Molecular Pharmacology of An Inherited Heart Disease

本申请中提出的研究的总体目标是了解由SCN5A基因遗传突变引起的心律失常的分子基础，并确定新的基因靶向治疗策略来治疗心律失常。中心假设是，这些心律失常发生的一个步骤是由SCN5A基因产物（主要的心脏Na+通道α亚基）的生物物理特性改变引起的膜电活动扰动，这是由疾病相关突变引起的，但类似的功能扰动可能与不同的临床疾病有关。离子通道特性的改变也可能赋予编码离子通道独特的药理学特性，使其成为治疗干预的独特靶点，但在揭示不同的遗传综合征方面可能效果较差。我们将重点关注与长QT综合征（LQT-3）和Brudaga综合征（BrS）相关的已鉴定的SCN5A突变，作为验证这一假设的范例。α亚基和位点定向突变的结构分析将补充遗传突变的分析，为解释通道功能的改变提供一个结构框架。这个项目有两个目的。目的1是验证与BrS或LQT-3相关的SCN5A基因遗传突变可能导致功能重叠的假设。目的2是验证遗传BrS和LQT-3 SCN5A突变特异性药理学可能由于突变诱导的表达通道门控改变的重叠而存在重叠的假设。提出的实验将结合膜片钳测量在哺乳动物细胞中瞬时表达的重组通道活性。我们的预测将通过结合膜片钳数据的离子通道门控和心脏动作电位的计算机模拟进行理论测试。我们假设从这些细胞和分子实验中获得的信息可以直接转化为基于突变基因产物的特定特性的人类治疗干预的改进，并且也揭示了这两种遗传性疾病可能的相互关系。

The overall goal of the research proposed in this application is to understand the molecular basis of cardiac arrhythmias caused, at least in part, by inherited mutations of the SCN5A gene, and to determine novel gene-targeted therapeutic strategies to treat them. The central hypothesis is that one step in the genesis of these arrhythmias is the perturbation of membrane electrical activity caused by alteration in the biophysical properties of the SCN5A gene product, the principal cardiac Na+ channel alpha subunit, by diseased-linked mutations, but that similar functional perturbations may be linked to distinct clinical disorders. Altered ion channel properties may also confer unique pharmacological properties upon the encoded ion channels making them unique targets for therapeutic intervention but, perhaps less effective in unmasking distinct inherited syndromes. We will focus on identified SCN5A mutations linked to the long QT syndrome (LQT-3) and Brudaga's syndrome (BrS) as paradigms to test this hypothesis. Structural analysis of the alpha subunit and site directed mutagenesis will complement the analysis of inherited mutations to provide a structural framework to interpret alteration in channel function. There are two aims of this project. Aim 1 is to test the hypothesis that there can be functional overlap caused by inherited mutations of the SCN5A gene linked either to BrS or LQT-3. Aim 2 is to test the hypothesis that there can be overlap in inherited BrS and LQT-3 SCN5A mutation-specific pharmacology due to overlap in mutation induced gating changes of expressed channels. Experiments that are proposed will combine patch clamp measurement of recombinant channel activity transiently expressed in mammalian cells. Theoretical testing of our predictions will be carried out using computer-based simulations of ion channel gating and cardiac action potentials that incorporate our patch clamp data. We hypothesize that information gained from these cellular and molecular experiments can be translated directly to improved therapeutic intervention in humans based on specific properties of mutant gene products, and also shed light on the possible interrelationship of these two inherited disorders.