Relevance of copper transporter 1 for cisplatin resistance in human ovarian carcinoma cells

Defects in intracellular accumulation of the antitumour drug cisplatin are a commonly observed feature in the cells selected for cisplatin resistance. Copper transporter 1 (CTR1) has been suggested to play an important role in drug uptake and resistance. Here, we describe a detailed investigation of the involvement of CTR1 in cisplatin uptake and its relevance for cisplatin resistance using a well characterised sensitive/cisplatin-resistant cell line pair: A2780 human ovarian carcinoma cell line and its cisplatin-resistant variant A2780cis. A2780cis cells showed decreased cisplatin accumulation and lower CTR1 expression compared to A2780 cells. Co-incubation with copper sulphate affected neither cisplatin accumulation (determined by nameless atomic absorption spectrometry) nor its cytotoxicity (determined using an MTT-assay, MTT=3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide). In both cell lines, CTR1 was localised near the nucleus as found using confocal fluorescence microscopy. The steady-state localisation of the protein in perinuclear region appears to involve its continuous endocytosis from cell surface. In contrast to copper, cisplatin exposure had no influence on the sub cellular localisation of CTR1. Co-localisation between CTR1 and a fluorescent cisplatin analogue labelled with carboxyfluorescein-diacetate could be observed in vesicular structures when continuous retrieval of the protein from cell membrane was inhibited. Our results strongly suggest that CTR1 mediates cisplatin uptake in the cell lines studied. Upon its transport across the plasma membrane by CTR1 the platinum drug is likely to be internalised along with the protein. Our findings imply that reduced CTR1 expression accounts for decreased cisplatin accumulation and represents one of the determinants of cisplatin resistance in A2780cis cell line. (c) 2012 Elsevier Inc. All rights reserved.