Optimized lentiviral vector to restore full-length dystrophin via a cell-mediated approach in a mouse model of Duchenne muscular dystrophy

Jinhong Meng; Marc Moore; John Counsell; Francesco Muntoni; Linda Popplewell; Jennifer Morgan

doi:10.1016/j.omtm.2022.04.015

Optimized lentiviral vector to restore full-length dystrophin via a cell-mediated approach in a mouse model of Duchenne muscular dystrophy

Mol Ther Methods Clin Dev. 2022 May 2:25:491-507. doi: 10.1016/j.omtm.2022.04.015. eCollection 2022 Jun 9.

Authors

Jinhong Meng^{1

2}, Marc Moore^{1

3

2}, John Counsell^{1

4

2}, Francesco Muntoni^{1

2}, Linda Popplewell³, Jennifer Morgan^{1

2}

Affiliations

¹ Dubowitz Neuromuscular Centre, Molecular Neurosciences Section, Developmental Neuroscience Programme, UCL Great Ormond Street Institute of Child Health, 30 Guilford Street, London WC1N 1EH, UK.
² National Institute for Health Research, Great Ormond Street Institute of Child Health Biomedical Research Centre, University College London, London WC1N 1EH, UK.
³ Department of Biological Sciences, School of Life Sciences and the Environment, Royal Holloway University of London, Egham Hill, Egham TW20 0EX, UK.
⁴ UCL Division of Surgery and Interventional Science, Charles Bell House, 43-45 Foley Street, London W1W 7TY, UK.

Abstract

Duchenne muscular dystrophy (DMD) is a muscle wasting disorder caused by mutations in the DMD gene. Restoration of full-length dystrophin protein in skeletal muscle would have therapeutic benefit, but lentivirally mediated delivery of such a large gene in vivo has been hindered by lack of tissue specificity, limited transduction, and insufficient transgene expression. To address these problems, we developed a lentiviral vector, which contains a muscle-specific promoter and sequence-optimized full-length dystrophin, to constrain dystrophin expression to differentiated myotubes/myofibers and enhance the transgene expression. We further explored the efficiency of restoration of full-length dystrophin in vivo, by grafting DMD myoblasts that had been corrected by this optimized lentiviral vector intramuscularly into an immunodeficient DMD mouse model. We show that these lentivirally corrected DMD myoblasts effectively reconstituted full-length dystrophin expression in 93.58% ± 2.17% of the myotubes in vitro. Moreover, dystrophin was restored in 64.4% ± 2.87% of the donor-derived regenerated muscle fibers in vivo, which were able to recruit members of the dystrophin-glycoprotein complex at the sarcolemma. This study represents a significant advance over existing cell-mediated gene therapy strategies for DMD that aim to restore full-length dystrophin expression in skeletal muscle.

Keywords: Duchenne muscular dystrophy; lentivirus; muscle-specific promoter; myoblasts; native full-length dystrophin; sequence-optimized full-length dystrophin.