Helper-dependent adenovirus vector (AdV)-mediated full-length dystrophin expression leads to significant mitigation of the dystrophic phenotype of the mdx mouse. However, dystrophin, as a neoantigen, elicits antibody formation. As an alternative approach, we evaluated gene transfer of full-length murine utrophin, a functional homologue of dystrophin that is normally present only at the neuromuscular junction. A single injection in the tibialis anterior (TA) muscle of the helper-dependent adenovirus vector encoding utrophin provided very good transduction, with 58% of fibers demonstrating sarcolemmal utrophin expression in the neonates, and 35% utrophin-positive (Utr(+)) fibers in adults. The presence of utrophin prevented extensive necrosis in the neonates, halted further necrosis in the adults, and led to restoration of sarcolemmal expression of dystrophin-associated proteins up to 1 year after injection. Marked physiological improvement was observed in both neonates and adults. Neither increased humoral responses nor cellular immune responses were evident. However, there was a time-related decline of the initial high utrophin expression. Although viral DNA persisted in animals that were injected in the neonatal stage, viral DNA levels decreased in muscles of adult mice. These results demonstrate that although utrophin gene transfer leads to amelioration of the dystrophic phenotype, the effects are not sustained upon loss of utrophin expression.
Adenoviral vectors that use the coxsackievirus and adenovirus receptor do not transduce mature muscle efficiently. Group B adenoviruses use CD46 as their cell attachment receptor. To evaluate the utility of vectors based on group B adenoviruses for gene transfer to human skeletal muscle, we assessed the expression of CD46 in biopsied normal skeletal muscle samples and in muscles from patients with Duchenne muscular dystrophy. Transcript levels of CD46 were extremely low in mature muscle and CD46 immunoreactivity was detected only on blood vessels in the muscle sections. Although myoblasts cultured from biopsied samples had robust cell surface CD46 expression by flow cytometry, CD46 transcript levels were barely detectable after differentiation of the myoblasts into myotubes. The myotubes were also much less susceptible to infection with an adenoviral vector carrying the fiber of serotype 35 adenovirus (AdF35). These results suggest that for skeletal muscle, vectors derived from group B adenoviruses may not be a suitable alternative to the commonly used Ad5 vectors.
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