Identification of the Hepatic Efflux Transporters of Organic Anions Using Double-Transfected Madin-Darby Canine Kidney II Cells Expressing Human Organic Anion-Transporting Polypeptide 1B1 (OATP1B1)/Multidrug Resistance-Associated Protein 2, OATP1B1/Multidrug Resistance 1, and OATP1B1/Breast Cancer R

Until recently, it was generally believed that the transport of various organic anions across the bile canalicular membrane was mainly mediated by multidrug resistance-associated protein 2 (MRP2/ABCC2). However, a number of new reports have shown that some organic anions are also substrates of multidrug resistance 1 (MDR1/ABCB1) and/or breast cancer resistance protein (BCRP/ABCG2), implying MDR1 and BCRP could also be involved in the biliary excretion of organic anions in humans. In the present study, we constructed new double-transfected Madin-Darby canine kidney II (MDCKII) cells expressing organic anion-transporting polypeptide 1B1 (OATP1B1)/MDR1 and OATP1B1/BCRP, and we investigated the transcellular transport of four kinds of organic anions, estradiol-17β-d-glucuronide (EG), estrone-3-sulfate (ES), pravastatin (PRA), and cerivastatin (CER), to identify which efflux transporters mediate the biliary excretion of compounds using double-transfected cells. We observed the vectorial transport of EG and ES in all the double transfectants. MRP2 showed the highest efflux clearance of EG among these efflux transporters, whereas BCRP-mediated clearance of ES was the highest in these double transfectants. In addition, two kinds of 3-hydroxy-3-methylglutaryl-CoA reductase inhibitors, CER and PRA, were also substrates of all these efflux transporters. The rank order of the efflux clearance of PRA mediated by each transporter was the same as that of EG, whereas the contribution of MDR1 to the efflux of CER was relatively greater than for PRA. This experimental system is very useful for identifying which transporters are involved in the biliary excretion of organic anions that cannot easily penetrate the plasma membrane.

[1]  V. Sasseville,et al.  A Novel Human Hepatic Organic Anion Transporting Polypeptide (OATP2) , 1999, The Journal of Biological Chemistry.

[2]  H. Yawo,et al.  Identification of a Novel Gene Family Encoding Human Liver-specific Organic Anion Transporter LST-1* , 1999, The Journal of Biological Chemistry.

[3]  Y. Sugiyama,et al.  Primary active transport of organic anions on bile canalicular membrane in humans. , 1999, The American journal of physiology.

[4]  L. Huang,et al.  Adenosine triphosphate-dependent transport of estradiol-17beta(beta-D-glucuronide) in membrane vesicles by MDR1 expressed in insect cells. , 1998, Hepatology.

[5]  Y. Sugiyama,et al.  Biliary excretion of pravastatin in rats: contribution of the excretion pathway mediated by canalicular multispecific organic anion transporter. , 1997, Drug metabolism and disposition: the biological fate of chemicals.

[6]  T. Friedberg,et al.  Endogenous drug transporters in in vitro and in vivo models for the prediction of drug disposition in man. , 2002, Biochemical pharmacology.

[7]  K. Maeda,et al.  Contribution of OATP2 (OATP1B1) and OATP8 (OATP1B3) to the Hepatic Uptake of Pitavastatin in Humans , 2004, Journal of Pharmacology and Experimental Therapeutics.

[8]  S. Thorgeirsson,et al.  Regulation of 2‐acetylaminofluorene‐and 3‐methylcholanthrene‐mediated induction of multidrug resistance and cytochrome P450IA gene family expression in primary hepatocyte cultures and rat liver , 1991, Molecular carcinogenesis.

[9]  S. Bates,et al.  Transport of methotrexate, methotrexate polyglutamates, and 17beta-estradiol 17-(beta-D-glucuronide) by ABCG2: effects of acquired mutations at R482 on methotrexate transport. , 2003, Cancer research.

[10]  M. Kool,et al.  Drug export activity of the human canalicular multispecific organic anion transporter in polarized kidney MDCK cells expressing cMOAT (MRP2) cDNA. , 1998, The Journal of clinical investigation.

[11]  Y. Sugiyama,et al.  Excretion of GSSG and Glutathione Conjugates Mediated by MRP1 and CM0AT/MRP2 , 1998, Seminars in liver disease.

[12]  M. Vijver,et al.  Subcellular localization and distribution of the breast cancer resistance protein transporter in normal human tissues. , 2001, Cancer research.

[13]  K. Bleasby,et al.  Rosuvastatin is a high affinity substrate of hepatic organic anion transporter OATP-C , 2001 .

[14]  D. Keppler,et al.  A novel human organic anion transporting polypeptide localized to the basolateral hepatocyte membrane. , 2000, American journal of physiology. Gastrointestinal and liver physiology.

[15]  D. Keppler,et al.  Vectorial transport by double-transfected cells expressing the human uptake transporter SLC21A8 and the apical export pump ABCC2. , 2001, Molecular pharmacology.

[16]  Y. Tanigawara.  Role of P-glycoprotein in drug disposition. , 2000, Therapeutic drug monitoring.

[17]  J. Tanaka,et al.  Construction of Epstein-Barr virus-based expression vector containing mini-oriP. , 1999, Biochemical and biophysical research communications.

[18]  Liyue Huang,et al.  Adenosine triphosphate–dependent transport of estradiol‐17β(β‐d‐glucuronide) in membrane vesicles by MDR1 expressed in insect cells , 1998 .

[19]  Y. Sugiyama,et al.  Species differences in the transport activity for organic anions across the bile canalicular membrane. , 1999, The Journal of pharmacology and experimental therapeutics.

[20]  T. Abe,et al.  Transcellular Transport of Organic Anions Across a Double-transfected Madin-Darby Canine Kidney II Cell Monolayer Expressing Both Human Organic Anion-transporting Polypeptide (OATP2/SLC21A6) and Multidrug Resistance-associated Protein 2 (MRP2/ABCC2)* , 2002, The Journal of Biological Chemistry.

[21]  M C Willingham,et al.  Cellular localization of the multidrug-resistance gene product P-glycoprotein in normal human tissues. , 1987, Proceedings of the National Academy of Sciences of the United States of America.

[22]  Yuichi Sugiyama,et al.  ABCG2 Transports Sulfated Conjugates of Steroids and Xenobiotics* , 2003, Journal of Biological Chemistry.

[23]  Y. Sugiyama,et al.  Functional Analysis of SNPs Variants of BCRP/ABCG2 , 2004, Pharmaceutical Research.

[24]  M. J. van de Vijver,et al.  Subcellular localization and distribution of the breast cancer resistance protein transporter in normal human tissues. , 2001, Cancer research.

[25]  Y. Sugiyama,et al.  Different biliary excretion systems for glucuronide and sulfate of a model compound; study using Eisai hyperbilirubinemic rats. , 1995, The Journal of pharmacology and experimental therapeutics.

[26]  T. Sudhop,et al.  Metabolism and drug interactions of 3-hydroxy-3-methylglutaryl coenzyme A-reductase inhibitors (statins) , 2001, European Journal of Clinical Pharmacology.

[27]  Y. Sugiyama,et al.  Multiple systems for the biliary excretion of organic anions in rats: liquiritigenin conjugates as model compounds. , 1994, The Journal of pharmacology and experimental therapeutics.

[28]  E. Wang,et al.  HMG-CoA Reductase Inhibitors (Statins) Characterized as Direct Inhibitors of P-Glycoprotein , 2001, Pharmaceutical Research.

[29]  Y. Sugiyama,et al.  Functional analysis of single nucleotide polymorphisms of hepatic organic anion transporter OATP1B1 (OATP-C). , 2004, Pharmacogenetics.

[30]  Y. Sugiyama,et al.  Biliary excretion of 17beta-estradiol 17beta-D-glucuronide is predominantly mediated by cMOAT/MRP2. , 2000, Pharmaceutical research.

[31]  O. H. Lowry,et al.  Protein measurement with the Folin phenol reagent. , 1951, The Journal of biological chemistry.

[32]  A. Guo,et al.  Delineating the contribution of secretory transporters in the efflux of etoposide using Madin-Darby canine kidney (MDCK) cells overexpressing P-glycoprotein (Pgp), multidrug resistance-associated protein (MRP1), and canalicular multispecific organic anion transporter (cMOAT). , 2002, Drug metabolism and disposition: the biological fate of chemicals.

[33]  Y. Sugiyama,et al.  Biliary Excretion of 17β-Estradiol 17β-d-Glucuronide Is Predominantly Mediated by cMOAT/MRP2 , 2000, Pharmaceutical Research.

[34]  G R Wilkinson,et al.  OATP and P-glycoprotein transporters mediate the cellular uptake and excretion of fexofenadine. , 1999, Drug metabolism and disposition: the biological fate of chemicals.

[35]  T. Abe,et al.  Human liver-specific organic anion transporter, LST-1, mediates uptake of pravastatin by human hepatocytes. , 2001, The Journal of pharmacology and experimental therapeutics.

[36]  G. Szakács,et al.  P glycoprotein and the mechanism of multidrug resistance. , 2002, Novartis Foundation symposium.

[37]  Yuichi Sugiyama,et al.  Inhibition of Transporter-Mediated Hepatic Uptake as a Mechanism for Drug-Drug Interaction between Cerivastatin and Cyclosporin A , 2003, Journal of Pharmacology and Experimental Therapeutics.

[38]  Y. Tanigawara,et al.  Simvastatin and lovastatin, but not pravastatin, interact with MDR1 , 2002, The Journal of pharmacy and pharmacology.

[39]  D. Keppler,et al.  Conjugate export pumps of the multidrug resistance protein (MRP) family: localization, substrate specificity, and MRP2-mediated drug resistance. , 1999, Biochimica et biophysica acta.