Phenotype rescue through the restoration of full-length dystrophin using CRISPR/Cas9 genome editing in Duchenne muscular dystrophy patient-derived iPSCs carrying the deletion of two exons

@article{rovina2026phenotype,
  title = {{Phenotype rescue through the restoration of full-length dystrophin using CRISPR/Cas9 genome editing in Duchenne muscular dystrophy patient-derived iPSCs carrying the deletion of two exons}},
  author = {Davide Rovina and Sofia Milasi and Giusy Di Giuseppe and Josè Manuel Pioner and Marianna Langione and Lucrezia Giammarino and Cecilia Ferrantini and Mattia Chiesa and Francesco Canonico and Domenico D’Amario and Eugenio Maria Mercuri and Giulio Pompilio and Elena Sommariva and Aoife Gowran and Angela Serena Maione},
  journal = {Genome Medicine},
  year = {2026},
  month = jul,
  doi = {10.1186/s13073-026-01738-3},
  url = {https://doi.org/10.1186/s13073-026-01738-3},
  note = {CC BY-NC-ND 4.0},
}

TY  - JOUR
TI  - Phenotype rescue through the restoration of full-length dystrophin using CRISPR/Cas9 genome editing in Duchenne muscular dystrophy patient-derived iPSCs carrying the deletion of two exons
AU  - Davide Rovina
AU  - Sofia Milasi
AU  - Giusy Di Giuseppe
AU  - Josè Manuel Pioner
AU  - Marianna Langione
AU  - Lucrezia Giammarino
AU  - Cecilia Ferrantini
AU  - Mattia Chiesa
AU  - Francesco Canonico
AU  - Domenico D’Amario
AU  - Eugenio Maria Mercuri
AU  - Giulio Pompilio
AU  - Elena Sommariva
AU  - Aoife Gowran
AU  - Angela Serena Maione
PY  - 2026
DA  - 2026/07/31
JO  - Genome Medicine
DO  - 10.1186/s13073-026-01738-3
UR  - https://doi.org/10.1186/s13073-026-01738-3
AB  - Background: Duchenne muscular dystrophy (DMD) is a X-linked disease affecting skeletal and cardiac muscle and is caused by mutations in the dystrophin gene (DMD). Patient-derived induced pluripotent stem cells (iPSCs) serve as reliable in vitro disease models. Their genetic correction by CRISPR/Cas9 allows the generation of isogenic controls and holds promises for gene therapy. However, restoring full-length dystrophin, especially when deletions involve multiple exons, constitutes a technological challenge. This study aimed to fully repair the dystrophin gene from a DMD iPSC line carrying the deletion of exons 49–50 and to characterize the rescue of the cardiac phenotype.  Methods: We developed an innovative CRISPR/Cas9-based approach involving the insertion of coding sequences of the deleted region, at the 3’ of exon 48, thereby generating a single continuous coding sequence encompassing exons 48-49-50. Subsequently, iPSCs were differentiated into cardiomyocytes and cardiac fibroblasts. Cardiac phenotypes were analysed by western blot, immunofluorescence, ELISA, FACS, Ionoptix, 3D engineered heart tissue (EHT) and single-nuclei RNA-seq.  Results: The correction of a two-exons DMD gene deletion in Duchenne iPSCs, using CRISPR/Cas9, enabled the re-expression of a stable and functional full-length dystrophin in cardiomyocytes resulting in the rescue of cardiac pathological phenotypes. Edited cardiomyocytes showed improved morphology, reduced release of the cardiomyocytes damage marker troponin I, and decreased ROS production. Moreover, dystrophin restoration enhanced contractility and ameliorated the Ca2+ kinetics. Notably, edited iPSC derived fibroblasts showed reduced pro-fibrotic stimuli response. In parallel, we also observed enhanced functioning of a 3D engineered heart tissue and profound change in the transcriptomic profile in both cardiomyocytes and fibroblasts after the re-expression of full-length dystrophin.  Conclusions: We developed an innovative approach that enabled the re-expression of full-length dystrophin in a DMD iPSC line with consequent complete rescue of in vitro DMD cardiac phenotypes. On the long term, these results could lay a foundation for future applications of cell therapy or in vivo CRISPR/Cas9-based intervention to treat DMD.  Supplementary Information: The online version contains supplementary material available at https://doi.org/10.1186/s13073-026-01738-3.
SN  - 1756-994X
ER  -