T Cell Receptor (TCR) Gene Transfer with Lentiviral Vectors Allows Efficient Redirection of Tumor Specificity in Naive and Memory T Cells Without Prior Stimulation of Endogenous TCR

Abstract

We investigated the possibility of introducing exogenous T cell receptor (TCR) genes into T cells by lentiviral transduction, without prior stimulation of endogenous TCR with anti-CD3. TCR transfer is used to impose tumor antigen specificity on recipient T cells, but sustained activation required for retroviral transduction may affect the clinical efficacy of engineered T cells. Cytokine stimulation makes T cells susceptible to lentiviral transduction in the absence of TCR triggering, but this advantage has never been exploited for TCR transfer. Autoimmune diseases are a source of high-affinity TCRs specific for self/tumor antigens. We selected, from a patient with vitiligo, a Mart1-specific TCR based on intrinsic interchain pairing properties and functional avidity. After lentiviral transduction of human peripheral blood mononuclear cells, preferential pairing of exogenous α and β chains was observed, together with effective recognition of Mart1⁺ melanoma cells. We tested transduction efficiency on various T cell subsets prestimulated with interleukin (IL)-2, IL-7, IL-15, and IL-21 (alone or in combination). Both naive and unfractionated CD8⁺ T cells could be transduced without requiring endogenous TCR triggering. IL-7 plus IL-15 was the most powerful combination, allowing high levels of transgene expression without inducing T cell differentiation (34 ± 5% Mart1-TCR⁺ cells in naive CD8⁺ and 16 ± 6% in unfractionated CD8⁺). Cytokine-prestimulated, Mart1-redirected naive and unfractionated CD8⁺ cells expanded better than CD3–CD28-prestimulated counterparts in response to both peptide-pulsed antigen-presenting cells and Mart1⁺ melanoma cells. This strategy allows the generation of tumor-specific T cells encompassing truly naive T cells, endowed with an intact proliferative potential and a preserved differentiation stage.