Publications (9) View all
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Article: A Novel Morpholino Oligomer Targeting ISS-N1 Improves Rescue of Severe SMA Transgenic Mice.
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ABSTRACT: In the search of the most efficacious antisense oligonucleotides (AO) aimed at inducing SMN2 exon 7 inclusion, we systematically assessed three AOs, PMO25 (-10, -34), PMO18 (-10, -27) and PMO20 (-10, -29), complementary to SMN2 intron 7 splicing silencer (ISS-N1). PMO25 was the most efficacious in augmenting exon7 inclusion in vitro in spinal muscular atrophy (SMA) patients' fibroblasts and in vitro splicing assays. PMO25 and PMO18 were compared further using a severe SMA mouse model. After a single intra cerebral ventricular (ICV) injection in neonatal mice, PMO25 increased the lifespan of severe SMA mice up to thirty-fold, with an average survival greater by three-fold than PMO18 at a dose of 20 µg/g and 2-fold at 40 µg/g. Exon7 inclusion was increased in the central nervous system (CNS) but not in peripheral tissues. Systemic delivery of PMO25 at birth achieved a similar outcome and produced increased exon7 inclusion both in the CNS and peripherally. Systemic administration of 10 µg/g of PMO25 conjugated to an octa-guanidine dendrimer (VMO25) increased the lifespan only two-fold in neonatal type I SMA mice, although prevented tail necrosis in mild SMA mice. Higher doses and ICV injection of VMO25 were associated with toxicity. We conclude that (1) the 25-mer AO is more efficient than the 18-mer and 20-mer in modifying SMN2 splicing in vitro; (2) it is more efficient in prolonging survival in SMA mice and (3) naked Morpholino oligomers are more efficient and safer than vivo-Morpholinos and have potential for future SMA clinical applications.Human gene therapy 01/2013; · 4.20 Impact Factor -
Article: Improved Antisense Oligonucleotide Design to Suppress Aberrant SMN2 Gene Transcript Processing: Towards a Treatment for Spinal Muscular Atrophy.
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ABSTRACT: Spinal muscular atrophy (SMA) is caused by loss of the Survival Motor Neuron 1 (SMN1) gene, resulting in reduced SMN protein. Humans possess the additional SMN2 gene (or genes) that does produce low level of full length SMN, but cannot adequately compensate for loss of SMN1 due to aberrant splicing. The majority of SMN2 gene transcripts lack exon 7 and the resultant SMNΔ7 mRNA is translated into an unstable and non-functional protein. Splice intervention therapies to promote exon 7 retention and increase amounts of full-length SMN2 transcript offer great potential as a treatment for SMA patients. Several splice silencing motifs in SMN2 have been identified as potential targets for antisense oligonucleotide mediated splice modification. A strong splice silencer is located downstream of exon 7 in SMN2 intron 7. Antisense oligonucleotides targeting this motif promoted SMN2 exon 7 retention in the mature SMN2 transcripts, with increased SMN expression detected in SMA fibroblasts. We report here systematic optimisation of phosphorodiamidate morpholino oligonucleotides (PMO) that promote exon 7 retention to levels that rescued the phenotype in a severe mouse model of SMA after intracerebroventricular delivery. Furthermore, the PMO gives the longest survival reported to date after a single dosing by ICV.PLoS ONE 01/2013; 8(4):e62114. · 4.09 Impact Factor -
Article: Analysis of HLA-DRB3 alleles and supertypical genotypes in the MHC Class II region in sporadic inclusion body myositis.
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ABSTRACT: We compared the carriage frequencies of HLA-DRB3 and its major alleles and of HLA-DRB4 and HLA-DRB5 in an Australian sIBM cohort and a population control group who had previously been genotyped for the HLA-DRB1*03:01 risk allele. There was a strong disease association with the carriage of the DRB3*01:01 allele which was accounted for by its linkage disequilibrium with DRB1*03:01. The carriage of HLA-DRB4 was found to be strongly protective and abrogated the risk effect of HLA-DRB1*03:01. The findings indicate that haplotypic combinations of alleles at the HLA-DRB1 and secondary HLA-DRB loci have important risk modifying effects in sIBM.Journal of neuroimmunology 09/2012; · 2.84 Impact Factor -
Article: A single administration of morpholino antisense oligomer rescues spinal muscular atrophy in mouse.
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ABSTRACT: Spinal muscular atrophy (SMA) is an autosomal-recessive disorder characterized by α-motor neuron loss in the spinal cord anterior horn. SMA results from deletion or mutation of the Survival Motor Neuron 1 gene (SMN1) and retention of SMN2. A single nucleotide difference between SMN1 and SMN2 results in exclusion of exon 7 from the majority of SMN2 transcripts, leading to decreased SMN protein levels and development of SMA. A series of splice enhancers and silencers regulate incorporation of SMN2 exon 7; these splice motifs can be blocked with antisense oligomers (ASOs) to alter SMN2 transcript splicing. We have evaluated a morpholino (MO) oligomer against ISS-N1 [HSMN2Ex7D(-10,-29)], and delivered this MO to postnatal day 0 (P0) SMA pups (Smn-/-, SMN2+/+, SMNΔ7+/+) by intracerebroventricular (ICV) injection. Survival was increased markedly from 15 days to >100 days. Delayed CNS MO injection has moderate efficacy, and delayed peripheral injection has mild survival advantage, suggesting that early CNS ASO administration is essential for SMA therapy consideration. ICV treatment increased full-length SMN2 transcript as well as SMN protein in neural tissue, but only minimally in peripheral tissue. Interval analysis shows a decrease in alternative splice modification over time. We suggest that CNS increases of SMN will have a major impact on SMA, and an early increase of the SMN level results in correction of motor phenotypes. Finally, the early introduction by intrathecal delivery of MO oligomers is a potential treatment for SMA patients.Human Molecular Genetics 12/2011; 21(7):1625-38. · 7.64 Impact Factor -
Article: Rational design of antisense oligomers to induce dystrophin exon skipping.
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ABSTRACT: Duchenne muscular dystrophy (DMD), one of the most severe neuromuscular disorders of childhood, is caused by the absence of a functional dystrophin. Antisense oligomer (AO) induced exon skipping is being investigated to restore functional dystrophin expression in models of muscular dystrophy and DMD patients. One of the major challenges will be in the development of clinically relevant oligomers and exon skipping strategies to address many different mutations. Various models, including cell-free extracts, cells transfected with artificial constructs, or mice with a human transgene, have been proposed as tools to facilitate oligomer design. Despite strong sequence homology between the human and mouse dystrophin genes, directing an oligomer to the same motifs in both species does not always induce comparable exon skipping. We report substantially different levels of exon skipping induced in normal and dystrophic human myogenic cell lines and propose that animal models or artificial assay systems useful in initial studies may be of limited relevance in designing the most efficient compounds to induce targeted skipping of human dystrophin exons for therapeutic outcomes.Molecular Therapy 04/2009; 17(8):1418-26. · 6.87 Impact Factor
