Duchenne muscular dystrophy is a fatal disorder caused by mutations in the DMD gene on the X chromosome, the largest known gene in humans. The mutations lead to the absence of the protein dystrophin, which is important for maintaining the cell membrane in muscle fiber. Duchenne is the most common inherited neuromuscular disorder, affecting almost 1 in 5000 male births. The ensuing progressive muscle loss in these boys leads to an inability to walk by their teens and heart failure in their twenties, resulting in death.

About 70% of Duchenne cases are caused by deletions in the DMD gene that create a misalignment in the reading frame when the gene is transcribed. This results in the wrong sequence of amino acids being produced instead of functioning dystrophin.

Selective treatments

Four therapies have been approved by the U.S. Food and Drug Administration for treating such cases. These work by using synthetic nucleic acid analogs called antisense oligonucleotides (AO) that force the cell’s transcription machinery to skip over the sections, or exons, where the mutations are found. Although this means that the resulting dystrophin protein is missing a portion, the rest is intact and may be able to carry out its role to some extent.

The current exon-skipping therapies are specific to single exons, meaning that they can only each treat up to 8 to 13 percent of all patients. Toshifumi Yokota, PhD, professor of medical genetics at the University of Alberta, and colleagues previously developed a mechanism to skip over a mutation hotspot that consists of 11 exons in a row. This broader approach could theoretically treat more than 40 percent of patients. However, this requires adding 11 different AOs to skip over all of the exons, which could limit the efficacy of the treatment and increase the risk of unintended effects.

Now, in a new publication, the researchers have reduced the number of AOs required and improved their uptake in mice. Some AOs can also cause skipping of the exons that follow the one that they are designed to target. By optimizing the mixture used, the team was able to skip over exons 44 to 55 in DMD using only five AOs. This reduced cocktail had a similar efficacy to the original when tested in cells derived from patients.

Improving delivery to muscle

Another limitation of AO therapies is that they are rapidly cleared from the bloodstream and have poor uptake into muscle. To counter this, the researchers added a cell-penetrating peptide to help improve the absorption of the AOs. The peptide, called DG9, is based on a domain of a human transcription factor protein and has been used in other research to improve delivery in a variety of tissues, including the heart.

Mice given repeated treatments of the DG9 and AO combination produced about three percent of healthy dystrophin levels. Although this is still a significantly reduced amount, there were noted improvements seen in the muscle tissue. This suggests that even minor increases in the production of dystrophin could make a difference to the quality of life and survival for people with Duchenne.

The authors of the paper wrote that their future work on repeated therapy would focus on its effects on cardiac function, given its importance for life expectancy.

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