Hypoxia regulation of phosphokinases and the prognostic value of pAKT in breast cancer.

Tumor hypoxia results in poor treatment response and is an indicator of poor outcome in cancer patients. TRIB3 is a hypoxia-upregulated protein involved in the ability of breast cancer cells to survive in hypoxic conditions. It is also involved in the prognosis of cancer patients, possibly by affecting several kinase-signaling pathways. We set out to establish which kinase-signaling pathways are regulated by hypoxia and whether these kinases are relevant for breast cancer prognosis. Using a phosphokinase antibody array comparing cells cultured under hypoxic conditions with those cultured during normoxia, we found that the phosphorylation status of ERK1/2, AKT, p70 S6 kinase, Lck and STAT3 was altered in both MCF7 and MDA-MB-231 breast cancer cells. Using Western blotting, we found that phosphorylated AKT (pAKT) increased in hypoxic conditions. Knockdown of TRIB3 attenuated this effect of hypoxia on AKT activation. Both pAKT and TRIB3 were expressed in pimonidazole-positive, hypoxic areas of human breast cancer tumors. In breast cancer patients significantly lower 5-year disease-free survival was observed for the pAKT-positive compared to the pAKT-negative group (64.6% vs 86.1%, p=0.03). In conclusion, the phosphorylation status of AKT is increased in hypoxic conditions and TRIB3 knockdown attenuates this response. Furthermore, pAKT expression denotes a worse prognosis in breast cancer patients. The hypoxia-related activation of AKT could explain the resistance to various treatments including chemotherapy and radiotherapy.