Immunization against the transgene but not the TetON switch reduces expression from gutless adenoviral vectors in the brain.

Immune responses against vectors or encoded transgenes can impose limitations on gene therapy. We demonstrated that tetracycline-regulated high-capacity adenoviral vectors (HC-Ads) sustain regulated transgene expression in the brain even in the presence of systemic pre-existing immune responses against adenoviruses. In this study we assessed whether systemic pre-existing immune responses against the transgene products, i.e., β-Gal or the tetracycline-dependent (TetON) regulatory transcription factors (rtTA2SM2 and the tTSKid), affect transgene expression levels and the safety profile of HC-Ads in the brain. We pre-immunized mice with plasmids encoding the TetON switch expressing rtTA2SM2 and the tTSKid or β-Gal. HC-Ads expressing β-Gal under the control of the TetON switch were then injected into the striatum. We assessed levels and distribution of β-Gal expression, and evaluated local inflammation and neuropathological changes. We found that systemic immunity against β-Gal, but not against the TetON switch, led to inflammation and reduction of transgene expression in the striatum. Therefore, the regulatory TetON switch appears to be safe to use, and capable of sustaining transgene expression in the brain even in the presence of an immune response against its components. Systemic immunity against the transgene had the effect of curtailing its expression, thereby affecting the efficacy and safety of gene delivery to the brain. This factor should be considered when developing gene therapies for neurological use.