Study of pathogenetic mechanisms and development of new therapeutic strategies for Myotonic Dystrophy type 1

Myotonic dystrophy type 1 (DM1) is a dominantly inherited neuromuscular disease caused by the abnormal expansion of CTG-triplets in the 3' untranslated region of the DMPK gene that is transcribed in toxic CUG-repeated mRNA. Although the molecular mechanisms of DM1 have been studied for many years, the role of recently identified regulatory RNAs, such as microRNAs and circular RNAs (circRNAs), is only starting to be clarified. Available therapeutic approaches aimed at neutralizing the toxic RNA provide only short-term effects and do not eliminate the repeats from the genome. Our research group is carrying out two parallel projects on DM1: 1)identify the role of circRNAs and their use as biomarkers (grant 23054 from AFM Telethon, France) New circRNAs dysregulated in DM1 patients were identified that might be used as DM1 biomarkers. Functional characterisation of one of these DM1-circRNAs, circARHGAP10, highlighted its potential role as a sponge for hsa-miR-409-3p. 2)develop a CRISPR/Cas9-mediated gene therapy to delete the expansion mutation (grant GGP19035, Telethon Italy) We have generated and transduced highly specific and inducible CRISPR/Cas9 components in myogenic cells derived from patients affected by DM1 and obtained the permanent removal of the pathogenetic CTG expansion and partial reversion of molecular alterations in edited cells. The occurrence of off-targets and on-target unintended editing events was evaluated. AAV-mediated transduction of the CRISPR/Cas9 components in DM1 mice led to CTG repeat deletions in the skeletal and cardiac muscles and in partial rescue of some DM1-associated molecular and phenotypic alterations.