Involvement of multidrug resistance-associated protein 2 in in vivo cisplatin resistance of rat hepatoma AH66 cells.

We previously showed that the rat ascites hepatoma cell line AH66 is cisplatin resistant in vivo, but not in vitro. We further found that the expression of multidrug resistance-associated protein 2 (MRP2) in AH66 cells harvested from tumor-bearing rats was markedly decreased after incubation for 48 hours in medium containing 5% fetal bovine serum (5% FBS DMEM), and that the expression recovered when the cells were re-inoculated into rats. To examine whether the in vivo cisplatin resistance of AH66 cells is related to modulation or the expression of MRP2, we used AH66 cells transfected with pBK-CMV expression vector containing MRP2 antisense DNA. The expression of MRP2 alone among the MRP transporter family was markedly decreased in the transfected cells. The uptake of cisplatin into these cells was significantly higher than that in AH66 parent cells or control vector-transfected AH66 cells. The uptake of cisplatin in AH66 parent cells and the vector control cells was increased by treatment with sodium azide or probenecid whereas the uptake in the MRP2 antisense transfectant cells was not affected. When AH66 parent cells and control transfectant cells were cultured with cisplatin in medium containing 5% ascites fluid (5% ASF DMEM) for 48 hours, the cisplatin sensitivity significantly decreased. In contrast, the cisplatin sensitivity of MRP2 antisense transfectant cells did not change after culture in 5% ASF DMEM. These results show that the cisplatin resistance of AH66 cells is dependent upon the enhanced expression of MRP2.