The successful application of gene therapy for the treatment of genetic diseases such as Fabry is reliant on the development of vectors that are safe and that facilitate sustained expression of therapeutic levels of the transgene product. Here, we report that intravenous administration of a recombinant AAV2 vector encoding human α-galactosidase A under the transcriptional control of a liver-restricted enhancer/promoter (AAV2/DC190-αgal) generated significantly higher levels of expression in BALB/c and Fabry mice than could be realized using the ubiquitous CMV promoter (AAV2/CMVHI-αgal). Moreover, AAV2/DC190-αgal-mediated hepatic expression of α-galactosidase A was sustained for 12 months in BALB/c mice and was associated with a significantly reduced immune response to the expressed enzyme. Subsequent challenge of the AAV2/DC190-αgal-treated animals with recombinant human α-galactosidase A at 6 months failed to elicit the production of anti-α-galactosidase A antibodies, suggesting the induction of immune tolerance in these animals. The levels of expression attained with AAV2/DC190-αgal in the Fabry mice were sufficient to reduce the abnormal accumulation of globotriaosylceramide in the liver, spleen, and heart to basal levels and in the kidney by approximately 40% at 8 weeks. Together, these results demonstrate that AAV2-mediated gene transfer that limits the expression of α-galactosidase A to the liver may be a viable strategy for treating Fabry disease.