An antisense method for efficient exon skipping and its application to Duchenne muscular dystrophy.

Abstract

Antisense-mediated exon skipping has been used to treat Duchenne muscular dystrophy (DMD) by restoring the <i>DMD</i> reading frame to express partially functional dystrophin. However, difficulty in designing effective antisense oligonucleotides (ASOs) has hindered the development of clinically effective therapies. Here we describe a robust bipartite ASO design, termed 5' splice site decoy (5D)-ASO, in which a short tail sequence is employed to enhance the splicing repression exerted by an antisense moiety. The tail carried by the antisense moiety to an exon of interest in a target pre-mRNA interferes with the recognition of the exon's 5' splice site by U1 snRNA and markedly enhances exon skipping, compared to tail-less ASOs. 5D-ASO has broad applicability, based on multiple genes tested. Particularly, an 8-nt tail, when appended to sequences targeting <i>DMD</i> exon 51, elicited a pronounced increase in exon skipping in mouse models, restored dystrophin expression in muscle tissues and improved the phenotype, without obvious signs of toxicity. The lead ASO further demonstrated a marked exon-skipping effect and an overall safe profile in monkeys. Our data establish a valuable platform technology for RNA-targeted therapeutics.