Graham McClorey | PubMed Health

PubMed2005

Splicing intervention for Duchenne muscular dystrophy.

The manipulation of pre-mRNA to alter gene transcript splicing patterns offers considerable potential for many genetic disorders. In particular, the targeted removal of one or more exons from a gene transcript can skip over, or compensate for, disease-causing mutations. Duchenne muscular dystrophy (DMD), the most common and severe form of muscular dystrophy, is one such disorder that could benefit from this strategy. Splicing modulation can convert a DMD phenotype into the less severe allelic Becker-like phenotype. Recent studies using antisense oligonucleotide-targeted exon skipping to induce near normal dystrophin in vivo in animal models, and in vitro in DMD cell lines, highlight the promise of this approach. On the basis of these successes, human clinical trials could be realized in the near future.

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PubMedDate not supplied

Improved antisense oligonucleotide induced exon skipping in the mdx mouse model of muscular dystrophy.

BACKGROUND: Duchenne muscular dystrophy (DMD) is a fatal genetic disorder caused by dystrophin gene mutations that preclude synthesis of a functional protein. One potential treatment of the disorder has utilised antisense oligoribonucleotides (AOs) to induce removal of disease-associated exons during pre-mRNA processing. Induced in-frame mRNA transcripts encode a shorter but functional dystrophin. We have investigated and improved the design of AOs capable of removing exon 23, and thus the disease-causing nonsense mutation, from mRNA in the mdx mouse model of DMD. METHODS: H-2K(b)-tsA58 mdx cultures were transfected with complexes of Lipofectin and AOs. Exon skipping was detected by RT-PCR and subsequent protein production was demonstrated by Western blotting. AOs were delivered at a range of doses in order to compare relative efficiencies. RESULTS: We describe effective and reproducible exon 23 skipping with several AOs, including one as small as 17 nucleotides. Furthermore, the location of a sensitive exon 23 target site has been refined, whilst minimum effective doses have been estimated in vitro. These doses are significantly lower than previously reported and were associated with the synthesis of dystrophin protein in vitro. CONCLUSIONS: These results demonstrate the increasing feasibility of an AO-based therapy for treatment of DMD. By refining AO design we have been able to reduce the size and the effective dose of the AOs and have dramatically improved the efficiency of the technique.

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