Evaluation of small compounds to enhance exon-skipping in Duchenne Muscular Dystrophy

My thesis project was focused on dysferlin, a protein mainly expressed in muscle. There is some evidence that dysferlin is involved in several processes like T-tubule homeostasis, myoblast-myotubes fusion and membrane repair, even if these roles are not clearly demonstrated yet. Mutations in DYSF have been associated with Limb Girdle Muscular Dystrophies (type 2B) and Miyoshi Myopathy. The wide spectrum of mutations in DYSF, the absence of mutational hotspot and the size of the coding sequence force us to consider different therapeutic strategies. The major part of DYSF encodes evolutionarily conserved C2 domains which are well characterized Ca2+ sensor domains that mainly play a role in vesicle fusion process. Nevertheless, very few evidences have been showed regarding the specific functions of each of the 7 dysferlin C2 domains. Based on two natural proofs of concept, we have designed an exon-skipping strategy and a mini-gene based approach. These strategies while having a therapeutic purpose allow us to evaluate the modularity of dysferlin. To further understand how the biochemical properties of each domain may influence the functionality of the full length protein, and thus how some of them are essential, I performed biological and biochemical analysis and identified dysferlin functions that are specific for some of the motifs.