Abstract
The human multidrug transporter MDR1 P-glycoprotein and the multidrug resistance proteins MRP1 and MRP2 transport a range of cytotoxic drugs, resulting in multidrug resistance in tumour cells. To overcome this form of drug resistance in patients, several inhibitors (reversal agents) of these transporters have been isolated. Using polarized cell lines stably expressing human MDR1, MRP1 or MRP2 cDNA, and 2008 ovarian carcinoma cells stably expressing MRP1 cDNA, we have investigated in this study the specificity of the reversal agents V-104 (a pipecolinate derivative), GF120918 (an acridone carboxamide derivative also known as GG918), and Pluronic L61 (a (poly)oxypropethylene and (poly)oxypropylene block copolymer). Transport experiments with cytotoxic drugs with polarized cell lines indicate that all three compounds efficiently inhibit MDR1 Pgp. Furthermore, V-104 partially inhibits daunorubicin transport by MRP1 but not vinblastine transport by MRP2. V-104 reverses etoposide resistance of 2008/MRP1 cells, whereas GF120918 does not reverse resistance due to MRP1. V-104 partially inhibits the export of the organic anion dinitrophenyl S -glutathione by MDCKII-MRP1 but not by MDCKII-MRP2 cells. Unexpectedly, export of the organic anion calcein by MDCKII-MRP1 and MDCKII-MRP2 cells is stimulated by Pluronic L61, probably because it relieves the block on entry of calcein AM into the cell by endogenous MDR1 Pgp. © 2000 Cancer Research Campaign
Highlights
Simultaneous resistance of tumour cells against a range of cytotoxic drugs is a serious problem in cancer chemotherapy
To further explore the suitability of transfected cell lines for the screening of reversal agents and to investigate the specificity of previously identified reversals, we describe in this report the effect of the inhibitors Pluronic L61, V-104, and GF120918 on MDR1 Pgp, MRP1- and MRP2mediated transport
At the highest concentration Pluronic L61 (0.1% (w/v)) tested, some inhibition (1.5-fold) of net vinblastine transport by MDCKII-MRP2 cells was observed, whereas transport mediated by MDR1 Pgp was completely blocked under these conditions (Figure 2)
Summary
Polarized canine (MDCKII) or porcine (LLC-PK1) kidney-cell lines stably expressing MDR1, MRP1, or MRP2 (cMOAT) have been described before (Schinkel et al, 1995; Evers et al, 1997; 1998; Bakos et al, 1998). The experiment was started (t = 0) by replacing the medium at either the apical or the basal side of the cell layer with 2 ml of complete medium containing 2 μM of drug (at 0.25 μCi ml–1), [14C]-labelled inulin (0.025μ Ci ml–1), and the amount of inhibitor indicated. Export of [14C]DNPGS was determined by incubating cells with 2 μM of [14C]CDNB (15 nCi ml–1) at room temperature as described previously (Evers et al, 1996; 1998), with the modification that the experiments were performed in Hanks Balanced Salt Solution (containing 1.3 mM CaCl2). If inhibitors were used in the experiment, cells were first incubated with HBSS containing inhibitor for 10 min before adding the substrate. Export of calcein was determined by incubating monolayers in HBSS containing calcein AM (1 μM) at room temperature. All transport experiments were performed in duplicate and repeated at least twice
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