Glutathione (GSH) is implicated in drug resistance mechanisms of several
cancers and is a key regulator of cell death pathways within cells. We studied
Ewing's sarcoma family of tumours (ESFT) cell lines and three mechanistically distinct
anticancer agents (
fenretinide,
doxorubicin, and
vincristine) to investigate whether the GSH
antioxidant system is involved in the reduced sensitivity to these chemotherapeutic agents in
hypoxia. Cell viability and death were assessed by the
trypan blue exclusion assay and
annexin V-PI staining, respectively.
Hypoxia significantly decreased the sensitivity of all ESFT cell lines to
fenretinide-induced death, whereas the effect of
doxorubicin or
vincristine was marginal and cell-line-specific. The response of the GSH
antioxidant system in ESFT cell lines to
hypoxia was variable and also cell-line-specific, although the level of GSH appeared to be most dependent on de novo biosynthesis rather than recycling. RNAi-mediated knockdown of key GSH regulatory
enzymes γ-
glutamylcysteine synthetase or
glutathione disulfide reductase partially reversed the
hypoxia-induced resistance to
fenretinide, and increasing GSH levels using
N-acetylcysteine augmented the
hypoxia-induced resistance in a cell line-specific manner. These observations are consistent with the conclusion that the role of the GSH
antioxidant system in modulating the sensitivity of ESFT cells to
fenretinide is heterogeneous depending on environment and cell type. This is likely to limit the value of targeting GSH as a therapeutic strategy to overcome
hypoxia-induced drug resistance in ESFT. Whether targeting the GSH
antioxidant system in conjunction with other
therapeutics may benefit some patients with ESFT remains to be seen.