LAUSR

Pompe disease is caused by mutations in the acid alpha-glucosidase gene (GAA) that is responsible for processing lysosomal glycogen. Patients with Pompe disease exhibit clinical phenotypes across a variety of tissues, including glycogen buildup in cells, deficits in cardiac, respiratory, and skeletal muscle function, and CNS pathology. Previously, we described novel multi-tissue GAA expressing rAAV vectors that reduced glycogen in skeletal muscle, heart, and brain 4 weeks after treatment of GAA-/- mice. We now describe longer term follow up of animals treated with hybrid liver/muscle/neuronal vectors. Mice were dosed with rAAV vectors at 1x1013, 3x1013, or 1x1014 vg/kg, and in-life measures and bioanalytical endpoints were assessed relative to vehicle-treated GAA -/- or WT littermate control mice. Assessment of diaphragm, heart, and skeletal muscle tissue indicated dose-dependent increases in hGAA expression and activity, and reduced glycogen accumulation in these tissues. Vectors with incremental expression in liver contributed to reduced antibody reactivity to hGAA, compared to mice dosed with a control vector containing a muscle-only promoter element, although the muscle-only promoter constructs achieved high levels of GAA activity and glycogen reduction in key tissues. Finally, we observed dose-dependent expression and activity of hGAA in nervous system tissue and demonstrated that a significant reduction in glycogen levels in spinal cord could be achieved in mice dosed with a systemically-administered rAAV8 vector construct. Our findings represent the first nonclinical application of engineered hybrid promoters providing tissue-specific neuromuscular transgene expression combined with immune-tolerizing liver expression and suggest key attributes for refinement of vector design for the clinical translation of an rAAV-based gene therapy for Pompe disease