Combinatorial glucose, nicotinic acid and N-acetylcysteine therapy has synergistic effect in preclinical C. elegans and zebrafish models of mitochondrial complex I disease.

Mitochondrial respiratory chain disorders are empirically managed with variable antioxidant, cofactor and vitamin 'cocktails'. However, clinical trial validated and approved compounds, or doses, do not exist for any single or combinatorial mitochondrial disease therapy. Here, we sought to pre-clinically evaluate whether rationally designed mitochondrial medicine combinatorial regimens might synergistically improve survival, health and physiology in translational animal models of respiratory chain complex I disease. Having previously demonstrated that gas-1(fc21) complex I subunit ndufs2-/-C. elegans have short lifespan that can be significantly rescued with 17 different metabolic modifiers, signaling modifiers or antioxidants, here we evaluated 11 random combinations of these three treatment classes on gas-1(fc21) lifespan. Synergistic rescue occurred only with glucose, nicotinic acid and N-acetylcysteine (Glu + NA + NAC), yielding improved mitochondrial membrane potential that reflects integrated respiratory chain function, without exacerbating oxidative stress, and while reducing mitochondrial stress (UPRmt) and improving intermediary metabolic disruptions at the levels of the transcriptome, steady-state metabolites and intermediary metabolic flux. Equimolar Glu + NA + NAC dosing in a zebrafish vertebrate model of rotenone-based complex I inhibition synergistically rescued larval activity, brain death, lactate, ATP and glutathione levels. Overall, these data provide objective preclinical evidence in two evolutionary-divergent animal models of mitochondrial complex I disease to demonstrate that combinatorial Glu + NA + NAC therapy significantly improved animal resiliency, even in the face of stressors that cause severe metabolic deficiency, thereby preventing acute neurologic and biochemical decompensation. Clinical trials are warranted to evaluate the efficacy of this lead combinatorial therapy regimen to improve resiliency and health outcomes in human subjects with mitochondrial disease.

Pre-clinical evaluation of cysteamine bitartrate as a therapeutic agent for mitochondrial respiratory chain disease.

Cysteamine bitartrate is a US Food and Drug Administration-approved therapy for nephropathic cystinosis also postulated to enhance glutathione biosynthesis. We hypothesized this antioxidant effect may reduce oxidative stress in primary mitochondrial respiratory chain (RC) disease, improving cellular viability and organismal health. Here, we systematically evaluated the therapeutic potential of cysteamine bitartrate in RC disease models spanning three evolutionarily distinct species. These pre-clinical studies demonstrated the narrow therapeutic window of cysteamine bitartrate, with toxicity at millimolar levels directly correlating with marked induction of hydrogen peroxide production. Micromolar range cysteamine bitartrate treatment in Caenorhabditis elegans gas-1(fc21) RC complex I (NDUFS2-/-) disease invertebrate worms significantly improved mitochondrial membrane potential and oxidative stress, with corresponding modest improvement in fecundity but not lifespan. At 10 to 100 µm concentrations, cysteamine bitartrate improved multiple RC complex disease FBXL4 human fibroblast survival, and protected both complex I (rotenone) and complex IV (azide) Danio rerio vertebrate zebrafish disease models from brain death. Mechanistic profiling of cysteamine bitartrate effects showed it increases aspartate levels and flux, without increasing total glutathione levels. Transcriptional normalization of broadly dysregulated intermediary metabolic, glutathione, cell defense, DNA, and immune pathways was greater in RC disease human cells than in C. elegans, with similar rescue in both models of downregulated ribosomal and proteasomal pathway expression. Overall, these data suggest cysteamine bitartrate may hold therapeutic potential in RC disease, although not through obvious modulation of total glutathione levels. Careful consideration is required to determine safe and effective cysteamine bitartrate concentrations to further evaluate in clinical trials of human subjects with primary mitochondrial RC disease.

Aberrant autolysosomal regulation is linked to the induction of embryonic senescence: differential roles of Beclin 1 and p53 in vertebrate Spns1 deficiency.

Spinster (Spin) in Drosophila or Spinster homolog 1 (Spns1) in vertebrates is a putative lysosomal H+-carbohydrate transporter, which functions at a late stage of autophagy. The Spin/Spns1 defect induces aberrant autolysosome formation that leads to embryonic senescence and accelerated aging symptoms, but little is known about the mechanisms leading to the pathogenesis in vivo. Beclin 1 and p53 are two pivotal tumor suppressors that are critically involved in the autophagic process and its regulation. Using zebrafish as a genetic model, we show that Beclin 1 suppression ameliorates Spns1 loss-mediated senescence as well as autophagic impairment, whereas unexpectedly p53 deficit exacerbates both of these characteristics. We demonstrate that 'basal p53' activity plays a certain protective role(s) against the Spns1 defect-induced senescence via suppressing autophagy, lysosomal biogenesis, and subsequent autolysosomal formation and maturation, and that p53 loss can counteract the effect of Beclin 1 suppression to rescue the Spns1 defect. By contrast, in response to DNA damage, 'activated p53' showed an apparent enhancement of the Spns1-deficient phenotype, by inducing both autophagy and apoptosis. Moreover, we found that a chemical and genetic blockage of lysosomal acidification and biogenesis mediated by the vacuolar-type H+-ATPase, as well as of subsequent autophagosome-lysosome fusion, prevents the appearance of the hallmarks caused by the Spns1 deficiency, irrespective of the basal p53 state. Thus, these results provide evidence that Spns1 operates during autophagy and senescence differentially with Beclin 1 and p53.

Supplement timing of cranberry extract plays a key role in promoting Caenorhabditis elegans healthspan.

Consumption of nutraceuticals is a major and potent dietary intervention for delaying aging. As the timing of administration is critical for the efficacy of bioactive compounds in medicine, the effectiveness of nutraceuticals may also be dramatically affected by the timing of supplementation. Cranberry exact (CBE), rich in polyphenols, is consumed as a nutraceutical, and possesses anti-aging properties. Here, we examined the influence of timing on the beneficial effects of CBE supplementation in C. elegans. The prolongevity effect of CBE in different aged worms, young adults, middle-age adults, and aged adults, was determined. Early-start intervention with CBE prolonged the remaining lifespan of worms of different ages more robustly than late-start intervention. The effectiveness of CBE on stress responses and physiological behaviors in different aged worms was also investigated. The early-start intervention prominently promoted motility and resistance to heat shocks and V. cholera infection, especially in aged worms. Together, these findings suggest that the timing of CBE supplementation critically influences its beneficial effects on C. elegans lifespan and healthspan. It is of interest to further investigate whether the similar results would occur in humans.

Nutraceutical interventions for promoting healthy aging in invertebrate models.

Aging is a complex and inevitable biological process that is associated with numerous chronically debilitating health effects. Development of effective interventions for promoting healthy aging is an active but challenging area of research. Mechanistic studies in various model organisms, noticeably two invertebrates, Caenorhabditis elegans and Drosophila melanogaster, have identified many genes and pathways as well as dietary interventions that modulate lifespan and healthspan. These studies have shed light on some of the mechanisms involved in aging processes and provide valuable guidance for developing efficacious aging interventions. Nutraceuticals made from various plants contain a significant amount of phytochemicals with diverse biological activities. Phytochemicals can modulate many signaling pathways that exert numerous health benefits, such as reducing cancer incidence and inflammation, and promoting healthy aging. In this paper, we outline the current progress in aging intervention studies using nutraceuticals from an evolutionary perspective in invertebrate models.