Systemic PPMO-mediated dystrophin expression in the Dup2 mouse model of Duchenne muscular dystrophy

Liubov V Gushchina; Tatyana A Vetter; Emma C Frair; Adrienne J Bradley; Kelly M Grounds; Jacob W Lay; Nianyuan Huang; Aisha Suhaiba; Frederick J Schnell; Gunnar Hanson; Tabatha R Simmons; Nicolas Wein; Kevin M Flanigan

doi:10.1016/j.omtn.2022.10.025

Systemic PPMO-mediated dystrophin expression in the Dup2 mouse model of Duchenne muscular dystrophy

Mol Ther Nucleic Acids. 2022 Nov 9:30:479-492. doi: 10.1016/j.omtn.2022.10.025. eCollection 2022 Dec 13.

Authors

Affiliations

¹ The Center for Gene Therapy, Nationwide Children's Hospital, Columbus, OH, USA.
² Sarepta Therapeutics, Inc., Cambridge, MA, USA.
³ Department of Pediatrics, The Ohio State University, Columbus, OH, USA.
⁴ Department of Neurology, The Ohio State University, Columbus, OH, USA.

Abstract

Duchenne muscular dystrophy (DMD) is a devastating muscle-wasting disease that arises due to the loss of dystrophin expression, leading to progressive loss of motor and cardiorespiratory function. Four exon-skipping approaches using antisense phosphorodiamidate morpholino oligomers (PMOs) have been approved by the FDA to restore a DMD open reading frame, resulting in expression of a functional but internally deleted dystrophin protein, but in patients with single-exon duplications, exon skipping has the potential to restore full-length dystrophin expression. Cell-penetrating peptide-conjugated PMOs (PPMOs) have demonstrated enhanced cellular uptake and more efficient dystrophin restoration than unconjugated PMOs. In the present study, we demonstrate widespread PPMO-mediated dystrophin restoration in the Dup2 mouse model of exon 2 duplication, representing the most common single-exon duplication among patients with DMD. In this proof-of-concept study, a single intravenous injection of PPMO targeting the exon 2 splice acceptor site induced 45% to 68% exon 2-skipped Dmd transcripts in Dup2 skeletal muscles 15 days post-injection. Muscle dystrophin restoration peaked at 77% to 87% average dystrophin-positive fibers and 41% to 51% of normal signal intensity by immunofluorescence, and 15.7% to 56.8% of normal by western blotting 15 to 30 days after treatment. These findings indicate that PPMO-mediated exon skipping is a promising therapeutic strategy for muscle dystrophin restoration in the context of exon 2 duplications.

Keywords: Duchenne muscular dystrophy, DMD; Dup2 mice; MT: oligonucleotides: therapies and applications; antisense oligonucleotide, AO; cell-penetrating peptide-conjugated PMO, PPMO; dystrophin; exon duplications; phosphorodiamidate morpholino oligomers, PMO.