Cytokine-induced killer cells are terminallydifferentiated activated CD8 cytotoxic T-EMRA lymphocytes

Objective: Cytokine-induced killer cells (CIK) are CD3⁺CD56⁺ T cells with natural killer (NK)-like cytotoxic activity used for the immunotherapy of tumors. We aimed to fully characterize CIK cells and define their ontogeny. Materials and Methods: CIK were generated in vitro by stimulation of peripheral blood mononuclear cells or T-cell subsets with interferon-γ, anti-CD3 and interleukin-2. They were fully characterized in terms of phenotype, cytotoxic activity, and gene expression with respect to circulating CD3⁺CD56⁺ cells, NK cells, and CD56^- T cells present in CIK cultures. Results: We demonstrate that CIK are terminally differentiated CD8 T cells that derive from proliferating CD3⁺CD56^-CD8⁺ T cells. They express polyclonal T-cell receptor Vβ chains and have acquired CD56, NKG2D, and large granular lymphocyte morphology, but lack expression of most NK-specific activating (NKp30, NKp44, NKp46) and inhibitory (KIR2DL1, KIR2DL2, KIR3DL1, NKG2A, CD94) receptors, and can kill K562 targets. Circulating CD3⁺CD56⁺ cells are also CD8⁺CD16^-, but are oligoclonal, poorly cytotoxic for K562, and express lower levels of CD56 and NKG2D. Gene profiling of CIK, CD56^- T and NK cells present at the end of culture shows that differences are much more limited between CIK and CD56^- T compared to CIK and NK cells. Most of the genes upregulated in CIK cells compared to CD56^- T cells are part of the tumor necrosis factor gene network. Conclusions: The CIK phenotype, that is CD45RA⁺, CCR7^-, CD62L-weakly positive, CD11a⁺, CD27⁺, CD28^-, macrophage inflammatory protein 1α⁺, perforin⁺, Fas ligand⁺ coincides almost exactly with that described for the T RA⁺ effector memory CD27 single positive subset of terminally differentiated human memory T cells.