Overexpression of glutathione S-transferase π enhances the adduct formation of cisplatin with glutathione in human cancer cells

In this paper, we provide direct evidence that glutathione S-transferase pi (GST pi) detoxifies cisplatin (CDDP). We used human colonic cancer HCT8 cells sensitive and resistant to CDDP, the level of cisplatin-glutathione adduct (DDP-GSH) being higher in the resistant cells. There was an overexpression of GST pi mRNA in these CDDP-resistant cells. Incubation of the cells with CDDP resulted in the formation of DDP-GSH dependent on the CDDP concentration and the incubation time. The formation of DDP-GSH was abolished when the cells were pre-treated with ethacrynic acid or ketoprofen, inhibitors of GST pi. Purified GST pi also catalyzed the formation of DDP-GSH in vitro, with an apparent K-m of 0.23 mM for CDDP and an apparent V-max of 4.9 nmol/min/mg protein. The increase in DDP-GSH produced by GST pi was linear with incubation time up to 3 h and optimal of pH 7.4. A GST ii transfectant cell line was constructed in HCT8 cells using a pcDNA3.1 (-)/Myc-His B with an expression vector containing cDNA for GST pi. Transfection of GST pi cDNA into HCT8 cells resulted in an increase in the expression of GST pi by 1.4-fold in parallel with an augmentation of the formation of DDP-GSH. These results suggest that GST pi Flays a role in the formation of DDP-GSH and the acquisition of resistance to CDDP in cancer cells.