Abstract 123 Preclinical CRISPR treatments to correct Duchenne Muscular Dystrophy in patient-specific cardiomyocytes and tissues

Clin Res Cardiol (2022). https://doi.org/10.1007/s00392-022-02002-5
Preclinical CRISPR treatments to correct Duchenne Muscular Dystrophy in patient-specific cardiomyocytes and tissues
O. Gutierrez-Gutierrez¹, O. Richter¹, M. Tiburcy², M. Shahriyari², E. Schoger², W.-H. Zimmermann², L. Cyganek¹, for the study group: DZHK
¹Herzzentrum Göttingen - Stem Cell Unit, Universitätsmedizin Göttingen, Göttingen; ²Institut für Pharmakologie und Toxikologie, Universitätsmedizin Göttingen, Göttingen;
Duchenne muscular dystrophy (DMD) is a rare disease characterized by progressive muscle weakness and muscle loss, which leads to respiratory disabilities and cardiomyopathies during adulthood and ultimately death. This X-linked recessive disorder has an incidence of 1 in 3,500 males and is caused by mutations in the dystrophin gene. Dystrophin is a giant protein expressed mainly in muscle cells whose function is to stabilize the muscle fibers by connecting the cytoskeleton to the extracellular matrix. It has been reported more than 5,000 different DMD mutations, mostly out-of-frame deletions of one or more exons, which give rise to frameshift, premature termination of translation and lack of dystrophin. Interestingly, in-frame deletions resulting in truncated functional dystrophins can give rise to milder muscular dystrophies or even asymptomatic clinical phenotypes. In our study, we aimed to develop personalized CRISPR/Cas-based gene therapy strategies in DMD patients, converting out-of-frame deletions in in-frame deletions by exon skipping. To achieve this, patient-specific induced pluripotent stem cells (iPSCs) from DMD patients were generated, differentiated into cardiac cells and phenotyped at the cellular and tissue level. Patients’ iPSC-derived cardiomyocytes (iPSC-CMs) confirmed disrupted reading frame, lack of dystrophin protein and severely reduced contractile performance. In order to induce exon skipping and reframing of the coding sequence, different CRISPR/Cas9 genome editing approaches targeting the acceptor splice site of adjacent exons were tested. Thus, corrected iPSC-CMs displayed efficient exon skipping and restoration of dystrophin expression. Excitingly, engineered heart muscle generated from corrected cardiomyocytes presented a restoration of the contractile function. Altogether, the use of patient-specific iPSC-CMs and engineered heart muscle represent an excellent preclinical model to test different clinically translatable CRISPR/Cas9-based exon skipping strategies for treating DMD.
https://dgk.org/kongress_programme/jt2022/aV940.html