Cisplatin rapidly down-regulates its own influx transporter hCTR1 in cultured human ovarian carcinoma cells

Purpose: Cisplatin (DDP)-resistant cells commonly exhibit reduced drug accumulation. Previous studies have shown that the major copper (Cu) influx transporter CTR1 controls the uptake of DDP in yeast and mammalian cells. The goal of this study was to examine the effect of Cu and DDP on the level and subcellular localization of hCTR1 protein in human ovarian carcinoma cells. Experimental Design: Cultured human ovarian carcinoma A2780 cells were exposed to DDP and Cu, and the effect on hCTR1 was determined using Western blot analysis and confocal digital deconvolution microscopy. Results: Loss of hCTR1 was triggered by DDP exposure in a concentration and time-dependent manner. Exposure to 0.5 mumol/L DDP for 5 minutes reduced hCTR1 levels and exposure to DDP concentrations greater than or equal to2 mumol/L caused almost complete disappearance. The loss of hCTR1 was observed within I minute of the start of exposure to 2 mumol/L DDP. Treatment of cells with 100 mumol/L Cu for 5 minutes produced a smaller effect. Pretreatment of cells with 2 mumol/L DDP for 5 minutes resulted in a 50% decrease in Cu-64 uptake, demonstrating that the DDP-induced loss of hCTR1 detected by Western blot analysis and imaging was functionally significant. Conclusions: DDP down-regulated the amount of its major influx transporter in cultured human ovarian carcinoma cells in a concentration- and time-dependent manner. The effect was observed at DDP concentrations within the range found in the plasma of patients being treated with DDP, and it occurred very quickly relative to the half-life of the drug.