Functional amounts of dystrophin produced by skipping the mutated exon in the mdx dystrophic mouse

Abstract

As a target for gene therapy, Duchenne muscular dystrophy (DMD) presents many obstacles but also an unparalleled prospect for correction by alternative splicing. The majority of mutations in the dystrophin gene occur in the region encoding the spectrin-like central rod domain, which is largely dispensable. Thus, splicing around mutations can generate a shortened but in-frame transcript, permitting translation of a partially functional dystrophin protein. We have tested this idea in vivo in the mdx dystrophic mouse (carrying a mutation in exon 23 of the dystrophin gene) by combining a potent transfection protocol with a 2-O-methylated phosphorothioated antisense oligoribonucleotide (2OMeAO) designed to promote skipping of the mutated exon*. The treated mice show persistent production of dystrophin at normal levels in large numbers of muscle fibers and show functional improvement of the treated muscle. Repeated administration enhances dystrophin expression without eliciting immune responses. Our data establishes the realistic practicality of an approach that is applicable, in principle, to a majority of cases of severe dystrophinopathy.

Dose-dependent restoration of dystrophin expression in cardiac muscle of dystrophic mice by systemically delivered morpholino

“…Specifically designed 2 0 O methyl phosphorothioate AONs delivered by intramuscular injections were able to skip the exon 23 effectively in tibialis anterior (TA) muscles and produce functional amount of proteins. 8 The same 2 0 O methyl phosphorothioate AON was shown to induce dystrophin expression in body-wide muscles when delivered systemically. 9 More recently, regular systemic administration of the phosphorodiamidate morpholino oligomers targeting the same mouse dystrophin exon 23 induced effective exon skipping and produced functional levels of dystrophin in skeletal muscles.…”

“…This can lead to the production of truncated but functional dystrophin, and can reverse a DMD to a milder Becker muscular dystrophy or near normal phenotypes. [8][9][10][11] The therapeutic potential of antisense therapy for DMD was initially showed in the dystrophic mdx mouse that harbors a nonsense point mutation in the exon 23, leading to general absence of dystrophin in the muscles. Specifically designed 2 0 O methyl phosphorothioate AONs delivered by intramuscular injections were able to skip the exon 23 effectively in tibialis anterior (TA) muscles and produce functional amount of proteins.…”

Long-term expression of full-length human dystrophin in transgenic mdx mice expressing internally deleted human dystrophins

“…6 There are many strategies for the induction of de novo dystrophin expression as a therapeutic approach for the treatment of DMD. These include systemic drug delivery, 7 gene modification at DNA and RNA levels, [8][9][10][11] cell transplantation [12][13][14] and gene delivery with a wide range of both viral [15][16][17] and nonviral [18][19][20] delivery systems. However, therapeutic levels, together with a stable longterm presence of dystrophin at the sarcolemma of the muscle fibres, are essential.…”

Le saut d’exon thérapeutique : un espoir pour les dystrophinopathies

“…Goyenvalle [5,6]. Mais ni l'efficacité, ni la stabilité n'atteignaient un niveau de significativité clinique.…”

“…Dès 1991, M. Matsuo [3] avait insisté sur l'importance des phénomènes d'épis-sage dans le déterminisme et la théra-peutique des dystrophinopathies. Plus récemment, d'autres groupes se sont engagés dans la voie de l'épissothéra-pie, comme alternative à la thérapie génique classique qui n'a pas encore remporté les succès escomptés [4][5][6]. Une restauration de la synthèse de dystrophine musculaire chez la souris mdx par exclusion post-transcriptionnelle de l'exon porteur de la mutation (stop sur l'exon 23) a été rapportée par plusieurs…”