Résumé :
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In Duchenne Muscular Dystrophy (DMD) the selective removal by exon skipping of exons flanking an out-of frame mutation in the dystrophin messenger can result in in-frame mRNA transcripts that are translated into shorter but functionally active dystrophin.The goal of our project is to determine in Golden Retriever Muscular Dystrophy (GRMD) dogs the effective dose of our therapeutic product defined as a recombinant Adeno-Associated Virus serotype 8 (rAAV8) expressing a optimized U7 snRNA specific for the correction by exon skipping of the GRMD dystrophin transcript. The mode of delivery is the locoregional high-pressure intravenous (IV) injection of a forelimb.We established a unique network of laboratories with complementary skills to deliver a comprehensive set of preclinical data to determine the therapeutic dose intended to be used in clinic. The primary outcomes are the restoration of dystrophin expression and the improvement of the tissue pathology in the injected limb compared to the controlateral limb. Among our secondary outcomes the biodistribution and shedding patterns are of particular importance to precisely determine the safety of our locoregional administration in skeletal muscle.Several groups of GRMD dogs were exposed to different rAAV8-U7snRNA doses (2,5E12 to 2,5E13 vg/kg). To assess the biodistribution and shedding patterns of the vector, biological fluids were repeatedly collected as soon as 6 hours post-injection and subsequently for weeks. Ultimately, all animals were sacrificed _3 months after injection of the vector. All muscles of the injected limb and of the contralateral limb were precisely sampled, as well as several skeletal muscles at distance (including heart and diaphragm) and major organs (including liver, spleen, kidneys, lungs, gonads and lymph nodes).The complete biodistribution and shedding data, obtained after quantitative PCR analysis of these different samples will be presented.This project is supported by AFM (Association Franse contre les Myopathies) and by ADNA (Advanced Diagnostics for New Therapeutic Approaches), a program dedicated to personalized medicine, coordinated by Institut Meux and supported by research and innovation aid from the French public agency, OSEO.
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