Adenoviral gene transfer to skeletal muscle: Characterization and manipulation of the adenoviral pathway of infection.

摘要

Adenoviruses have the capacity to deliver large amounts of DNA to the host cell nucleus in a very efficient manner. For this reason, they have been widely exploited for gene therapy applications. In addition, Ad vectors can infect non-dividing cells and accommodate very large transgenes, making them particularly well suited for gene therapy for Duchenne muscular dystrophy, a devastating neuromuscular disorder caused by mutations in the dystrophin gene. The utility of the Ad vector for muscle gene therapy would be greatly enhanced by targeting the virus to muscle specific proteins. In light of recent studies that have introduced evidence suggesting that the Ad classical binding and internalization receptors (coxsackievirus and adenovirus receptor (CAR) and αvβ3 or αvβ5 integrins, respectively) may not be the sole means of Ad entry into many cell types, it is clear that in order to optimize muscle-targeting strategies, a solid understanding of the normal mechanisms of Ad binding and internalization into muscle cells must be attained.; In this study, we used a novel set of Ad vectors that are ablated for binding to CAR, αv integrins, or both to test the degree to which these classical receptors contribute to Ad transduction in muscle cells. We found that (1) Ad can infect muscle cells by alternate pathways, (2) the utilization of these non-classical pathways differs depending on the maturation state and developmental stage of the muscle, and (3) the in vitro muscle cell culture provides a good model in which to further dissect the mechanistic details of Ad infection in myogenic cells.; We redirected Ad to α7β1 integrin, a relatively muscle-specific integrin expressed at high levels in mature and dystrophin-deficient muscle, with the dual goals of increasing efficiency and specificity of muscle transduction. We altered Ad tropism by linking the fiber-ablated and double-ablated Ad vectors to an anti-α7 integrin antibody using bispecific targeting complexes and found that the α7-targeted vectors were able to significantly enhance transduction in C2C12 myoblasts. However, targeting efficacy was subject to variability and future research should examine the mechanisms by which the α7-targeted Ad vectors infect cells in order to identify the factors governing the efficiency of transduction. The continued characterization of the classical and alternate pathways of Ad infection in the context of both native and modified tropism will facilitate the successful design of efficient muscle-targeted Ad vectors capable of transducing muscle cells in a specific and reproducible manner.