The ligands for natural cytotoxicity receptors NKp30, NKp44 and NKp46 are currently unknown. However; we postulate that at least NKp46 and NKp30 may be involved in autologous gastric tumor cell recognition since lytic activity was abrogated in the presence of blocking antibody against these receptors. Since no significant change was observed in NKG2D expression on expanded NK cells, we did not directly test the involvement of this activating

receptor in autologous gastric tumor cell cytotoxicity. The fact that autologous cytotoxicity was not completely inhibited by a combination of anti-DNAM-1, NKp46, NKp44 and NKp30 may indicate that NKG2D or other Selleckchem Palbociclib unidentified receptors may also be involved. Importantly, the interaction between NK cell receptors and their ligands has recently been shown to abrogate NK cell mediated cytotoxicity of human and mouse melanoma cell lines [32]. Of note, both tumor cell lines also expressed high levels of Fas which is recognized to establish cell death upon interaction with its ligand, Fas-ligand [33]. In order to test the possibility of target cell-induced killing of the expanded NK cells, all NK cells were evaluated for Fas and Fas-ligand high throughput screening compounds expression before and after ex-vivo expansion. Although expanded NK cells up-regulated high levels of Fas, they

did not express Fas-ligand (data not shown). It has been GPX6 suggested that in order to overcome self tolerance, multiple activating receptor-ligand interactions should be engaged [31]. Indeed, multiple

activating interactions appear to be involved in autologous cytotoxicity of tumor cells derived from patient 1 when the inhibition of cytotoxicity, in the presence of all 4 antibodies, is compared with DNAM-1 or NKp30 alone (P = 0.0356 and P = 0.0165, respectively). In contrast, no significant additional decline in autologous cytotoxicity was observed for patient 2 when cytolytic activity of all four activating receptors was compared to NKp46 alone (P = 0.7359). We postulate that these data reflect variation in expression of receptor-ligand combination in humans that are known to be operative in the control of NK cell cytotoxic activity. These variations include HLA and KIR polymorphism as well as tumor type and tumor origin (e.g. primary versus metastatic tumor cells). This is illustrated in a recent report on studies in patients with multiple myeloma [34] where the investigators demonstrated no specific association of autologous NK cell cytotoxicity with a single activating NK cell receptor. In fact, autologous cytotoxic effects were more likely mediated by several activating NK cell receptors which is also in agreement with a previous report [35] demonstrating that natural cytotoxicity of resting NK cells requires co-activation by more than one receptor.