{"paper_id":"42537b4a-14a9-4e57-97a5-fb5eebd544ba","body_text":"Abstract\nExon skipping is a leading therapeutic approach for Duchenne muscular dystrophy (DMD), a progressive muscle wasting disorder caused by pathogenic variants in the DMD gene that typically disrupt the translation reading frame. This approach aims to modulate DMD pre-mRNA splicing to re-frame the transcript and generate an internally deleted but partially functional quasi-dystrophin protein. Four exon skipping drugs have received FDA accelerated approval, although their clinical efficacy is very limited. To investigate how treatment timing influences exon skipping outcomes, dystrophin-deficient mdx mice were injected with peptide-phosphorodiamidate morpholino oligonucleotide (PPMO) exon skipping conjugates beginning at either the adult (12-week) or aged (75-week) stages, and tibialis anterior muscles harvested for biochemical and transcriptomic analyses. Mean Dmd exon 23 skipping was 79% in adults and 44% in aged PPMO-treated mdx mice, whereas dystrophin protein restoration was 35% and 8%, respectively. Histopathological improvements were only evident in the adult-treated mice. PPMO-treatment in adult mdx mice induced a broad transcriptomic shift towards a wild-type signature, whereas treatment in aged mice resulted in negligible gene expression changes, indicating that late intervention is ineffective at reversing disease-associated pathologies despite low-level dystrophin restoration. Dystrophin transcript imbalance was corrected only in adult-treated mdx mice. Increased expression of the dystrophin-repressing microRNA miR-31-5p, which was more strongly upregulated in aged mdx muscle, provides a potential mechanistic explanation. In conclusion, PPMO-mediated exon skipping is substantially more effective when initiated in adult rather than aged dystrophic muscle, supporting early therapeutic intervention in DMD-affected individuals.\nCompeting Interest Statement\nMJAW is a founder, advisor, and shareholder in Pepgen Ltd, a company which aims to commercialize PPMO technologies similar to those utilized in this manuscript. The remaining authors declare no competing interests.","source_license":"CC-BY-4.0","license_restricted":false}