The Cooperative Induction of Hypoxia-Inducible Factor-1α and STAT3 during Hypoxia Induced an Impairment of Tumor Susceptibility to CTL-Mediated Cell Lysis

Hypoxia is an essential component of tumor microenvironment. In this study, we investigated the influence of hypoxia (1% PO2) on CTL-mediated tumor cell lysis. We demonstrate that exposure of target tumor cells to hypoxia has an inhibitory effect on the CTL clone (Heu171)-induced autologous target cell lysis. Such inhibition correlates with hypoxia-inducible factor-1 alpha (HIF-1 alpha) induction but is not associated with an alteration of CTL reactivity as revealed by granzyme B polarization or morphological change. Western blot analysis indicates that although hypoxia had no effect on p53 accumulation, it induced the phosphorylation of STAT3 in tumor cells by a mechanism at least in part involving vascular endothelial growth factor secretion. We additionally show that a simultaneous nuclear translocation of HIF-1 alpha and phospho-STAT3 was observed. Interestingly, gene silencing of STAT3 by small interfering RNA resulted in HIF-1 alpha inhibition and a significant restoration of target cell susceptibility to CTL-induced killing under hypoxic conditions by a mechanism involving at least in part down-regulation of AKT phosphorylation. Moreover, knockdown of HIF-1 alpha resulted in the restoration of target cell lysis under hypoxic conditions. This was further supported by DNA microarray analysis where STAT3 inhibition resulted in a partly reversal of the hypoxia-induced gene expression profile. The present study demonstrates that the concomitant hypoxic induction of phopho-STAT3 and HIF-1 alpha are functionally linked to the alteration of non-small cell lung carcinoma target susceptibility to CTL-mediated killing. Considering the eminent functions of STAT3 and HIF-1 alpha in the tumor microenvironment, their targeting may represent novel strategies for immunotherapeutic intervention. The Journal of Immunology, 2009, 182: 3510-3521.