Analysis of muscle creatine kinase regulatory elements in recombinant adenoviral vectors

Adenoviral gene transfer holds promise for gene therapy, but effective tran sduction of id large and distributed tissue such as muscle will almost cert ainly require systemic delivery. In this context, the use of muscle-specifi c regulatory elements such as the muscle creatine kinase (MCK) promoter and enhancer will avoid potentially harmful ectopic expression of transgenes. We describe here the development and testing of adenoviral vectors containi ng small, striated muscle-specific, highly active MCK expression cassettes. One of these regulatory elements (CK6) is less than 600 bp in length and i s 12% as active as the CMV promoter/enhancer in muscle. A recombinant adeno viral vector containing this regulatory element retains very high muscle sp ecificity, expressing 600-fold higher levels of transgene in muscle than in liver. Muscle-specific regulatory elements may also increase persistence o f transduced muscle cells. Adenoviral transduction of dendritic cells has b een shown to stimulate cytotoxic T-lymphocyte (CTL) responses directed agai nst transgene epitopes. We show that human dendritic cells infected in vitr o with MCK-containing adenoviruses do not express significant levels of tra nsgene. Furthermore, while adenoviral vectors containing nonspecific promot ers are normally cleared from muscle tissue within 1 month, we show that MC K-containing vectors express significant levels of transgene 4 months after intramuscular injection.