Copper transporter 2 regulates the cellular accumulation and cytotoxicity of Cisplatin and Carboplatin.
OBJECTIVE
Copper transporter 2 (CTR2) is known to mediate the uptake of Cu(+1) by mammalian cells. Several other Cu transporters, including the influx transporter CTR1 and the two efflux transporters ATP7A and ATP7B, also regulate sensitivity to the platinum-containing drugs. We sought to determine the effect of CTR2 on influx, intracellular trafficking, and efflux of cisplatin and carboplatin.


METHODS
The role of CTR2 was examined by knocking down CTR2 expression in an isogenic pair of mouse embryo fibroblasts consisting of a CTR1(+/+) line and a CTR1(-/-) line in which both CTR1 alleles had been deleted. CTR2 levels were determined by quantitative reverse transcription-PCR and Western blot analysis. Cisplatin (DDP) was quantified by inductively coupled plasma mass spectrometry and (64)Cu and [(14)C]carboplatin (CBDCA) accumulation by gamma and scintillation counting.


RESULTS
Deletion of CTR1 reduced the uptake of Cu, DDP, and CBDCA and increased resistance to their cytotoxic effects by 2- to 3-fold. Knockdown of CTR2 increased uptake of Cu only in the CTR1(+/+) cells. In contrast, knockdown of CTR2 increased whole-cell DDP uptake and DNA platination in both CTR1(+/+) and CTR1(-/-) cells and proportionately enhanced cytotoxicity while producing no effect on vesicular accumulation or efflux. A significant correlation was found between CTR2 mRNA and protein levels and sensitivity to DDP in a panel of six ovarian carcinoma cell lines.


CONCLUSIONS
CTR2 is a major determinant of sensitivity to the cytotoxic effects of DDP and CBDCA. CTR2 functions by limiting drug accumulation, and its expression correlates with the sensitivity of human ovarian carcinoma cell lines to DDP.
Blair
Brian G
BG
Moores Cancer Center and Department of Medicine, University of California, San Diego, La Jolla, California.
Larson
Christopher A
CA
Safaei
Roohangiz
R
Howell
Stephen B
SB
D000818:Animals; D016190:Carboplatin / Q000493:pharmacokinetics* / Q000494:pharmacology / Q000627:therapeutic use; D002277:Carcinoma / Q000188:drug therapy / Q000235:genetics / Q000473:pathology; D027682:Cation Transport Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D002478:Cells, Cultured; D002945:Cisplatin / Q000493:pharmacokinetics* / Q000494:pharmacology / Q000627:therapeutic use; D003603:Cytotoxins / Q000493:pharmacokinetics* / Q000494:pharmacology / Q000627:therapeutic use; D018736:DNA Adducts / Q000187:drug effects / Q000378:metabolism; D019008:Drug Resistance, Neoplasm / Q000235:genetics; D005260:Female; D005347:Fibroblasts / Q000378:metabolism; D055785:Gene Knockdown Techniques; D006801:Humans; D051379:Mice; D018345:Mice, Knockout; D010051:Ovarian Neoplasms / Q000188:drug therapy / Q000235:genetics / Q000473:pathology; D000080083:SLC31 Proteins; D014018:Tissue Distribution / Q000235:genetics
D027682:Cation Transport Proteins; D003603:Cytotoxins; D018736:DNA Adducts; D000080083:SLC31 Proteins; C524078:SLC31A2 protein, human; D016190:Carboplatin; D002945:Cisplatin