. 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.
Mitochondrial respiratory chain dysfunction occurs in an astonishingly frequent, varied and largely untreatable group of multi-systemic genetic disorders that afflict all ages and ethnicities. Translational research investigations in Caenorhabditis elegans (invertebrate worms) and Danio rerio (vertebrate zebrafish) animal models of RC disease may demonstrate effective combinations of pharmacologic and nutritional therapies, and their specific mechanisms, that restore cell and organ function toward normal to improve overall animal health.
