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T.P.2.03 Characterisation of a complex dystrophin mutation: Assume nothing when designing exon skipping strategies

Fletcher, S., Davis, M., MADDEN, H. and Wilton, S. (2008) T.P.2.03 Characterisation of a complex dystrophin mutation: Assume nothing when designing exon skipping strategies. Neuromuscular Disorders, 18 (9-10). p. 757.

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We report an unusual and complex dystrophin gene re-arrangement that causes Duchenne muscular dystrophy, and an antisense oligonucleotide (AO) exon skipping strategy that restores the reading frame. All dystrophin exons were present, as determined by MLPA and multiplex PCR studies, while RNA analysis indicated an anomaly involving exons 49 and 50. An inversion of 28 kb was identified that lead to the omission of these exons from the mature gene transcript. In addition, multiple mRNAs arising from the variable inclusion of at least six pseudo-exons were identified. The inversion arose from an unequal recombination event, and upon characterization of the precise breakpoints in introns 48 and 50, a small duplication was identified at the proximal intron 48 breakpoint and an 11 kb deletion was found at the distal intron 50 breakpoint. Several AOs were evaluated against the two most commonly incorporated pseudo-exons (PE1 and PE2) which, when used in conjunction with an AO targeting exon 51 for removal, should restore the reading frame in the majority of the patient’s dystrophin gene transcripts. AOs directed to PE1 were very efficient at excising both pseudo-exons, while surprisingly PE2 was not dislodged by any AOs designed to target this pseudo-exon. Furthermore, the AO designed to excise exon 51 removed the target exon and also PE1. Spatial arrangements of the pseudo-exons in this dystrophin pre-mRNA may play a role in exon definition and inclusion. The obscuring of authentic splicing motifs in exon 51 presumably compromised pseudo-exon recognition and led to removal of two exons with a single AO. This has implications for the skipping of multiple exons and also demonstrates that one of the most unusual dystrophin gene re-arrangements may be treatable with the first compound to enter DMD clinical trials in the United Kingdom.

Item Type: Journal Article
Publisher: Elsevier BV
Copyright: © 2008 Published by Elsevier B.V.
Notes: Abstract: 13th International Congress of the World Muscle Society, Newcastle Upon Tyne, UK. 29 September - 2 October 2008
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