitro studies in iPSC- derived cardiomyocytes (iPSC-CMs) and in vivo studies in HCM mutant mice. We will begin by generating functional genomics probes of the intracellular arrhythmogenic signaling in iPSC-derived cardiomyocytes carrying HCM causative variants in MYBPC3, MYH7 and TNNT2. The probes will be identified based on screening synthetic miRNAs (syn-miRs) which collectively suppress nearly all proteins in the cell and therefore make ideal probes of complex biology. Analysis of probe selectivity for gene variants will indicate the existence of common and/or distinct signaling mechanisms. In parallel, we will identify and characterize candidate pathways indicated from analysis of myectomy samples from MYBPC3 mutant HCM patients. Once we have obtained pathway information and probes from these two approaches, we will comprehensively determine the protein mediators by high throughput functional evaluation in the MYBPC3 mutant iPSC-CMs. Based on the effect in the iPSC-CMs, selectivity for MYBPC3 and potential as a drug target, we will prioritize the most promising candidate targets for in vivo evaluation by AAV9 knockdown in HCM transgenic mice carrying a Mybpc3 mutation homologous to that in the iPSC-CMs. We expect that modulating the function of the candidate mediators will suppress arrhythmia and tachycardia in the Mybpc3 mutant mice. In summary, these studies will increase our understanding of the arrhythmogenic signaling caused by HCM mutations and promote the development of improved prognostic and mechanism-based therapeutics for familial HCM patients. It will also increase our understanding of fundamental cardiomyocyte biology that might underlie other cardiac diseases. The Specific Aims are: 1) Determine if discrete signaling mechanisms cause arrhythmic susceptibility across “high” propensity HCM gene variants, and 2) Comprehensively define the proteins that dictate electrical remodeling by functional screening in MYBPC3 mutant iPSC-CMs and test their efficacy as therapeutic targets in an Mybpc3 mutant mouse model of HCM.