Antigen-specific T cells are crucial components for antitumor or antivirus immunity. Adoptive transfer of antigen specific T-cells can induce remission of disease; however, the quality of the T cells that are selected for expansion and adoptive transfer has been identified as a critical factor that determines the persistence of transferred cells. Memory stem (TSCM) T-cells demonstrate evidence of stem cell like properties, such as differentiation potential as well as self-renewal. Thus far, there are no data characterizing antigen-specific TSCM cells with respect to their differentiation pattern, TCR clonotype, or function in comparison to different memory T-cell subsets. In this study, we demonstrate successful generation of CMV-specific T cells derived from naïve (TN) and memory T-cell subsets (TSCM, TCM and TEM) from HLA*A0201 CMV seropositive donors. T cells were sensitized using artificial antigen presenting cells transduced to express HLA-A*0201 and CMVpp65 in the presence of the cytokines IL-7 and IL-15. We then characterized CMVpp65-specific T cells isolated by their binding to tetramers of HLA-A*0201 bearing the immunodominant CMVpp65495-503 peptide, NLVPMVATV (NLV). Upon in vitro expansion in the presence of CMV antigen, both Tet+ TN and TSCM-derived cells differentiated into TCM and TEM. However, during prolonged stimulation with CMV antigen, Tet+ TSCM-derived cells sustained the population of the less differentiated Tcm memory phenotype, as well as a higher proliferative capacity leading to superior expansion of Tet+ CMV-specific T-cells compared to CMV-specific TCM and TEM memory T cell subsets. In functional assays, both TN and TSCM-derived Tet+ cells were able to secrete high level of IFN-γ and TNF-α, as well as CD107a in an antigen-dependent manner. Over 30 days of antigen-specific stimulation, T cell receptor (TCR) gene analysis of the Tet+ T-cells demonstrated a graduated clonal overlap from TSCM to TCM to TEM among the CMV specific T-cells. Furthermore, certain TCR clonotypes were shared within CMV specific T-cells derived from all memory T-cell subsets, indicating that T-cells recognizing the same epitope were derived from the same mother clones. In conclusion, these data provide evidence that antigen-specific TSCM cells constitute an earlier differentiation stage within memory T-cells compared to TCM and TEM cells, with a superior expansion capacity. TSCM cells may therefore serve not only as a durable reservoir, but also an initiator of the memory immune response upon antigen challenge resulting from reactivation of a latent infection. These findings have implications for the design of adoptive immunotherapy strategies employing virus specific CTLs to maximize generation of T-cell population, optimized for in vivo expansion, function and persistence.
Citation Format: Tzu-yun Kuo, Aisha N. Hasan, Richard J. O'Reilly. Functional characterization of virus specific stem cell like memory T cells, their maturation in vitro, and potential contribution to secondary T cell responses [abstract]. In: Proceedings of the Second CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; 2016 Sept 25-28; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(11 Suppl):Abstract nr B050.
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