Currently approved exon skipping phosphorodiamidate morpholino oligomer (PMO) therapies for Duchenne muscular dystrophy (DMD) produce modest amounts of dystrophin due to poor cell entry and limited escape from the endosome. In order to overcome these limitations, we developed a family of therapeutics consisting of Endosomal Escape Vehicle (EEV™) cyclic cell penetrating peptides conjugated to exon skipping PMOs.
ENTR-601-45 is an exon 45 skipping PMO conjugated to the EEV platform and is in development for the treatment of exon 45 skip-amenable DMD. Here, we examined the efficacy of ENTR-601-45 in cell and animal models. Treatment of exon 45 skip-amenable, DMD patient-derived skeletal and cardiac muscle cells with ENTR-601-45 demonstrated robust, dose-dependent exon 45 skipping and dystrophin production. To assess the therapeutic potential of ENTR-601-45 in vivo, we utilized del44hDMD.mdx mice containing a human dystrophin-expressing transgene (hDMD) with DMD exon 44 deleted on an mdx background. These mice are amenable to human DMD exon 45 skipping, allowing for quantification of both exon 45 skipping and dystrophin production in vivo. Three doses of ENTR-601-45 administered every six weeks (Q6W) to del44hDMD.mdx mice produced robust, dose-dependent exon 45 skipping, dystrophin expression, and proper localization to the sarcolemma in skeletal muscle 6 weeks after the third dose. In addition, three Q6W doses of ENTR-601-45 produced a dose-dependent reduction to damage after repeated eccentric contractions in the gastrocnemius skeletal muscle which was maintained in all dose groups 5 weeks after the third dose.
These findings demonstrate the efficacy of ENTR-601-45 in both cell and animal models of exon 45 skip-amenable DMD. Improved skeletal muscle function in an exon 45 skip–amenable DMD mouse model suggests that ENTR-601-45 can produce functional dystrophin protein in vivo. Together, these results show the therapeutic potential of ENTR-601-45 and support further study in DMD patients amenable to exon 45 skipping.