Therapeutic targeting of mitochondrial dysfunction in FSHD

Facioscapulohumeral dystrophy (FSHD) is an incurable, hereditary disease primarily affecting skeletal musculature. Clinically, FSHD is characterized by progressive muscle weakness and wasting. The genetic cause of FSHD are mutations that lead to production of the protein DUX4, which is usually not made in muscle. When DUX4 is erroneously present in muscle it triggers muscle degeneration. To date, it is unclear precisely how DUX4 causes muscle disease. However, several studies suggest that FSHD patients present with impaired muscle metabolism, suffering from a condition termed metabolic stress. From the metabolic perspective, skeletal muscle is a highly active tissue and generating enough energy to fulfill its many functions is of crucial importance for muscle health. Consequently, any chronic perturbance of the complex bioenergetic mechanisms necessary for working muscles to meet their metabolic demand causes functional muscle impairment over time. Mitochondria, small energy factories inside cells, are essential to supply muscle with enough energy so it can perform work (for example contraction). Any failure of mitochondria to function efficiently causes a state of energy crisis over time, leading to subsequent muscle degeneration. We have investigated the role of DUX4 in muscle cells from FSHD patients and found that DUX4 changes muscle cell metabolism so that large amounts of radicals, small but highly reactive and potentially damaging molecules, are made. Radicals are an inevitable byproduct of metabolism, which have to be properly controlled and detoxified by the muscle cells to avoid damage to the cells. Increased radical levels in FSHD interfere with muscle metabolism and lead to muscle impairment, especially when they disturb how muscle utilizes oxygen. Oxygen is an important metabolite in muscle, and local oxygen availability can fluctuate greatly, for example when we exercise. A complex interplay between mitochondria, radicals and oxygen sensing enables muscle to adapt to metabolic and bioenergetic fluctuations, and our preliminary analyses hint at a deregulation of mitochondrial function through radical-induced impairment of oxygen sensing. Importantly, we found that antioxidants, substances which can scavenge radicals and prevent them from causing damage in the cell, are effective in restoring FSHD muscle function. Antioxidant therapy has previously been tested in FSHD patients and a recent clinical trial found moderate muscle functional improvement. However, therapeutic efficacy of antioxidants could likely be improved if we better understand the mechanisms causing metabolic stress in FSHD. This project will investigate the bioenergetic and metabolic differences between healthy and FSHD muscle models to find novel therapeutic entry points. Specifically, we aim at deciphering the relationship between mitochondria, radicals, oxygen sensing and muscle function. Many antioxidants are already available as approved dietary supplements, so could be readily translated into clinics. Thus, our investigation of FSHD muscle metabolism will both broaden our understanding of how DUX4 causes FSHD, and identify new refined antioxidant-based therapeutics to complement and support more experimental therapies directed at reducing DUX4 levels.

Facioscapulohumeral muscular dystrophy (FSHD) is a prevalent, incurable genetic muscle disease characterised by progressive muscle wasting and loss. While the genetic mutations underlying FSHD are identified, the biological mechanisms driving the disease are not fully understood. However, it is known that FSHD muscles suffer from impaired energy-producing processes, a condition called metabolic stress. This stress can arise when mitochondria, the cell's powerhouses, malfunction, leading to an imbalance in highly reactive molecules called reactive oxygen species (ROS). Excessive ROS production results in oxidative stress, damaging cells and ultimately causing cell death. Muscles affected by FSHD show metabolic disruptions, and while mitochondrial dysfunction and oxidative stress contribute to muscle damage in the disease, particularly driven by FSHD-causative DUX4 protein, the specific underlying mechanisms are not well-defined. This project aimed to elucidate how metabolic stress contributes to FSHD. We focused on how changes in muscle metabolism linked to the DUX4 protein influence disease progression and sought to identify new therapeutic strategies. Using muscle cells from FSHD patients and cells with inducible DUX4 expression, we examined the role of mitochondrial reactive oxygen species (mitoROS) in disrupting mitochondrial metabolism and increasing sensitivity to low oxygen availability (hypoxia). Our work revealed that DUX4 expression in muscle cells leads to immediate mitochondrial changes, notably increased mitoROS production. This effect precedes cell death and is pronounced in hypoxia. Supplementation with mitochondria-targeted antioxidants significantly alleviated the effects of DUX4, such as increased cell death and muscle cell shrinkage (hypotrophy), particularly in hypoxia. These targeted antioxidants were more effective than conventional ones, highlighting the importance of directly acting on mitochondrial dysfunction. Further, we identified mitochondria as the primary source of oxidative stress in FSHD, as they excessively produce mitoROS from specific sites within the mitochondrial respiratory chain. This finding links mitochondrial dysfunction directly to oxidative stress. We explored the therapeutic potential of respiratory chain-targeted antioxidants like mitoQ10, which proved effective in reducing oxidative stress and preserving mitochondrial function compared to other antioxidant approaches. Additionally, we investigated the impact of disrupted nitric oxide (NO) signalling in FSHD. Excessive mitoROS production in FSHD mitochondria was found to decrease NO levels, impairing the muscle's ability to produce new mitochondria. Reduced NO signalling was associated with diminished mitochondrial mass and function in FSHD muscle cells. Targeting NO-responsive pathways with approved drugs showed promise in alleviating oxidative stress and improving mitochondrial function, offering a new therapeutic avenue. In conclusion, this research identified oxidative stress through mitochondrial dysfunction as a key mechanism in FSHD. Importantly, this mechanism can be targeted with drugs. Our findings are further supported by transcriptomic analyses of FSHD muscle biopsies, which revealed widespread impairment of genes related to mitochondrial function, oxidative stress, and NO signalling, laying the groundwork for novel therapeutic strategies.