Dual Role of Oxidative Stress in Head and Neck Cancer Chemotherapy: Cytotoxicity and Pro-survival Autophagy

Increased intracellular glutathione (GSH) and thioredoxin (Trx) metabolism are mechanisms that are widely implicated in chemotherapy resistance. The current study tests if simultaneous inhibition of both GSH and Trx systems would induce cytotoxicity via oxidative stress in human head and neck cancer (HNSCC) cells and sensitize HNSCC cells to the EGFR inhibitor erlotinib. Inhibition of GSH and Trx with buthionine sulfoximine (BSO) and auranofin (AUR) respectively induced significant decreases in cell viability and clonogenic survival compared to either drug alone in FaDu, Cal-27 and SCC-25 HNSCC cells in vitro and in a Cal-27 xenograft model. BSO+AUR significantly induced parameters of oxidative stress such as increased glutathione and thioredoxin oxidation compared to the other treatment groups in vitro. Glutathione reductase and glutathione peroxidase activity were unaffected by BSO+AUR, but peroxiredoxin activity was significantly suppressed in HNSCC cells treated with AUR and BSO+AUR. Pretreatment with N-acetylcysteine completely reversed BSO+AUR-induced cell killing in FaDu and Cal-27 cells, however, catalase and selenium significantly reversed BSO+AUR-induced cell killing in only FaDu cells. AUR significantly increased caspase 3/7 activity and reduced the viability of Bax/Bak double knockout (DKO) mouse hematopoietic that were reconstituted with Bax (DKO-Bax), but did not affect the viability of DKO cells supporting the role of apoptosis in AUR-induced cell killing. However BSO+AUR decreased caspase 3/7 activity and significantly reduced the viability of both DKO and DKO-Bax MEFs suggesting that that necrosis was involved. The combination of BSO and AUR significantly sensitized FaDu, Cal-27, SCC-25 and SQ20B cells to the EGFR inhibitor Erlotinib in vitro. These results support the