NK cell function in cancer patients is severely impaired, but the mechanism underlying this impairment is not clearly understood. In this study we show evidence that TGF-β1 secreted by tumors is responsible for the poor NK lytic activity via down-regulating an NK-activating receptor, NKG2D. The plasma level of TGF-β1 in human lung cancer or colorectal cancer patients was elevated compared with that in normal volunteers, and this elevation was inversely correlated with surface expression of NKG2D on NK cells in these patients. Incubation of NK cells with plasma obtained from cancer patients specifically down-modulated surface NKG2D expression, whereas addition of neutralizing anti-TGF-β1 mAbs completely restored surface NKG2D expression. Likewise, incubation of NK cells and lymphokine-activated killer cells with TGF-β1 resulted in dramatic reduction of surface NKG2D expression associated with impaired NK cytotoxicity. Modulation of NKG2D by TGF-β1 was specific, as expression of other NK receptors, CD94/NKG2A, CD44, CD16, 2B4, or CD56, was not affected by TGF-β1. Impaired NK cytotoxicity by TGF-β1 was not due to alteration of lytic moieties, such as perforin or Fas, or apoptotic pathway, but, rather, appeared to be due to lack of NKG2D expression. Taken together, our data suggest that impaired NK function in cancer patients can be attributed to down-modulation of activating receptors, such as NKG2D, via secretion of TGF-β1.

The presence of membrane-bound TGF-β1 (mTGF-β1) has been recently observed in regulatory T cells, but only a few studies have reported the same phenomenon in cancer cells. In this study, we investigate the regulation of mTGF-β1 expression in five head and neck squamous cell carcinoma cell lines using FACS analysis. Through blocking Ab and exogenous cytokine treatment experiments, we found that expression of mTGF-β1 is significantly induced by the activated immune cell-derived factor IFN-γ. In addition, IFN-γ and TNF-α are shown to have a synergistic effect on mTGF-β1 expression. Moreover, we found that exogenous TNF-α induces endogenous TNF-α mRNA expression in an autocrine loop. In contrast to previous reports, we confirm that, in this model, mTGF-β1 is neither a rebound form of once-secreted TGF-β1 nor an activated form of its precursor membrane latency-associated peptide. Inhibitors of transcription (actinomycin D), translation (cycloheximide), or membrane translocation (brefeldin A) effectively block the induction of mTGF-β1, which suggests that induction of mTGF-β1 by IFN-γ and/or TNF-α occurs through de novo synthesis. These findings suggest that some cancer cells can detect immune activating cytokines, such as IFN-γ and TNF-α, and actively block antitumor immunity by induction of mTGF-β1.