Uptake of Compounds That Selectively Kill Multidrug-Resistant Cells: The Copper Transporter SLC31A1 (CTR1) Increases Cellular Accumulation of the Thiosemicarbazone NSC73306

Acquired drug resistance in cancer continues to be a challenge in cancer therapy, in part due to overexpression of the drug efflux transporter P-glycoprotein (P-gp, MDRI, ABCB1). NSC73306 is a thiosemicarbazone compound that displays greater toxicity against cells expressing functional P-gp than against other cells. Here, we investigate the cellular uptake of NSC73306, and examine its interaction with P-gp and copper transporter 1 (CTRL SLC31A1). Overexpression of P-gp sensitizes LLC-PKI cells to NSC73306. Cisplatin (IC50 = 77 mu M), cyclosporin A (IC50 = 500 mu M), and verapamil (IC50 = 700 mu M) inhibited cellular accumulation of [H-3]NSC73306. Cellular hypertmdcity of NSC73306 to P-gp-expressing cells was inhibited by cisplatin in a dose-dependent manner. Cells transiently expressing the cisplatin uptake transporter CTRI (SLC3IA1) showed increased[H-3]NSC73306 accumulation. In contrast, CTR1 knockdown decreased [H-3]NSC73306 accumulation. The presence of NSC73306 reduced CTR1 levels, similar to the negative feedback of CTRI levels by copper or cisplatin. Surprisingly, although cisplatin is a substrate of CTR1, we found that CTR1 protein was overexpressed in high-level cisplatin-resistant KB-CP20 and BEL7404-CP20 cell lines. We confirmed that the CTR1 protein was functional, as uptake of NSC73306 was increased in KB-CP20 cells compared to their drug-sensitive parental cells, and downregulation of CTR1 in KB-CP20 cells reduced [3H]NSC73306 accumulation. These results suggest that NSC73306 is a transport substrate of CTR1.