Développement d'une stratégie thérapeutique combinée pour le traitement de la dystrophie musculaire de Duchenne

Duchenne muscular dystrophy (DMD) is a severe X-linked neuromuscular disorder caused by mutations in the DMD gene, leading to the absence of functional dystrophin and progressive muscle degeneration. Antisense oligonucleotides (ASOs) can restore the reading frame of the DMD transcript and promote the production of a truncated but functional dystrophin. However, their therapeutic efficacy remains limited, partly due to poor distribution to target tissues.The aim of this thesis was to enhance angiogenesis, which is impaired in DMD, in order to facilitate ASO distribution and potentiate their therapeutic effect. First, we evaluated a combined strategy associating an AAV9 vector expressing a novel VEGF-A isoform (LVRF) with a palm-tcDNA ASO targeting exon 23 in the mdx mouse model. Pretreatment with AAV-LVRF followed by 12 weeks of antisense therapy significantly improved the vascular network, increased ASO biodistribution by 3.8-fold, and enhanced exon skipping and dystrophin restoration by 80% and 50%, respectively, compared to ASO monotherapy. These effects were associated with an overall improvement of muscle pathology.In a second approach, we explored a non-viral strategy by targeting Flt-1, a decoy receptor of VEGF that limits angiogenesis. An ASO designed to induce a frameshift mutation and a siRNA directed against the soluble isoform sFlt1-i13 were evaluated in murine endothelial C166 cells. Both approaches efficiently downregulated Flt-1 at RNA and protein levels and promoted beneficial effects on cell proliferation, survival, and migration in vitro.Finally, we investigated a pharmacological approach using 20-hydroxyecdysone (20E), a molecule with pleiotropic effects on muscle and potential angiogenic activity. In the mdx model, the combination of ASO and 20E for eight weeks significantly improved functional performance (treadmill, inverted grid, and electrophysiology tests), without increasing dystrophin levels but inducing a marked anabolic effect on muscle fibers.Together, these findings highlight the relevance of combined therapeutic approaches in DMD and demonstrate that targeting the vascular and metabolic environment can enhance the efficacy of antisense-based therapies.