T cell activation modulates retrovirus-mediated gene expression.
Important considerations for T lymphocyte-based gene therapy include efficient gene delivery and expression in primary, human T cells. In this study, retrovirus-mediated gene transfer and the fate of proviral gene expression were evaluated in human T cells activated using (1) immobilized anti-CD3 monoclonal antibody (MAb) plus interleukin 2, or (2) cis costimulation using beads carrying coimmobilized anti-CD3 and anti-CD28 MAbs. By cross-linking the CD3 and CD28 receptors, these MAbs mimic in vivo signaling events, leading to cytokine production and proliferation. A modified human interleukin 1beta (IL-1beta) cDNA inserted into the MFG retroviral vector served as an indicator gene. Retroviral transduction frequencies were similar for T lymphocytes activated by the respective methods. However, early after MAb stimulation and virus exposure, proviral gene expression was greater at the RNA and protein levels in optimized anti-CD3/anti-CD28 bead-activated T cells, corresponding with augmented endogenous cytokine responses and mitogenesis. Proviral gene expression was not regulated by extrinsic cell factors present in activated T cell supernatants. Regardless of the MAb stimulation method, proviral IL-1beta expression declined in later T cell cultures concomitant with a decrease in cellular cytokines. Restimulation by either method reinduced both T cell activity and vector expression. Our finding that proviral gene regulation is downmodulated in the absence of T cell signaling events has implications for clinical strategies using retrovirus-modified T cells.
Vince Lombardi Gene Therapy Laboratory, Immunotherapy, Research and Treatment Institute, St. Luke's Medical Center, Milwaukee, WI 53201, USA.
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D015496:CD4-Positive T-Lymphocytes / Q000378:metabolism; D018414:CD8-Positive T-Lymphocytes / Q000378:metabolism; D002478:Cells, Cultured; D016207:Cytokines / Q000378:metabolism; D005786:Gene Expression Regulation; D006801:Humans; D007375:Interleukin-1 / Q000235:genetics / Q000378:metabolism*; D008213:Lymphocyte Activation; D011533:Proviruses / Q000235:genetics; D012190:Retroviridae / Q000235:genetics; D013601:T-Lymphocytes / Q000276:immunology / Q000378:metabolism* / Q000821:virology; D014162:Transfection; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*
D016207:Cytokines; D007375:Interleukin-1; D014764:Viral Proteins