Regulation and Dysregulation of Cardiac EC coupling by Calmodulin

项目总结/摘要 钙释放通道、兰尼碱受体（RyR 2s）的遗传和获得性缺陷是一系列 致死性心脏病，从心律失常到心力衰竭（即瑞安病）。虽然RyR 2 被认为是治疗这些疾病的逻辑靶点，基于以下的有效疗法 缺乏RyR 2功能的正常化。这部分是由于RyR 2调节的复杂性。在 这个建议，我们将测试的假设，智能工程蛋白与AAV介导的偶联 基因转移为合理设计治疗瑞安病的疗法提供了策略。基于 钙调素（CaM）在调节RyR 2中的作用，我们提出了一种新的心脏基因治疗方法， 使用RyR 2特异性和多靶点CaM（即治疗性CaM（TCaM））的ryanopathies。这一战略是 基于我们最初的研究提出的概念，即RyR 2功能的各种调节剂都集中在一个 控制RyR 2活性动力学和钙信号稳定性的共同机制 心动周期遗传和获得性RyR 2缺陷，包括RyR 2、CASQ 2和CaM的突变，改变RyR 2 不应性导致钙循环紊乱、过早的异常钙释放和随后的心律失常。 设计用于重置RyR 2不应性的治疗性CaM（TCaM），无论潜在病因如何， 为瑞安病提供了一种通用的治疗策略。为了实现这一目标，我们将使用多尺度研究 (from分子到整个动物），该联合收割机结合了蛋白质生物化学（戴维斯）实验室和细胞 生理学（Gyorke）实验室使用新的和尖端的蛋白质递送方法工程钙调素和 最先进的多室和多细胞成像工具，用于测试其功能效果。 此外，使用遗传小鼠模型和临床相关犬心脏制备物的研究 一个失败（HF）模型将为基于去硬化的新治疗策略提供“原理证明” 通过钙调素介导的钙信号传导不应性的减缓而导致的异常钙释放。我们建议：1）测试 不同遗传形式的CPVT通过共同机制损害RyR 2功能的假设： 缩短Ca信号传导不应性;和2）工程化“治疗性”（T）CaM，用于治疗 瑞安病

Project Summary/Abstract Genetic and acquired defects in Ca release channels, ryanodine receptors (RyR2s), underlie a spectrum of lethal cardiac disorders ranging from arrhythmias to heart failure (i.e. ryanopathies). Although RyR2 is considered to be a logical target for the treatment of these disorders, effective therapies based on normalization of RyR2 function are lacking. This is in part due to the complex nature of RyR2 regulation. In this proposal, we will test the hypothesis that intelligently-engineered proteins coupled with AAV-mediated gene transfer provide a strategy for the rational design of therapies to treat ryanopathies. Based on calmodulin's (CaM) role in regulating RyR2, we propose a novel cardiac gene therapy approach against ryanopathies using RyR2-specific and multi-target CaMs, i.e therapeutic CaMs (TCaMs). This strategy is based on the concept suggested by our initial studies that various regulators of RyR2 function converge on a common mechanism of refractoriness that controls RyR2 activity dynamics and Ca signaling stability during the cardiac cycle. Genetic and acquired RyR2 defects, including mutations in RyR2, CASQ2 and CaM, alter RyR2 refractoriness resulting in disturbed Ca cycling, premature aberrant Ca release and consequent arrhythmias. Therapeutic CaMs (TCaMs) designed to reset RyR2 refractoriness, regardless of the underlying etiology, will provide a general treatment strategy for ryanopathies. To accomplish this goal, we will use multi-scale studies (from molecule to whole animal) that combine the expertise of a protein biochemistry (Davis) lab and a cellular physiology (Gyorke) lab using novel and cutting edge protein delivery approaches for engineered CaMs and state-of-the art multi-compartmental and multicellular imaging tools for testing their functional effects. Additionally, studies using genetic mouse models and preparations from a clinically relevant canine heart failure (HF) model will provide a “proof-of-principle” for new therapeutic strategies based on desynchronization of aberrant Ca release by CaM-mediated slowing of Ca signaling refractoriness. We propose to: 1) Test the hypothesis that different genetic forms of CPVT impair RyR2 function through a common mechanism: shortening Ca signaling refractoriness; and 2) Engineer “therapeutic”(T)CaMs for the treatment of ryanopathies.