Pharmacologic Treatment of Mitochondrial Complex I Dysfunction in C. Elegans

项目总结。线粒体呼吸链(RC)病导致高血压病患者多系统衰竭 能量需求器官，导致神经发育、心脏、肌肉、视力和/或听力障碍 全球新陈代谢不稳定。而减少线粒体内疾病异常的经验性方法 已经被证明是普遍无效的，我们建议靶向线粒体外的生理效应 由原发RC功能障碍引起的可能改善健康结局。在母公司R01奖励期间，我们 优化了线虫、线虫和斑马鱼D. RERIO用于询问RC的发病机制并评价候选药物的疗效和毒性 活体动物的疗法。最重要的是，我们发现中枢结节的药物靶向 一个整合的营养感应信号网络，以及调节蛋白毒素的基本细胞过程 压力，可以显著改善整体细胞、器官和动物的健康。我们已经确定了一种新的武器库 在显著改善复合体I(CI)缺乏气体的短暂寿命的17种单独药物和营养物质中- 1条(Fc21)突变蠕虫。本R01竞争更新提案的总体目标是阐明准确的 药物调节RC功能障碍的代谢后果的机制， 利用线虫(无脊椎动物)和斑马鱼(脊椎动物)的固有优势 模特们。我们假设，线粒体RC的细胞后果的药物调节 功能障碍将为RC疾病的常见亚组提供有效的治疗方法，无论个体 致病原因。这项提议的具体目标是：[目标1]调查多种药物治疗的疗效 在线虫主要RC疾病亚型的模型中；[AIM 2]以定义特定的NSSN节点和调节 在GAS-1(FC21)线虫模型中介导治疗效果的细胞蛋白毒性应激的成分 [目的]评价CI缺陷型斑马鱼器官水平的联合治疗效果。 模特。方法包括测试铅药物疗法的合理组合对线虫菌株寿命的影响 缺乏不同的RC复合体，以及组合的RC突变和中心NSSN节点突变的蠕虫。 健康跨度效应将通过生物途径的转录组图谱和体内定量的 线粒体生理学，NAD+和NADH水平，以及中间代谢流量。靶向表达 活体动物的图谱和荧光分析将用于量化NSSN节点的活动和 调节蛋白毒性应激的主要细胞通路。器官层面的治疗效果将在#年进行评估 斑马鱼RC病模型的显微镜和体内线粒体生理学主要集中在脑， 心脏和肌肉。综上所述，拟议的研究对于阐明药物和 RC病的营养性治疗及其合理组合，最终将全面改善 人类线粒体疾病表现多样的患者的健康和临床结局。

PROJECT SUMMARY. Mitochondrial respiratory chain (RC) diseases cause multi-system failure of high- energy demand organs, with resulting neurodevelopmental, cardiac, muscle, vision, and/or hearing disabilities with global metabolic instability. While empiric approaches to reduce intra-mitochondria disease aberrations have proven generally ineffective, we propose that targeting the extra-mitochondrial physiologic effects induced by primary RC dysfunction may improve health outcomes. During the parent R01 award period we optimized a robust cadre of disease models and in vivo assays in the nematode C. elegans and zebrafish D. rerio with which to interrogate RC disease mechanisms and evaluate the efficacy and toxicity of candidate drug therapies in living animals. Most importantly, we discovered that pharmacologic targeting of central nodes in an integrated nutrient-sensing signaling network, and of basic cellular processes that regulate proteotoxic stress, can significantly improve overall cellular, organ, and animal health. We have identified a novel arsenal of 17 individual drugs and nutrients that significantly improve the short lifespan of complex I (CI) deficient gas- 1(fc21) mutant worms. The overall goal of this R01 Competing Renewal proposal is to elucidate precise mechanisms by which pharmacologic agents modulate the metabolic consequences of RC dysfunction, capitalizing on the inherent advantages of both C. elegans (invertebrate) and zebrafish (vertebrate) animal models. We hypothesize that pharmacologic modulation of the cellular consequences of mitochondrial RC dysfunction will offer effective therapies for common subgroups of RC disease, irrespective of individual pathogenic cause. Specific Aims of this proposal are: [AIM 1] To investigate efficacy of multi-drug treatments in C. elegans models of major RC disease subtypes; [AIM 2] To define specific NSSN nodes and regulatory components of cellular proteotoxic stress that mediate treatment efficacy in the gas-1(fc21) C. elegans model of CI disease; and [AIM 3] To evaluate organ-level combinatorial therapy effects in the CI-deficient zebrafish model. Methods include testing rational combinations of lead drug therapies on lifespan in C. elegans strains deficient in different RC complexes, as well as in combined RC mutant and central NSSN node mutant worms. Healthspan effects will be studied by transcriptome profiling of biological pathways and in vivo quantitation of mitochondrial physiology, NAD+ and NADH levels, and intermediary metabolic flux. Targeted expression profiling and fluorescence analyses in living animals will be used to quantify activities of the NSSN nodes and major cellular pathways that regulate proteotoxic stress. Organ-level treatment effects will be evaluated in zebrafish models of RC disease by microscopy and in vivo mitochondrial physiology focused primarily on brain, heart, and muscle. In summary, the proposed studies are essential to elucidate mechanisms of drug and nutrient-based treatments for RC disease, and their rational combinations, which will ultimately improve overall health and clinical outcomes in human patients with the diverse manifestations of mitochondrial disease.