Gene therapy is a potential route for the delivery of secreted therapeutic proteins, but pharmacologic control of expression will generally be required for optimal safety and efficacy. Previous attempts to achieve regulated expression in large animal models have been thwarted by transient expression or immune responses to regulatory proteins. We evaluated the ability of the dimerizer-regulated gene expression system to achieve controlled, long-term production of erythropoietin (Epo) following intramuscular administration of adeno-associated virus (AAV) vectors to 16 primates. All animals showed dose-responsive and completely reversible elevation of Epo and hematocrit in response to the dimerizer rapamycin, or analogs with reduced immunosuppressive activity, administered intravenously or orally. Animals that received optimized dual vectors showed persistent regulated expression for the duration of the study, with no apparent immune response to Epo or the regulatory proteins. Similar results were obtained with single vectors incorporating both the Epo and regulatory genes, including those packaged into serotype 1 AAV vectors to allow use of lower viral doses. For the longest-studied animal, regulated expression has persisted for more than 6 years and 26 induction cycles. These data indicate that one-time or infrequent gene transfer followed by dimerizer regulation is a promising approach for delivery of therapeutic proteins.