Overexpression of glutathione S-transferase pi enhances the adduct formation of cisplatin with glutathione in human cancer cells.
In this paper, we provide direct evidence that glutathione S-transferase pi (GSTpi) 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 GSTpi 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 GSTpi. Purified GSTpi also catalyzed the formation of DDP-GSH in vitro, with an apparent Km of 0.23 mM for CDDP and an apparent Vmax of 4.9 nmol/min/mg protein. The increase in DDP-GSH produced by GSTpi was linear with incubation time up to 3 h and optimal of pH 7.4. A GSTpi transfectant cell line was constructed in HCT8 cells using a pcDNA3.1 (-)/Myc-His B with an expression vector containing cDNA for GSTpi. Transfection of GSTpi cDNA into HCT8 cells resulted in an increase in the expression of GSTpi by 1.4-fold in parallel with an augmentation of the formation of DDP-GSH. These results suggest that GSTpi plays a role in the formation of DDP-GSH and the acquisition of resistance to CDDP in cancer cells.
Goto
S
S
Department of Biochemistry and Molecular Biology in Disease, Atomic Bomb Disease Institute, Nagasaki University School of Medicine, Japan.
Iida
T
T
Cho
S
S
Oka
M
M
Kohno
S
S
Kondo
T
T
D002945:Cisplatin / Q000378:metabolism / Q000493:pharmacokinetics* / Q000494:pharmacology; D003110:Colonic Neoplasms / Q000188:drug therapy / Q000201:enzymology*; D019008:Drug Resistance, Neoplasm; D005260:Female; D005978:Glutathione / Q000378:metabolism*; D051549:Glutathione S-Transferase pi; D005982:Glutathione Transferase / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008658:Inactivation, Metabolic; D007527:Isoenzymes / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D010051:Ovarian Neoplasms / Q000188:drug therapy / Q000201:enzymology*; D014162:Transfection; D014407:Tumor Cells, Cultured
D007527:Isoenzymes; C496556:GSTP1 protein, human; D051549:Glutathione S-Transferase pi; D005982:Glutathione Transferase; D005978:Glutathione; D002945:Cisplatin