Abstract
Splice-switching antisense oligonucleotides (SSOs) offer great potential for RNA-targeting therapies, and two SSO drugs have been recently approved for treating Duchenne Muscular Dystrophy (DMD) and Spinal Muscular Atrophy (SMA). Despite promising results, new developments are still needed for more efficient chemistries and delivery systems. Locked nucleic acid (LNA) is a chemically modified nucleic acid that presents several attractive properties, such as high melting temperature when bound to RNA, potent biological activity, high stability and low toxicity in vivo. Here, we designed a series of LNA-based SSOs complementary to two sequences of the human dystrophin exon 51 that are most evolutionary conserved and evaluated their ability to induce exon skipping upon transfection into myoblasts derived from a DMD patient. We show that 16-mers with 60% of LNA modification efficiently induce exon skipping and restore synthesis of a truncated dystrophin isoform that localizes to the plasma membrane of patient-derived myotubes differentiated in culture. In sum, this study underscores the value of short LNA-modified SSOs for therapeutic applications.
Highlights
Antisense oligonucleotides are powerful tools to modulate gene expression
One region contains a sequence of 15 nucleotides that is conserved across mammalian species ranging from primates (Human, Orangutan, Macaque, Tarsier, Bushbaby), to Tree-Shrew, Megabat, Hedgehog, Alpaca, Dolphin, Ferret, and rodents (Mouse and Rat), as well as bird species such as Chicken, Duck, Flycatcher, and Zebra-finch
Locked nucleic acid (LNA)-modified SSOs targeting either the acceptor or donor splice sites of the Duchenne Muscular Dystrophy (DMD) exon 58 were shown to be effective in inducing exon skipping
Summary
Antisense oligonucleotides are powerful tools to modulate gene expression. Antisense oligonucleotides can be used to induce RNA interference or RNase H based mechanisms of gene down-regulation, inhibit the function of microRNAs, or modulate splicing (for a recent review see [1]). Despite early clinical trial failures, new developments are fueling optimism in the antisense field. The design of novel chemical modifications and delivery systems is improving the potency and efficacy of these drugs in RNA-targeting therapeutic. Short LNA SSOs restore dystrophin production in DMD myotubes ulisboa.pt/investigacao/gapic/. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript
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