ADENOVIRUS-MEDIATED DELIVERY INTO MYOCYTES OF MUSCLE GLYCOGEN-PHOSPHORYLASE, THE ENZYME-DEFICIENT IN PATIENTS WITH GLYCOGEN-STORAGE-DISEASETYPE-V

The feasibility of using adenovirus as a vector for the introduction o f glycogen phosphorylase activity into myocytes has been examined. We used the C2C12 myoblast cell line to assay the impact of phosphorylase gene transfer on myocyte glycogen metabolism and to reproduce in vitr o the two strategies proposed for the treatment of muscle genetic dise ases, myoblast transplantation and direct DNA delivery. In this study, a recombinant adenovirus containing the muscle glycogen phosphorylase cDNA transcribed from the cytomegalovirus promoter (AdCMV-MGP) was us ed to transduce both differentiating myoblasts and nondividing mature myotube cells. Muscle glycogen phosphorylase mRNA levels and total pho sphorylase activity were increased in both cell types after viral trea tment although more efficiently in the differentiated myotubes. The in crease in phosphorylase activity was transient (15 days) in myoblasts whereas in myotubes higher levels of phosphorylase gene expression and activity were reached, which remained above control levels for the du ration of the study (20 days). The introduction of muscle phosphorylas e into myotubes enhanced their glycogenolytic capacity. AdCMV-MGP-tran sduced myotubes had lower glycogen levels under basal conditions. In a ddition, these engineered cells showed more extensive glycogenolysis i n response to both adrenaline, which stimulates glycogen phosphorylase phosphorylation, and carbonyl cyanide m-chlorophenylhydrazone, a meta bolic uncoupler. In conclusion, transfer of the muscle glycogen phosph orylase cDNA into myotubes confers an enhanced and regulatable glycoge nolytic capacity. Thus this system might be useful for delivery of mus cle glycogen phosphorylase and restoration of glycogenolysis in muscle cells from patients with muscle phosphorylase deficiency (McArdle's d isease).z