Retrovirus-mediated gene transfer in polyclonal T cells results in lower apoptosis and enhanced ex vivo cell expansion of CMV-reactive CD8 T cells as compared with EBV-reactive CD8 T cells

Abstract

AbstractTo modulate alloreactivity after hematopoietic stem cell transplantation, “suicide” gene-modified donor T cells (GMCs) have been administered with an allogeneic T-cell–depleted marrow graft. We previously demonstrated that such GMCs, generated after CD3 activation, retrovirusmediated transduction, and G418 selection, had an impaired Epstein-Barr virus (EBV) reactivity, likely to result in an altered control of EBV-induced lymphoproliferative disease. To further characterize the antiviral potential of GMCs, we compared the frequencies of cytomegalovirus (CMV)–specific CD8+T (CMV-T) cells and EBV-specific CD8+T (EBV-T) cells within GMCs from CMV- and EBV-double seropositive donors. Unlike anti-EBV responses, the anti-CMV responses were not altered by GMC preparation. During the first days of culture, CMV-T cells exhibited a lower level of CD3-induced apoptosis than did EBV-T cells. In addition, the CMV-T cells escaping initial apoptosis subsequently underwent a higher expansion rate than EBV-T cells. The differential early sensitivity to apoptosis could be in relation to the “recent activation” phenotype of EBV-T cells as evidenced by a higher level of CD69 expression. Furthermore, EBV-T cells were found to have a CD45RA–CD27+CCR7–effector memory phenotype, whereas CMV-T cells had a CD45RA+CD27–CCR7–terminal effector phenotype. Such differences could be contributive, because bulk CD8+CD27–cells had a higher expansion than did bulk CD8+CD27+cells. Overall, ex vivo T-cell culture differentially affects apoptosis, long-term proliferation, and overall survival of CMV-T and EBV-T cells. Such functional differences need to be taken into account when designing cell and/or gene therapy protocols involving ex vivo T-cell manipulation.