Pharmacologic treatment of mitochondrial complex I dysfunction in C. elegans

描述（由申请人提供）：线粒体呼吸链（RC）复合体I功能障碍是一种非常常见的多系统疾病的基础，这种疾病折磨着所有年龄和种族。相对无法客观评估介导多种疾病表现的细胞机制，这在很大程度上阻碍了对受影响患者进行有效治疗的评估和实施。我们假设，无论个体致病原因如何，对线粒体RC功能障碍的细胞后果进行药理学调节将为RC功能障碍的常见亚群提供有效的治疗方法。本提案的总体目标是阐明药物调节RC功能障碍代谢后果的机制，利用秀丽隐杆线虫动物模型提供的固有研究优势。本提案的具体目的是：SA1：确定秀丽隐杆线虫中PPAR/SIRT1通路的药理学调节是否会减弱甚至逆转复合物I功能障碍的代谢后果；和SA2：在翻译秀丽隐杆线虫复合体I功能障碍动物模型中，表征“线粒体鸡尾酒”成分在体内的影响。我们将分别在SA1和SA2的RC复合物I突变体中研究直接调节PPAR/ sirt1相关信号通路的五种药物，以及四类（维生素、抗氧化剂、复合物I翻译后激活剂和中间代谢调节剂）中的十五种常见的“线粒体鸡尾酒”成分。将进行生物信息学整合，以辨别每种药物对5种体内表型的总体功效，以实现两个目标：(a)寿命；(b)表达微阵列分析代谢途径谱；(c)利用稳定同位素通过中间代谢途径的通量；(d)荧光显微镜下线粒体膜电位；(e)荧光显微镜下线粒体相对氧化负荷。通过RNA干扰诱导PPAR/SIRT1通路基因敲低的复合物I突变体的体内功能分析，将探索可识别表型反应的机制。通过这种方式，目前缺乏有效或有害的科学或临床证据的假定对人类RC疾病有益的药物可以在秀丽隐杆线虫中以低成本和高通量的方式客观和非侵入性地筛选。这项转化研究可能为减轻人类线粒体疾病的全球后遗症提供特定的治疗靶点和潜在有效的药物。

DESCRIPTION (provided by applicant): Dysfunction of complex I in the mitochondrial respiratory chain (RC) underlies an astonishingly frequent group of multi-systemic disorders that afflict all ages and ethnicities. A relative inability to objectively assess cellular mechanisms that mediate widely variable disease manifestations has largely prohibited evaluation and implementation of effective therapies in affected patients. We hypothesize that pharmacologic modulation of the cellular consequences of mitochondrial RC dysfunction will offer effective therapies for common subgroups of RC dysfunction, irrespective of individual pathogenic cause. The overall goal of this proposal is to elucidate mechanisms by which pharmacologic agents modulate the metabolic consequences of RC dysfunction, capitalizing on the inherent investigative advantages that the C. elegans animal model provides. The Specific Aims of this proposal are to: SA1: Determine whether pharmacologic modulation of the PPAR/SIRT1 pathway in C. elegans will attenuate, or even reverse, the metabolic consequences of complex I dysfunction; and SA2: Characterize in vivo consequences of "mitochondrial cocktail" components in a translational C. elegans animal model of complex I dysfunction. Five pharmacologic agents that directly modulate PPAR/SIRT1-related signaling pathways as well as fifteen common "mitochondrial cocktail" components in four major classes (vitamins, antioxidants, complex I post- translational activator, and intermediary metabolic modifiers) will be studied in RC complex I mutants in SA1 and SA2, respectively. Bioinformatics integration will be performed to discern the overall efficacy of each pharmacologic agent on 5 in vivo phenotypes for both aims: (a) lifespan; (b) metabolic pathway profiling by expression microarray analysis; (c) flux through intermediary metabolic pathways using stable isotopes; (d) mitochondrial membrane potential by fluorescence microscopy; and (e) relative mitochondrial oxidant burden by fluorescence microscopy. Mechanisms underlying discernible phenotypic responses will be explored by in vivo functional analyses in complex I mutants harboring RNA interference-induced knockdown for individual PPAR/SIRT1 pathway genes. In this manner, drugs having postulated benefit for human RC disease that currently lack scientific or clinical evidence of either efficacy or harm can be objectively and non- invasively screened in a low-cost and high-throughput fashion in C. elegans. This translational research may suggest specific therapeutic targets and potentially effective pharmacologic agents to mitigate the global sequelae of human mitochondrial disease. PUBLIC HEALTH RELEVANCE: Mitochondrial complex I dysfunction occurs in an astonishingly frequent, varied, and largely untreatable group of genetic disorders afflicting all ages and ethnicities. Caenorhabditis elegans offers a robust genetic model in which to assess the global impact of complex I dysfunction and therapeutic candidates. This translational research may demonstrate effective pharmacologic therapies, and their specific mechanisms, to potentially mitigate the secondary consequences of human mitochondrial disease.