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In vivo restoration of dystrophin expression

Fletcher, S., Honeyman, K., Fall, A., Harding, P., Johnsen, R. and Wilton, S. (2005) In vivo restoration of dystrophin expression. In: 4th Australasian Gene Therapy Society Meeting, 27 - 29 April 2005, Rydges Hotel, Carlton, VIC.


Duchenne muscular dystrophy (DMD) is the most common, serious form of muscular dystrophy and is caused by mutations in the large dystrophin gene. We are evaluating the potential of antisense oligonucleotides (AOs) to bypass defects in the dystrophin gene that preclude the synthesis of a functional protein. In-frame dystrophin gene rearrangements typically result in the milder allelic disorder, Becker muscular dystrophy, clearly demonstrating that some regions of the dystrophin protein can be lost with relatively minor consequences. The huge size of the dystrophin gene, its complex expression patterns and expression of the predominant isoforms in non-dividing cells, have hindered gene replacement therapies for DMD, but render the dystrophin gene product an ideal candidate to evaluate AO therapies directed at splicing intervention. We report in vitro and in vivo studies, comparing morpholino and 2_-O-methyl AOs of identical sequence and peptide nucleic acids (PNAs) directed at the same dystrophin splice site, in a mouse model of muscular dystrophy. PNAs were inefficient at inducing specific exon skipping in vitro and in vivo under all delivery conditions tested. The 2OMeAO was able to restore dystrophin expression in vitro and in vivo when complexed with delivery agents. The ability of the uncomplexed morpholino to induce exon skipping was unexpectedly high in vivo, whereas this same preparation appeared essentially ineffective in vitro. Intramuscular administration of the morpholino to juvenile mice reduced central nucleation and resulted in near-normal dystrophin expression and muscle architecture. The efficient uptake of the uncomplexed morpholino in vivo may be considered an advantage when applying these compounds in a clinical setting. While testing in cell culture is a critical step in the design of AOs for induced exon skipping, delivery and the ultimate efficacy of AOs can only be evaluated in a relevant and appropriate animal model.

Publication Type: Conference Item
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