Specific DMPK-promoter targeting by CRISPRi reverses myotonic dystrophy type 1-associated defects in patient muscle cells

Florent Porquet (1, 2) , Lin Weidong (3) , Kévin Jehasse (2) , Hélène Gazon (3) , Maria Kondili (1) , Silvia Blacher (3) , Laurent Massotte (2) , Emmannuel Di Valentin (3) , Denis Furling (1) , Nicolas Albert Gillet (4) , Arnaud François Klein (1) , Vincent Seutin (3) , Luc Willems (2)

1 Centre de recherche en Myologie – U974 SU-INSERM
2 GIGA-Neurosciences [Université Liège]
3 GIGA [Université Liège]
4 NARILIS - Namur Research Institute for Life Sciences

Myotonic dystrophy type 1 (DM1) is a neuromuscular disease that originates from an expansion of CTG microsatellites in the 3 0 untranslated region of the DMPK gene, thus leading to the expression of transcripts containing expanded CUG repeats (CUGexp). The pathophysiology is explained by a toxic RNA gain of function where CUGexp RNAs form nuclear aggregates that sequester and alter the function of MBNL splicing factors, triggering splicing misregulation linked to the DM1 symptoms. There is currently no cure for DM1, and most therapeutic strategies aim at eliminating CUGexp-DMPK transcripts. Here, we investigate a DMPK-promoter silencing strategy using CRISPR interference as a new alternative approach. Different sgRNAs targeting the DMPK promoter are evaluated in DM1 patient muscle cells. The most effective guides allowed us to reduce the level of DMPK transcripts and CUGexp-RNA aggregates up to 80%. The CUGexp-DMPK repression corrects the overall transcriptome, including spliceopathy, and reverses a physiological parameter in DM1 muscle cells. Its action is specific and restricted to the DMPK gene, as confirmed by genomewide expression analysis. Altogether, our findings highlight DMPK-promoter silencing by CRISPRi as a promising therapeutic approach for DM1.

Mots clés: Myotonic Dystrophy; gene therapy; CRISPRi; Cellular model

Mots clés: Dystrophie Mytotonique de type 1; Thérapie génique; Modèles cellulaires

Domaines: Sciences du Vivant [q-bio]; Biologie moléculaire

Florent Porquet, Lin Weidong, Kévin Jehasse, Hélène Gazon, Maria Kondili, et al.. Specific DMPK-promoter targeting by CRISPRi reverses myotonic dystrophy type 1-associated defects in patient muscle cells. Molecular Therapy - Nucleic Acids, 2023, 32, pp.857 - 871. ⟨10.1016/j.omtn.2023.05.007⟩. ⟨hal-04287597⟩