5-aza-2-deoxycytidine improves skeletal muscle function in a mouse model for recessive RYR1-related congenital myopathy

RYR1 congenital myopathies are rare disorders severely impacting muscle function and impairing the quality of life of patients and their families, yet to date, no pharmacological therapies are available to treat the severe muscle weakness of affected patients. The recessive forms of RYR1-related congenital myopathies caused by nonsense/ frameshift mutations in one allele and a missense mutation in the other allele, resulting in the homozygous and reduced expression of the missense mutation are the most severe forms. They are often accompanied by a reduction of RyR1 protein and significant changes in the biochemical composition of muscles. Using a mouse model carrying the RyR1 p.Q1970fsX16+p.A4329D compound heterozygous mutations (dHT mice),we performed a pre-clinical study and injected mice for 15 weeks with 0.05 mg/kg of the FDA approved drug 5-aza-2-deoxycytidine which targets the epigenetic enzymes DNA methyltransferases. We evaluated muscle strength, calcium homeostasis and proteome and report that drug treatment improves all investigated muscle function in drug treated dHT mice. Importantly, the beneficial effects were particularly significant in fast twitch muscles which are the first muscles to be impaired in patients. In conclusion, this study provides proof of concept for the pharmacological treatment of patients with congenital myopathies linked to recessive RYR1 mutations, leading the path to a phase 1 clinical study by repurposing an FDA approved drug. We aimed to investigate the effects of 5-aza-2'-deoxycytidine on the methylation status of skeletal muscles in a congenital myopathy model. A mouse model was developed, including both wild-type (WT) and heterozygous RYR1 mutation (dHT) carriers. Genomic DNA from 45 samples was analyzed using the Infinium Mouse Methylation Array (Illumina). These samples consisted of 15 from WT mice and 30 from dHT mice, divided into vehicle-treated and TMP269-5Aza-treated groups. Methylation patterns were profiled across three distinct muscle types: extensor digitorum longus (EDL), flexor digitorum brevis (FDB), and soleus (SOL).