CRISPR/Cas13-mediated Dynamin 2 reduction therapy in a canine model of DNM2-related centronuclear myopathy

Abstract

We determined the potential of CRISPR/Cas13 technology as a therapeutic approach for centronuclear myopathies (CNMs) by reducing the expression of a single protein, DNM2. CNMs are severe congenital rare muscle disorders that result in muscle hypotrophy and weakness, with no cure. CNMs frequently result from mutations in either BIN1, MTM1, or DNM2 genes, with DNM2 being a key GTPase that plays a pivotal role in muscle membrane interactions with MTM1 and BIN1. Previous studies indicate that reducing DNM2 transcript expression by half could correct CNM phenotypes regardless the genetic forms, paving the way for a broad-spectrum CNM-therapy. We evaluated CRISPR/Cas13X.1-mediated DNM2 transcript knockdown, as a therapeutic application in a unique naturally-occurring canine CNM model harboring the DNM2R465W/+ mutation, the most frequent pathogenic variant in patients. We show that in vivo intramuscular AAV-mediated CRISPR/Cas13X.1 injections, led to a reduction in DNM2 transcript and protein levels at one and two months post-treatment. Our results demonstrate the feasibility of CRISPR/Cas13-based therapy for CNM in a large animal model, paving the way for advancing this approach towards clinical trials.