Reengineering a transmembrane protein to treat muscular dystrophy using exon skipping

Exon skipping uses antisense oligonucleotides as a treatment for genetic diseases. The antisense oligonucleotides used for exon skipping are designed to bypass premature stop codons in the target RNA and restore reading frame disruption. Exon skipping is currently being tested in humans with dystrophin gene mutations who have Duchenne muscular dystrophy. For Duchenne muscular dystrophy, the rationale for exon skipping derived from observations in patients with naturally occurring dystrophin gene mutations that generated internally deleted but partially functional dystrophin proteins. We have now expanded the potential for exon skipping by testing whether an internal, in-frame truncation of a transmembrane protein gamma-sarcoglycan is functional. We generated an internally truncated gamma-sarcoglycan protein that we have termed Mini-Gamma by deleting a large portion of the extracellular domain. Mini-Gamma provided functional and pathological benefits to correct the loss of gamma-sarcoglycan in a Drosophila model, in heterologous cell expression studies, and in transgenic mice lacking gamma-sarcoglycan. We generated a cellular model of human muscle disease and showed that multiple exon skipping could be induced in RNA that encodes a mutant human gamma-sarcoglycan. Since Mini-Gamma represents removal of 4 of the 7 coding exons in gamma-sarcoglycan, this approach provides a viable strategy to treat the majority of patients with gamma-sarcoglycan gene mutations.