Introduction and Objective: The potential role of natural killer (NK) and lymphokine activated killer (LAK) cells in cancer immunotherapy is expanding and subsequent investigation of tumor escape mechanisms of innate immunity is an exciting area of basic science and clinical inquiry. Tumour hypoxia has been shown to be associated with malignant progression in cancer cells and an independent prognostic indicator of poor clinical outcome for patients with various cancers. We have previously shown that this hypoxia induced progression can be attenuated by low concentrations of nitric oxide (NO) mimetic agents. We therefore postulate that a hypoxic environment may also have a significant effect on the ability of immune cells to target and kill tumor cells. We determine the effect of hypoxia on natural killer cell cytotoxicity. Furthermore, we determine the role of NO to improve prostate cancer cell killing by circulating immune cells. Methods: Two human prostate cancer cell lines PC3 and DU145, as well as a natural killer cell (NK) sensitive leukemia cell line K562, were used for cytotoxicity experiments. Cells were cultured in hypoxic (1% O2) or standard (20% O2) conditions for 24 hours prior to exposure to effector immune cells. Cytotoxicity was assessed using a standard four hour chromium release assay to measure NK activity of fresh human peripheral blood lymphocytes, as well as IL-2 stimulated LAK cells. Furthermore, we investigated the effect of hypoxia on the tumor expression of ligands (MHC class 1 chain related molecules (MIC)) for NK activating receptors with flow cytometry. To determine the ability of NO to modulate effects of tumor hypoxia, we cultured the cell lines with an NO mimetic, GTN, to determine if it could inhibit the hypoxic effects on tumor cell killing. Results: Results of the cytotoxicity assays reveal that culturing cell lines in a low oxygen environment will significantly decrease NK and LAK cell killing. This reduction of cytotoxicity was found to be associated with a tumor cell shedding of MIC A, B molecules. As well, in both the leukemia and prostate cancer cell lines, the addition of low molar concentrations of GTN did significantly attenuate the hypoxia-associated decreased cell killing. Conclusions: These findings indicate hypoxia leads to cellular adaptive responses of prostate cancer cells which interfere with immune effector cell killing, due at least in part by tumor shedding of MIC. These hypoxia related effects may be inhibited by the addition of low concentrations of nitric oxide mimetics. These novel findings suggest that manipulation of nitric oxide signalling may be a beneficial adjuvant in cell based adoptive immunotherapy protocols for prostate cancer.

[Proc Amer Assoc Cancer Res, Volume 47, 2006]