Decreased DNA interstrand cross-linking and cytotoxicity induced in human brain tumor cells by 1,3-bis(2-chloroethyl)-1-nitrosourea after in vitro reaction with glutathione

Abstract

Although both direct and glutathione S-transferase (GST)-catalyzed interactions between many electrophiles and GSH generally result in inactivation of the former, there are several reports of compounds whose electrophilic, alkylating, and cytotoxic activities are potentiated by GSH. This study investigates the effects of direct in vitro interaction between GSH and BCNU at physiological pH (7.2) and temperature (37 degrees C) and how this affects the cytotoxic and DNA cross-linking activity of 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) in target human malignant brain tumor cells. The kinetics and dose-response relationship of this interaction were determined by measuring residual GSH and residual BCNU-cytotoxicity in aGSH/BCNU mixture over a 45-min period and at varying BCNU concentrations. The results demonstrate that reaction of BCNU with four times its molar concentration of GSH for 45 min significantly inactivates BCNU, as expressed by a 32% decrease in induction of cellular DNA cross-linking, a 21% increase in DNA synthesis, and a 15% increase in clonogenic survival of human brain tumor cells compared to incubates of BCNU alone. Equine liver (EL)-GST increased the inactivation of BCNU only slightly (insignificant at p = 0.05). These results suggest that, in contrast to agents such as the alkyl-N-nitro-N'-nitrosoguanidines which become more potent alkylators after reacting with GSH, the 2-chloroethylnitrosoureas (CENUs) undergo inactivation by GSH. We propose that such interactions between GSH and the CENUs may constitute an important aspect of CENU metabolism and provide a potential means by which brain tumor cells can circumvent CENU toxicity and exhibit resistance to this class of agents.