Dual exon skipping in myostatin and dystrophin for Duchenne muscular dystrophy

We targeted myostatin exon 2 using antisense oligonucleotides (AON) in healthy and DMD-derived myotubes cultures. We assessed the exon skipping level, transcriptional expression of myostatin and its target genes, and combined myostatin and several dystrophin AONs. These AONs were also applied in the mdx mice models via intramuscular injections.Myostatin AON induced exon 2 skipping in cell cultures and to a lower extent in the mdx mice. It was accompanied by decrease in myostatin mRNA and enhanced MYOG and MYF5 expression. Furthermore, combination of myostatin and dystrophin AONs induced simultaneous skipping of both genes.We conclude that two AONs can be used to target two different genes, MSTN and DMD, in a straightforward manner. Targeting multiple ligands of TGF-beta family will be more promising as adjuvant therapies for DMD.Duchenne muscular dystrophy (DMD) is an X-linked recessive neuromuscular disorder, which is caused by dystrophin deficiency in muscle fibers. DMD fibers are more sensitive to muscle damage, leading to degeneration and replacement of muscle fibers by fat and connective tissue (fibrosis). Monaco et al found that frame shift mutations in the DMD gene will lead to a truncated and non-functional form of dystrophin [1], which become the primary cause of the disease. However, mutations which maintain the DMD open reading frame result in shorter dystrophin proteins that retain the essential actin binding-, cysteine rich- and carboxy terminal domains, and thus are partly functional [1]. Patients with such mutations develop the less severe Becker muscular dystrophy. This reading frame rule holds true for ~91% of DMD cases [2] and has inspired the development of the exon skipping strategy, which employs antisense oligonucleotides (AONs). These small synthetic RNA molecules are complimentary to exonic or splice site sequences, thereby upon hybridization are able to modulate exon inclusion by the splicing machinery (recently reviewed in [3-5]).Comprehens