For the majority of Duchenne muscular dystrophy (DMD) mutations, antisense oligonucleotide (AON)-mediated exon skipping has the potential to restore a functional protein. Here we show that weekly intravenous injections of morpholino phosphorodiamidate (morpholino) AONs induce expression of functional levels of dystrophin in body-wide skeletal muscles of the dystrophic mdx mouse, with resulting improvement in muscle function. Although the level of dystrophin expression achieved varies considerably between muscles, antisense therapy may provide a realistic hope for the treatment of a majority of individuals with DMD.

Antisense oligonucleotide-mediated alternative splicing has great potential for treatment of Duchenne muscular dystrophy (DMD) caused by mutations within nonessential regions of the dystrophin gene. We have recently shown in the dystrophic mdx mouse that exon 23, bearing a nonsense mutation, can be skipped after intramuscular injection of a specific 2 -O-methyl phosphorothioate antisense oligoribonucleotide (2OMeAO). This skipping created a shortened, but in-frame, transcript that is translated to produce near-normal levels of dystrophin expression. This expression, in turn, led to improved muscle function. However, because DMD affects muscles body-wide, effective treatment requires dystrophin induction ideally in all muscles. Here, we show that systemic delivery of specific 2OMeAOs, together with the triblock copolymer F127, induced dystrophin expression in all skeletal muscles but not in cardiac muscle of the mdx dystrophic mice. The highest dystrophin expression was detected in diaphragm, gastrocnemius, and intercostal muscles. Large numbers of fibers with near-normal level of dystrophin were observed in focal areas. Three injections of 2OMeAOs at weekly intervals enhanced the levels of dystrophin. Dystrophin mRNA lacking the targeted exon 23 remained detectable 2 weeks after injection. No evidence of tissue damage was detected after 2OMeAO and F127 treatment either by serum analysis or histological examination of liver, kidney, lung, and muscles. The simplicity and safety of the antisense protocol provide a realistic prospect for treatment of the majority of DMD mutations. We conclude that a significant therapeutic effect may be achieved by further optimization in dose and regime of administration of antisense oligonucleotide.exon skipping ͉ muscular dystrophy