Glycogen storage disease II (GSD-II;Pompe disease;MIM 232300) is a lethal muscular dystrophy caused by the deficiency of acid alpha-glucosidase (GAA;acid maltase). No curative therapy is available for GSD-II. Obstacles to gene therapy in GSD-II include very high GAA requirements and the occurence of neutralizing anti-GAA antibodies. Gene therapy with adeno-associated virus (AAV) vectors was advanced by the availability of AAV serotypes 6-9, termed myotrophic AAV vectors in this application. The general hypothesis states that myotrophic AAV vectors will express efficacious GAA in the striated muscles of GSD-II mice, including the heart, diaphragm, and skeletal muscles. Myotrophic AAV2 vectors pseudotyped as AAV7, AAV8, and AAV9 will be delivered intravenously to analyze the systemic delivery of these vectors to striated muscle. Specific Hypothesis 1: Pseudotyped AAV vectors containing a muscle-specific promoter will express high levels of GAA in the striated muscle of GSD-II mice. Correction of GAA deficiency and glycogen storage in the heart, diaphragm, and skeletal muscle will be analyzed for each serotype in GAA- knockout (GAA-KO) mice. Myotrophic AAV vectors will be administered to infantile GAA-KO mice, demonstrating efficacy in Pompe disease. Specific Hypothesis 2: An AAV vector will evade cellular and humoral immune responses to achieve long-term efficacy in GSD-II mice. Cellular and humoral immune responses to hGAA expression restricted to striated muscles will be analyzed. ELISpot assays will be performed to analyze antigen-specific T cell responses to hGAA and AAV capsids, and flow cytometric detection of additional lymphocyte markers will evaluate the immune response to AAV vectors. Specific Hypothesis 3: Liver-specific expression will Induce tolerance to introduced GAA and enhance the efficacy of myotrophic AAV vectors in GSD-II mice. Tolerance to GAA will be induced by liver-specific expression of GAA in immunocompetent GAA-KO mice. T regulatory cells will be quantitated in tolerant GAA-KO mice;furthermore, T regulatory cells will be depleted to demonstrate their role in establishing tolerance to GAA. A novel strategy will deliver an AAV vector encoding chimeric GAA by intravenous injection and isolated limb delivery to convert skeletal muscle to a depot for secreted GAA. Efficacious muscle-targeted gene therapy in GSD-II will have implications for gene therapy in other muscular dystrophies and myopathies.

National Institute of Health (NIH)
National Heart, Lung, and Blood Institute (NHLBI)
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Special Emphasis Panel (ZRG1-GTIE-A (01))
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Skarlatos, Sonia
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Duke University
Schools of Medicine
United States
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