Enhanced γ-glutamylcysteine synthetase activity decreases drug-induced oxidative stress levels and cytotoxicity

Multidrug resistance of cancer cells can be intrinsic or acquired and occurs due to various reasons, including increased repair of genotoxic damage, an enhanced ability to remove/detoxify chemical agents, or reactive oxygen species (ROS), and repression of apoptosis. Human A2780/100 ovarian carcinoma cells exhibit resistance to DNA cross-linking agents, chlorambucil (Cbl), cisplatin (Cpl), melphalan (Mel), and ionizing radiation (IR) compared to the parental cell line, A2780. In the present study, we show that when A2780/100 and A2780 cells were treated with Cbl, GSH was extruded via methionine or cystathionine-inhibitable transporters of intact plasma membrane. GSH loss was followed by a rapid increase in ROS levels. The resistant, but not drug-sensitive cells normalized the intracellular GSH concentration along with ROS levels within 4-6 h after Cbl addition, and survived drug treatment. Normalization of GSH and ROS levels in A2780/100 cells correlated well with elevated γ-glutamylcysteine synthetase (γ-GCS) activity (10 ± 1.8-fold over A2780 cells). Ectopic overexpression of the γ-GCS heavy subunit in drug-sensitive cells nearly restored GSH and ROS to pre-treatment levels consequently increased cellular resistance to genotoxic agents (Cbl, Cpl, and IR), while overexpression of γ-GCS light subunit had no such effects. Thus, in our model system, drug-resistant cells have the inherent ability to maintain increased γ-GCS activity, reestablish physiological GSH, and cellular redox state and maintain increased cellular resistance to DNA cross-linking agents and IR.