Empirical and Computational Comparison of Alternative Therapeutic Exon Skip Repairs for Duchenne Muscular Dystrophy

Ma KM, Thomas ES, Wereszczynski J, Menhart N. Empirical and Computational Comparison of Alternative Therapeutic Exon Skip Repairs for Duchenne Muscular Dystrophy. Biochemistry 2019;58:2061-2076

Ma_2019_Biochemistry. Apr 16;58(15);2061-2076



Duchenne muscular dystrophy is a common and devastating genetic disease primarily caused by exon deletions that create a genetic frameshift in dystrophin. Exon skipping therapy seeks to correct this by masking an exon during the mRNA maturation process, restoring dystrophin expression, but creating an edited protein missing both the original defect and the therapeutically skipped region. Crucially, it is possible to correct many defects in alternative ways, by skipping an exon either before, or after the patient’s defect. This results in alternatively edited, hybrid proteins of possibly different properties and therapeutic consequences. We examined three such dystrophin exon skipped edits, Deltae45-53, Deltae46-54 and Deltae47-55, comprising two pairs of alternative repairs of Deltae46-53 and Deltae47-54 DMD defects. We found that in both cases, Deltae46-54 was the more stable repair as determined by a variety of thermodynamic and biochemical measurements. We also examined the origin of these differences with molecular dynamics simulations, which showed that these stability differences were the result of different types of structural perturbations. For example, in one edit there was partial unfolding at the edit site which caused domain-localized perturbations, while in another there was unfolding at the protein domain junctions distal to the edit site which increased molecular flexibility. These results demonstrate that alternative exon skip repairs of the same underlying defect can have very different consequences at the level of protein structure and stability, and furthermore that these can arise by different mechanisms, either locally, or by more subtle long-range perturbations.

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