Transport of amphipathic anions by human multidrug resistance protein 3

• Published in: Cancer research (Chicago, Ill.) Vol. 60; no. 17; pp. 4779 - 4784
• Main Authors: HAO ZENG, GUOSHENG LIU, REA, Philip A, KRUH, Gary D
• Format: Journal Article
• Language: English
• Published: Philadelphia, PA American Association for Cancer Research 01-09-2000
• Subjects: Adenosine Triphosphate - metabolism; Anion Transport Proteins; Antineoplastic agents; ATP-Binding Cassette Transporters - metabolism; ATP-Binding Cassette, Sub-Family B, Member 1 - metabolism; Biological and medical sciences; Biological Transport, Active; Carbon Radioisotopes; Carrier Proteins - metabolism; Cell Line; Cell Membrane - metabolism; Estradiol - analogs & derivatives; Estradiol - pharmacokinetics; General aspects; Glutathione - analogs & derivatives; Glutathione - pharmacokinetics; Glycocholic Acid - pharmacokinetics; Humans; Kidney - metabolism; Kinetics; Leukotriene C4 - pharmacokinetics; Medical sciences; Methotrexate - pharmacokinetics; Multidrug Resistance-Associated Proteins; Osmosis; Pharmacology. Drug treatments; Substrate Specificity; Tritium; Antineoplastic agent; Amphipathic compound; Ion transport; Subgroup; Selectivity; Estradiol; In vitro; Glucuroconjugate; Protein; Organic anion; Antimetabolic; Membrane transport; Bile acid; Multiple resistance; Conjugated compound; Mechanism of action; Methotrexate; Carrier protein; Glutathione
• ISSN: 0008-5472; 1538-7445

The multidrug resistance-associated protein 1 (MRP1) and the canalicular multispecific organic anion transporter (cMOAT or MRP2) are ATP-binding cassette transporters that confer resistance to some anticancer drugs and efflux glutathione and glucuronate conjugates from the cell. The MRP subfamily of ABC transporters, however, contains at least four other members of which MRP3 (MOAT-D) bears the closest structural resemblance to MRP1. Although transfection studies have established that human MRP3 confers increased resistance to several anticancer agents, neither the substrate selectivity nor physiological functions of this transporter have been determined. Here we report the results of investigations of the in vitro transport properties of cloned human MRP3 using membrane vesicles prepared from MRP3-transfected HEK293 cells. It is shown that the expression of MRP3 is specifically associated with enhancement of the MgATP-dependent transport into membrane vesicles of the glucuronide estradiol 17-beta-D-glucuronide (E(2)17betaG), the glutathione conjugates 2,4-dinitrophenyl S-glutathione (DNP-SG) and leukotriene C4 (LTC4), the antimetabolite methotrexate, and the bile acid glycocholate. DNP-SG, LTC4, and E(2)17betaG are transported at moderate affinity and low capacity with Km and Vmax values of 5.7 +/- 1.7 microM and 3.8 +/- 0.1 pmol/mg/min, 5.3 +/- 2.6 microM and 20.2 +/- 5.9 pmol/mg/min, and 25.6 +/- 5.4 microM and 75.6 +/- 5.9 pmol/mg/min, respectively. Methotrexate and glycocholate are transported at low affinity and high capacity with Km and Vmax values of 776 +/- 319 microM and 288 +/- 54 pmol/mg/min and 248 +/- 113 microM and 183 +/- 34 pmol/mg/min, respectively. On the basis of these findings, the osmotic dependence of the transport measured and its inability to transport taurocholate, MRP3, like MRP1 and cMOAT, is concluded to be competent in the transport of glutathione S-conjugates, glucuronides, and methotrexate, albeit at low to moderate affinity. In contrast to MRP1, cMOAT, and all other characterized mammalian ABC transporters, however, MRP3 is active in the transport of the monoanionic human bile constituent glycocholate.