NK cell subsets that express appropriate receptors to positively recognize and respond to tumors are relatively abundant compared to antigen-specific T or B cells and thus this can provide a mechanism for rapid response against malignant cells. Considering tumor-induced suppression of NK cell activity it is of interest to study the effect of NK-cell stimulatory cytokines, such as interferon-alpha or IL-2 applied in immunotherapy of melanoma (MM), on the functional and receptor characteristics of the less investigated CD16-defined NK cells and their dim and bright subsets. In this study 40 patients with MM in clinical stage IV were investigated prior to therapy. NK cytotoxicity against K562 cell line and the expression of its activating, NKG2D and CD161, as well as inhibitory KIR, CD158a and CD158b receptors on CD3-CD16+ NK cells and their CD16dim and CD16bright subsets, as well as mRNA of transcription molecule IRF-1 were evaluated after 18 h in vitro treatment of PBL in medium RPMI alone, with interferon alpha (IFN, 250U/ml) and rh IL-2 (200U/ml). In vitro treatments with IFN and IL-2 induced significant enhancement of NK cell cytotoxic activity and the expression of NKG2D and CD161, after both treatments and after IFN treatment, respectively, CD3-CD16+ NK cells. We show for the first time in MM patients that IL-2-induced increase in NK cytotoxicity correlates with its induction of NKG2D. Moreover, we further show that upregulation of NKG2D with IFN and IL-2 obtained on the cytotoxic CD16bright subset correlates with enhancement of NK cytotoxicity with these cytokines. Contrary to this, IFN-induced upregulation of CD161 in this subset did not correlate with enhancement of NK cytotoxicity by this cytokine. Analyses of the regulatory CD16dim NK subset, except for the effect of IFN on NKG2D expression, shows the same type of cytokine-induced increase in NKG2D and CD161 as in the CD16bright subset, although, without correlation with obtained increase in cytotoxicity. The investigated cytokines had no effect on the expression of CD158a and CD158b on CD3-CD16+ NK cells and on their bright and dim subsets. Evaluation of mRNA of transcription molecule IRF-1 shows that, unlike IL-2, IFN\#945; signficantly up-regulates its expression. Considering the importance of CD16 in both direct and ADCC cytotoxicity, in this study we show for the first time that in vitro enhancement of NK cell cytotoxic activity with investigated cytokines in MM patients correlates to a grater extent with the induction of the activating NKG2D receptor on the CD16bright subset than on CD3-CD16+ NK cells. As NK cell mediated killing of tumor cells may depend on the balance between stimulatory and inhibitory signaling, induction of stimulatory NKG2D receptor expression through the use of these cytokines, especially on the CD16bright NK cell subset, may enhance tumor cytolysis.
Citation Information: In: Proc Am Assoc Cancer Res; 2009 Apr 18-22; Denver, CO. Philadelphia (PA): AACR; 2009. Abstract nr 4162.
100th AACR Annual Meeting-- Apr 18-22, 2009; Denver, CO