Antisense-induced exon skipping and synthesis of dystrophin in the mdx mouse

Duchenne muscular dystrophy (DMD) is a severe muscle wasting disease arisin g from defects in the dystrophin gene, typically nonsense or frameshift mut ations, that preclude the synthesis of a functional protein. A milder, alle lic version of the disease, Becker muscular dystrophy, generally arises fro m in-frame deletions that allow synthesis of a shorter but still semifuncti onal protein. Therapies to introduce functional dystrophin into dystrophic tissue through either cell or gene replacement have not been successful to date. We report an alternative approach where 2'-O-methyl antisense oligori bonucleotides have been used to modify processing of the dystrophin pre-mRN A in the mdx mouse model of DMD. By targeting 2'-O-methyl antisense oligori bonucleotides to block motifs involved in normal dystrophin pre-mRNA splici ng, we induced excision of exon 23, and the mdx nonsense mutation, without disrupting the reading frame. Exon 23 skipping was first optimized in vitro in transfected H-2K(b)-tsA58 mdx myoblasts and then induced in vivo. Immun ohistochemical staining demonstrated the synthesis and correct subsarcolemm al localization of dystrophin and gamma -sarcoglycan in the mdx mouse after intramuscular delivery of antisense oligoribonucleotide:liposome complexes . This approach should reduce the severity of DMD by allowing a dystrophic gene transcript to be modified, such that it can be translated into a Becke r-dystrophin-like protein.