Activation-Dependent Expression of an Interleukin-2 Transgene Mediated by Transduction with a Lentiviral Vector Enhances T Cell Functions.

Adoptive T cell immunotherapy (ATCI) is an evolving strategy that explores antigen-specific T cells manipulated ex vivo as therapeutic agents. Although the concept of ATCI has been tested clinically, with success in the treatment of post-transplant EBV induced lymphoproliferative disease, one of the major obstacles hindering its application to other malignancies is the procurement of tumor-specific T cells that possess potent anti-tumor functions even in the inhibitory environment at the tumor sites. This study aims to genetically engineer enriched viral specific T cells for improved immune functions. A self-inactivating lentiviral vector (SIN) CD69p-IL2 was constructed to encode the transgene interleukin-2 (IL2) under the control of a human CD69 promoter (CD69p), and this vector was tested in ex vivo cultivated EBV-specific T cells. SIN vector allows a high degree of autonomy for the internal promoter, and CD69 expression in the T cells is closely associated with T cell activation. Experiments showed that the SIN vectors efficiently transduced EBV-specific T cells, both CD4 and CD8. Furthermore, the newly cloned CD69p exhibited a higher degree of responsiveness to physiological antigen stimulation than the early promoter from the cytomegalovirus (CMVp). In response to stimulation by EBV-infected B cells, the percentage of IL2 expressing cells was 2 fold higher for the activated CD69p-IL2 transduced T cells than the non-transduced, or the CMVp-IL2 transduced, counterparts. In correlation with the stronger IL2 expression, 3 fold more T cells expressed the anti-viral cytokine interferon-γ (IFN-γ) in the CD69p-IL2 transduced T cells than the CMVp-IL2 transduced, and the IFN-γ expression at the single cell level was 2 fold higher in the former, indicating an enhanced functionality. While the culture supernatant from the CMVp-IL2 transduced T cells contained IL2 at a concentration 2000 fold higher than the non-transduced T cells, the IL2 level in the media from the CD69p-IL2 transduced T cells was comparable to that in the control, suggesting the IL2 expression mediated by the CD69p more relevant to T cell functions than the CMVp. These results may serve as a foundation for the further development and clinical application of specific T cells engineered for enhanced immune functions.