EX-VIVO GENE-TRANSFER USING ADENOVIRUS-MEDIATED FULL-LENGTH DYSTROPHIN DELIVERY TO DYSTROPHIC MUSCLES

Duchenne muscular dystrophy (DMD) is an X-linked recessive muscle dise ase characterized by a lack of dystrophin expression. Myoblast transpl antation and gene therapy have the potential of restoring dystrophin, thus decreasing the muscle weakness associated with the disease. In th is study we present data on the myoblast mediated ex vivo gene transfe r of full-length dystrophin to mdx (dystrophin deficient) mouse muscle as a model for autologous myoblast transfer. Both isogenic primary md x myoblasts and an immortalized mdx cell line were transduced with an adenoviral vector that has all viral coding sequences deleted and enco des beta-galactosidase and full-length dystrophin. Subsequently, these transduced myoblasts were injected into dystrophic max muscle, where the injected cells restored dystrophin, as well as dystrophin-associat ed proteins. A greater amount of dystrophin replacement occurred in md x muscle following transplantation of mdx myoblasts isolated from a tr ansgenic mouse overexpressing dystrophin suggesting that engineering a utologous myoblasts to express high amounts of dystrophin might be ben eficial. The ex vivo approach possesses attributes that make it useful for gene transfer to skeletal muscle including: (1) creating a reserv oir of myoblasts capable of regenerating and restoring dystrophin to d ystrophic muscle; and (2) achieving a higher level of gene transfer to dystrophic muscle compared with adenovirus-mediated direct gene deliv ery. However, as observed in direct gene transfer studies, the ex vivo approach also triggers a cellular immune response which limits the du ration of transgene expression.