Susceptibility of Human Head and Neck Cancer Cells to Combined Inhibition of Glutathione and Thioredoxin Metabolism

Arya Sobhakumari,Elise V M Fletcher,Sean M Martin,C Michael Knudson,Andrean L Simons,Arlene D Parsons,Douglas R Spitz,Laurie Love-Homan,Jinah Choi

doi:10.1371/journal.pone.0048175

Abstract

Increased glutathione (GSH) and thioredoxin (Trx) metabolism are mechanisms that are widely implicated in resistance of cancer cells to chemotherapy. The current study determined if simultaneous inhibition of GSH and Trx metabolism enhanced cell killing of human head and neck squamous cell carcinoma (HNSCC) cells by a mechanism involving oxidative stress. Inhibition of GSH and Trx metabolism with buthionine sulfoximine (BSO) and auranofin (AUR), respectively, induced significant decreases in clonogenic survival compared to either drug alone in FaDu, Cal-27 and SCC-25 HNSCC cells in vitro and in vivo in Cal-27 xenografts. BSO+AUR significantly increased glutathione and thioredoxin oxidation and suppressed peroxiredoxin activity in vitro. Pre-treatment with N-acetylcysteine completely reversed BSO+AUR-induced cell killing in FaDu and Cal-27 cells, while catalase and selenium supplementation only inhibited BSO+AUR-induced cell killing in FaDu cells. BSO+AUR decreased caspase 3/7 activity in HNSCC cells and significantly reduced the viability of both Bax/Bak double knockout (DKO) and DKO-Bax reconstituted hematopoietic cells suggesting that necrosis was involved. BSO+AUR also significantly sensitized FaDu, Cal-27, SCC-25 and SQ20B cells to cell killing induced by the EGFR inhibitor Erlotinib in vitro. These results support the conclusion that simultaneous inhibition of GSH and Trx metabolism pathways induces oxidative stress and clonogenic killing in HNSCCs and this strategy may be useful in sensitizing HNSCCs to EGFR inhibitors.

Highlights

Acquired resistance to chemotherapy is a major obstacle to successful head and neck squamous cell carcinoma (HNSCC) treatment
These results suggest that buthionine sulfoximine (BSO)+AUR induced oxidative stress via increased GSH and Trx oxidation in HNSCC cells
Increased GSH and Trx metabolism have been known for years to be correlated with high tumor aggression and resistance to chemotherapy [4,5,6,7]

Summary

Introduction

Acquired resistance to chemotherapy is a major obstacle to successful head and neck squamous cell carcinoma (HNSCC) treatment. Numerous studies over the years have explored strategies of individually inhibiting GSH or Trx metabolism in addition to conventional chemotherapy agents, but have yielded variable results [14,15,16] probably due to the redundant protective functions of these systems [17,18,19,20] Given that both systems detoxify H2O2 and use NADPH as reducing equivalents, it is logical that both GSH and Trx systems have overlapping and redundant functions in the detoxification of ROS. To overcome the redundancy in these pathways as they relate to resistance to therapy in HNSCC, the current study determined the effect of simultaneously inhibiting both the GSH and Trx metabolism using buthionine sulfoximine (BSO; an inhibitor of GSH synthesis), and auranofin This strategy was found to be very effective at enhancing oxidative stressmediated tumor cell killing and enhancing sensitivity to Erlotinib chemotherapy

Materials and Methods

Results

Findings

Discussion