Augmentation of Transport for Cisplatin-glutathione Adduct in Cisplatinresistant Cancer Cells

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We studied the outward transport of cisplatin (CDDP) -glutathione (GSH) adduct (DDP-GSH) in CDDP-resistant cancer cells. Incubating the cells in the presence of CDDP resulted in the formation of an adduct with GSH and subsequent transport outside the cells. We used human colonic cancer cells sensitive (HCT8) and resistant (HCT8DDP) to CDDP,and human ovarian cancer cells sensitive (A2780) and resistant (A2780DDP) to CDDP as materials.The concentration of intracellular GSH was higher in the resistant cells (118.7<plus-minus>5.9nmol/10^6 HCT8DDP vs 19.0<plus-minus>1.0nmol/10^6 HCT8 and 24.1<plus-minus>1.2nmol/10^6 A2780DDP vs 9.4<plus-minus>0.5nmol/10^6 A2780, respectively). The activity of the GSH synthesizing enzyme, gamma-glutamylcysteine synthetase (gamma-GCS) was higher in the CDDP-resistant cells (7.1<plus-minus>0.2milliunits/10^6 HCT8DDP vs 2.2<plus-minus>0.1milliunits/10^6 HCT8, and 2.9<plus-minus>0.1milliunits/10^6 A2780DDP vs 1.4<plus-minus>0.1milliunits/10^6 A2780 respectively) … More . Furthermore, immunological levels of gamma-GCS and the expression of gamma-GCS mRNA were higher in these CDDP-resistant cells than those in the control cells in accordance with the change in the concentration of GSH.DDP-GSH transport increased in the CDDP-resistant colonic cancer cells by 219% (324<plus-minus>12fmol/10^6 HCT8DDP cells/min vs 148<plus-minus>11fmol/10^6 HCT8 cells/min) and the CDDP-resistant ovarian cancer cells by 126% (127<plus-minus>7fmol/10^6 A2780DDP cells/min vs 101<plus-minus>8fmol/10^6 A2780 cells/min).DDP-GSH transport was also estimated using inside-out vesicles from these cells. The active transport of DDP-GSH was 243% of HCT8 in HCT8DDP and 121% of A2780 in A2780DDP.These data suggest that the acquisition of CDDP-resistance in cancer cells is due partly to the increase in the transport of DDP-GSH outside the cells as well as the increase in the concentration of GSH.Immunological estimation of the membrane proteins against human erythrocyte glutathione Sconjugate-stimulated Mg^<2+>-ATPase sera resulted in no apparent cross reactivity, suggesting that there are several transport systems for DDP-GSH. Less