Assessment of the feasibility of exon 45–55 multiexon skipping for duchenne muscular dystrophy

Laura Van Vliet,Judith Ct Van Deutekom,Christa L De Winter,Gert-Jan B Van Ommen,Annemieke Aartsma-Rus

doi:10.1186/1471-2350-9-105

Laura Van Vliet, Judith Ct Van Deutekom + Show 3 more

Open Access

https://doi.org/10.1186/1471-2350-9-105

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Abstract

BackgroundThe specific skipping of an exon, induced by antisense oligonucleotides (AON) during splicing, has shown to be a promising therapeutic approach for Duchenne muscular dystrophy (DMD) patients. As different mutations require skipping of different exons, this approach is mutation dependent. The skipping of an entire stretch of exons (e.g. exons 45 to 55) has recently been suggested as an approach applicable to larger groups of patients. However, this multiexon skipping approach is technically challenging. The levels of intended multiexon skips are typically low and highly variable, and may be dependent on the order of intron removal. We hypothesized that the splicing order might favor the induction of multiexon 45–55 skipping.MethodsWe here tested the feasibility of inducing multiexon 45–55 in control and patient muscle cell cultures using various AON cocktails.ResultsIn all experiments, the exon 45–55 skip frequencies were minimal and comparable to those observed in untreated cells.ConclusionWe conclude that current state of the art does not sufficiently support clinical development of multiexon skipping for DMD.

Highlights

The specific skipping of an exon, induced by antisense oligonucleotides (AON) during splicing, has shown to be a promising therapeutic approach for Duchenne muscular dystrophy (DMD) patients
We hypothesized that multiexon 45–55 skipping might be induced or enhanced using different AONs and combination strategies (Figure 1)
To enhance the chance that both AONs hybridize to the same pre-mRNA transcript we included an AON where AONs targeting exon 45 and exon 55 are linked by 10 uracil nucleotides

Summary

Introduction

The specific skipping of an exon, induced by antisense oligonucleotides (AON) during splicing, has shown to be a promising therapeutic approach for Duchenne muscular dystrophy (DMD) patients. Antisense-mediated exon skipping is emerging as a very promising therapeutic approach for Duchenne muscular dystrophy (DMD) [1] The aim of this approach is to restore the disrupted reading frame of DMD transcripts, and allow synthesis of partly functional, internally deleted Becker-like dystrophins, rather than prematurely truncated non-functional Duchenne dystrophins. This can be achieved by antisense oligonucleotides (AONs) that target specific exons and hide them from the splicing machinery during pre-mRNA splicing, resulting in the skipping of said exons [2]. Most mutations involve deletions (page number not for citation purposes)

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Conclusion