Gene therapy for progressive muscular dystrophy using a new generation adenovirus vector and transposase

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Duchenne muscular dystrophy(DMD) is a fatal progressive muscle wasting disease caused by defects in the dystrophin gene. No viral vector except the helper-dependent adenovirus vector(HDAdv) can package 14kb full-length dystrophin cDNA and HDAdv is considerably safer than old-generation adenovirus vectors due to the large-size deletion in its genome. We have generated HDAdv that carries myc-tagged murine full-length dystrophin cDNA(HDAdv-myc-mFLdys). We injected it into the multiple proximal muscles of 7-day-old utrophin/dystrophin double knockout mice (dko mice), which typically show symptoms quite similar to human DMD because the proximal muscles are organs affected in DMD patients. Eight weeks after injections, the transduced dystrophin was widely expressed and we found a significant reduction of centrally nucleated myofibers and the restoration of dystrophin associated proteins, p-dystroglycan (p-DG) and a-sarcoglycan (a-SG), as well as neuronal nitric oxide synthase. (nNOS). The injected dko mice also showed an increase in body weight, an improvement in motor performances, and prolonged lifespan. Using HDAdv, we could treat DMD model mice, even when the therapeutic gene was transferred into multiple skeletal muscles. Our results suggest that multiple intramuscular administrations of HDAdv carrying full-length dystrophin may reduce symptoms and compensate for lost functions in DMD patients.For the long expression of target gene product, we tried to integrate the dystrophin gene into chromosome by using the sleeping beauty-transposone system and HDAdv. However, it revealed that the DNA construct of HDAdv is linear and it does not fit the circular DNA of the sleeping beauty-transposone system. So, further study is necessary to integrate the dystrophin gene into chromosome.