Talk abstracts
Talk on Wednesday 04:45-05:00pm submitted by Tabatha Simmons
Exon 2 skipping strategies to treat DMD 5’ mutations: Intramuscular delivery of rAAV9.snRNA mediated skipping and antisense morpholino oligomers.
Tabatha Simmons (The Center for Gene Therapy, Nationwide Childrens Hospital, Molecular Cellular, and Developmental Biology, The Ohio State University), Nicolas Wein (The Center for Gene Therapy, Nationwide Childrens Hospital), Adeline Vulin-Chaffiol (The Center for Gene Therapy, Nationwide Childrens Hospital), Kristin Heller (The Center for Gene Therapy, Nationwide Childrens Hospital), Louise Rodino-Klapac (The Center for Gene Therapy, Nationwide Childrens Hospital), Kevin Flanigan (The Center for Gene Therapy, Nationwide Childrens Hospital, Departments of Pediatrics and Neurology, The Ohio State University)
Abstract:
Exon skipping therapies for Duchenne muscular dystrophy (DMD) have to date been directed toward patients with out-of-frame exon deletions, resulting in larger but in-frame internal deletions and translation of an internally truncated but partially functional dystrophin protein. We are developing exon-skipping therapies for duplication mutations, which account for around 6% of all mutations by testing both virally (AAV) mediated skipping induced by a modified U7snRNA (rAAV9.U7.ACCA) and antisense oligomer-induced skipping in the Dup2 mouse. Intramuscular (IM) injections of rAAV9.U7.ACCA at 6 doses between 1x1010 and 1x1012 total vector genomes were delivered to the tibialias anterior (TA) (N=6) and analyzed for exon skipping and dystrophin production. Additionally, peptide-morpholino conjugates (PPMO) containing an antisense to exon 2 was delivered by IM injection to the TA at doses of either 10 or 20 ug total PPMO (N=3 each) and analyzed for exon skipping and dystrophin production. Treatment with either modality results in significant expression of apparently full-length dystrophin. Physiology has been assessed in the AAV-treated mice, in which correction of absolute force deficits in comparison to the background Bl6 strain are seen, along with a partial yet significant correction of eccentric contraction injury in comparison to untreated Dup2 mice. These data suggest that skipping of a duplicated exon 2 may be a feasible therapeutic approach, particularly because skipping of exon 2 may be associated with an apparently unlimited therapeutic window. Complete skipping of exon 2 results in activation of an internal ribosome entry site (IRES) located in exon 5 of dystrophin allowing for cap-independent translation from an alternative initiation site within exon 6. This alternate dystrophin isoform is highly functional despite being N-truncated, suggesting a potential route to therapy for any of the approximately 5% of patients with mutations in the 5’ end of the gene.
Keywords: DMD, exon-skipping, gene therapy