Combined 20-Hydroxyecdysone and Antisense-Mediated Exon Skipping Improve Functional Outcomes in a Mouse Model of Duchenne Muscular Dystrophy.

Abstract

Duchenne muscular dystrophy (DMD) is a severe X-linked disorder caused by mutations in the DMD gene, resulting in a lack of dystrophin protein. This leads to progressive muscle wasting, cardiac and respiratory dysfunction, and premature death. Antisense oligonucleotide (ASO)-based therapies represent a promising approach to treating DMD, with several already approved by the FDA. However, the levels of dystrophin restoration achieved in clinical trials are often insufficient for meaningful therapeutic impact, highlighting the urgent need to enhance ASO efficacy. One potential strategy is to improve muscle pathophysiology, which is compromised in DMD due to cycles of necrosis and regeneration, chronic inflammation, and fibrotic and adipose tissue replacement. These disease characteristics may limit ASO efficiency. In this study, we evaluated the combination of tricyclo-DNA-ASO targeting the Dmd exon 23 with 20-hydroxyecdysone (20-E), a steroid hormone known to activate the protective arm of the renin-angiotensin-aldosterone system, enhance protein and ATP synthesis, and exhibit anti-inflammatory and antifibrotic properties. Mdx mice were treated with ASO alone or in combination with 20-E for 8 weeks. While both treatments restored similar levels of dystrophin and significantly improved functional outcomes such as the distance run and maximum speed in the treadmill exhaustion test, other improvements like the specific force and the decrease in the force drop after eccentric contraction were observed only with the combination therapy. Importantly, the cotreatment was well tolerated without liver or kidney toxicity. These findings provide proof of concept that combining 20-E with ASO therapy can ameliorate dystrophic pathology and improve muscle function in a DMD mouse model. By targeting both dystrophin restoration and muscle pathophysiology, this combined approach may offer a therapeutic strategy with the potential for meaningful clinical benefits, warranting further investigation and potential translation to patients.