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. 2000 Jul 3;83(3):375–383. doi: 10.1054/bjoc.2000.1262

Vinblastine and sulfinpyrazone export by the multidrug resistance protein MRP2 is associated with glutathione export

R Evers 1,4, M de Haas 1, R Sparidans 2, J Beijnen 2, P R Wielinga 3, J Lankelma 3, P Borst 1
PMCID: PMC2374564  PMID: 10917554

Abstract

The multidrug resistance proteins MRP1 and MRP2 are members of the same subfamily of ATP-binding cassette transporters. Besides organic molecules conjugated to negatively charged ligands, these proteins also transport cytotoxic drugs for which no negatively charged conjugates are known to exist. In polarized MDCKII cells, MRP1 routes to the lateral plasma membrane, and MRP2 to the apical plasma membrane. In these cells MRP1 transports daunorubicin, and MRP2 vinblastine; both transporters export reduced glutathione (GSH) into the medium. We demonstrate that glutathione transport in MDCKII-MRP1 cells is inhibited by the inhibitors of organic anion transporters sulfinpyrazone, indomethacin, probenecid and benzbromarone. In MDCKII-MRP2 cells, GSH export is stimulated by low concentrations of sulfinpyrazone or indomethacin, whereas export is inhibited down to control levels at high concentrations. We find that unmodified sulfinpyrazone is a substrate for MRP2, also at concentrations where GSH export is inhibited. We also show that GSH export in MDCKII-MRP2 cells increases in the presence of vinblastine, and that the stochiometry between drug and GSH exported is between two and three. Our data indicate that transport of sulfinpyrazone and vinblastine is associated with GSH export. However, at high sulfinpyrazone concentrations this compound is transported without GSH. Models of MRP action are discussed that could explain these results. © 2000 Cancer Research Campaign

Keywords: multidrug resistance protein, polarized cell, glutathione, organic anion, drug transport, GS-X pump

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Selected References

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  1. Bakos E., Evers R., Sinkó E., Váradi A., Borst P., Sarkadi B. Interactions of the human multidrug resistance proteins MRP1 and MRP2 with organic anions. Mol Pharmacol. 2000 Apr;57(4):760–768. doi: 10.1124/mol.57.4.760. [DOI] [PubMed] [Google Scholar]
  2. Bakos E., Evers R., Szakács G., Tusnády G. E., Welker E., Szabó K., de Haas M., van Deemter L., Borst P., Váradi A. Functional multidrug resistance protein (MRP1) lacking the N-terminal transmembrane domain. J Biol Chem. 1998 Nov 27;273(48):32167–32175. doi: 10.1074/jbc.273.48.32167. [DOI] [PubMed] [Google Scholar]
  3. Büchler M., König J., Brom M., Kartenbeck J., Spring H., Horie T., Keppler D. cDNA cloning of the hepatocyte canalicular isoform of the multidrug resistance protein, cMrp, reveals a novel conjugate export pump deficient in hyperbilirubinemic mutant rats. J Biol Chem. 1996 Jun 21;271(25):15091–15098. doi: 10.1074/jbc.271.25.15091. [DOI] [PubMed] [Google Scholar]
  4. Chu X. Y., Kato Y., Sugiyama Y. Multiplicity of biliary excretion mechanisms for irinotecan, CPT-11, and its metabolites in rats. Cancer Res. 1997 May 15;57(10):1934–1938. [PubMed] [Google Scholar]
  5. Cole S. P., Deeley R. G. Multidrug resistance mediated by the ATP-binding cassette transporter protein MRP. Bioessays. 1998 Nov;20(11):931–940. doi: 10.1002/(SICI)1521-1878(199811)20:11<931::AID-BIES8>3.0.CO;2-J. [DOI] [PubMed] [Google Scholar]
  6. Cole S. P., Sparks K. E., Fraser K., Loe D. W., Grant C. E., Wilson G. M., Deeley R. G. Pharmacological characterization of multidrug resistant MRP-transfected human tumor cells. Cancer Res. 1994 Nov 15;54(22):5902–5910. [PubMed] [Google Scholar]
  7. Cui Y., König J., Buchholz J. K., Spring H., Leier I., Keppler D. Drug resistance and ATP-dependent conjugate transport mediated by the apical multidrug resistance protein, MRP2, permanently expressed in human and canine cells. Mol Pharmacol. 1999 May;55(5):929–937. [PubMed] [Google Scholar]
  8. Evers R., Kool M., van Deemter L., Janssen H., Calafat J., Oomen L. C., Paulusma C. C., Oude Elferink R. P., Baas F., Schinkel A. H. Drug export activity of the human canalicular multispecific organic anion transporter in polarized kidney MDCK cells expressing cMOAT (MRP2) cDNA. J Clin Invest. 1998 Apr 1;101(7):1310–1319. doi: 10.1172/JCI119886. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Evers R., Zaman G. J., van Deemter L., Jansen H., Calafat J., Oomen L. C., Oude Elferink R. P., Borst P., Schinkel A. H. Basolateral localization and export activity of the human multidrug resistance-associated protein in polarized pig kidney cells. J Clin Invest. 1996 Mar 1;97(5):1211–1218. doi: 10.1172/JCI118535. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Feller N., Broxterman H. J., Währer D. C., Pinedo H. M. ATP-dependent efflux of calcein by the multidrug resistance protein (MRP): no inhibition by intracellular glutathione depletion. FEBS Lett. 1995 Jul 17;368(2):385–388. doi: 10.1016/0014-5793(95)00677-2. [DOI] [PubMed] [Google Scholar]
  11. Gottesman M. M., Hrycyna C. A., Schoenlein P. V., Germann U. A., Pastan I. Genetic analysis of the multidrug transporter. Annu Rev Genet. 1995;29:607–649. doi: 10.1146/annurev.ge.29.120195.003135. [DOI] [PubMed] [Google Scholar]
  12. Heijn M., Hooijberg J. H., Scheffer G. L., Szabó G., Westerhoff H. V., Lankelma J. Anthracyclines modulate multidrug resistance protein (MRP) mediated organic anion transport. Biochim Biophys Acta. 1997 May 22;1326(1):12–22. doi: 10.1016/s0005-2736(97)00003-5. [DOI] [PubMed] [Google Scholar]
  13. Higgins C. F. ABC transporters: from microorganisms to man. Annu Rev Cell Biol. 1992;8:67–113. doi: 10.1146/annurev.cb.08.110192.000435. [DOI] [PubMed] [Google Scholar]
  14. Holló Z., Homolya L., Hegedüs T., Sarkadi B. Transport properties of the multidrug resistance-associated protein (MRP) in human tumour cells. FEBS Lett. 1996 Mar 25;383(1-2):99–104. doi: 10.1016/0014-5793(96)00237-2. [DOI] [PubMed] [Google Scholar]
  15. Horio M., Chin K. V., Currier S. J., Goldenberg S., Williams C., Pastan I., Gottesman M. M., Handler J. Transepithelial transport of drugs by the multidrug transporter in cultured Madin-Darby canine kidney cell epithelia. J Biol Chem. 1989 Sep 5;264(25):14880–14884. [PubMed] [Google Scholar]
  16. Ishikawa T. The ATP-dependent glutathione S-conjugate export pump. Trends Biochem Sci. 1992 Nov;17(11):463–468. doi: 10.1016/0968-0004(92)90489-v. [DOI] [PubMed] [Google Scholar]
  17. Ito K., Suzuki H., Hirohashi T., Kume K., Shimizu T., Sugiyama Y. Functional analysis of a canalicular multispecific organic anion transporter cloned from rat liver. J Biol Chem. 1998 Jan 16;273(3):1684–1688. doi: 10.1074/jbc.273.3.1684. [DOI] [PubMed] [Google Scholar]
  18. Jedlitschky G., Leier I., Buchholz U., Barnouin K., Kurz G., Keppler D. Transport of glutathione, glucuronate, and sulfate conjugates by the MRP gene-encoded conjugate export pump. Cancer Res. 1996 Mar 1;56(5):988–994. [PubMed] [Google Scholar]
  19. Jedlitschky G., Leier I., Buchholz U., Hummel-Eisenbeiss J., Burchell B., Keppler D. ATP-dependent transport of bilirubin glucuronides by the multidrug resistance protein MRP1 and its hepatocyte canalicular isoform MRP2. Biochem J. 1997 Oct 1;327(Pt 1):305–310. doi: 10.1042/bj3270305. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Leier I., Jedlitschky G., Buchholz U., Cole S. P., Deeley R. G., Keppler D. The MRP gene encodes an ATP-dependent export pump for leukotriene C4 and structurally related conjugates. J Biol Chem. 1994 Nov 11;269(45):27807–27810. [PubMed] [Google Scholar]
  21. Loe D. W., Almquist K. C., Cole S. P., Deeley R. G. ATP-dependent 17 beta-estradiol 17-(beta-D-glucuronide) transport by multidrug resistance protein (MRP). Inhibition by cholestatic steroids. J Biol Chem. 1996 Apr 19;271(16):9683–9689. doi: 10.1074/jbc.271.16.9683. [DOI] [PubMed] [Google Scholar]
  22. Loe D. W., Deeley R. G., Cole S. P. Characterization of vincristine transport by the M(r) 190,000 multidrug resistance protein (MRP): evidence for cotransport with reduced glutathione. Cancer Res. 1998 Nov 15;58(22):5130–5136. [PubMed] [Google Scholar]
  23. Masuda M., I'izuka Y., Yamazaki M., Nishigaki R., Kato Y., Ni'inuma K., Suzuki H., Sugiyama Y. Methotrexate is excreted into the bile by canalicular multispecific organic anion transporter in rats. Cancer Res. 1997 Aug 15;57(16):3506–3510. [PubMed] [Google Scholar]
  24. Müller M., Meijer C., Zaman G. J., Borst P., Scheper R. J., Mulder N. H., de Vries E. G., Jansen P. L. Overexpression of the gene encoding the multidrug resistance-associated protein results in increased ATP-dependent glutathione S-conjugate transport. Proc Natl Acad Sci U S A. 1994 Dec 20;91(26):13033–13037. doi: 10.1073/pnas.91.26.13033. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Oude Elferink R. P., Meijer D. K., Kuipers F., Jansen P. L., Groen A. K., Groothuis G. M. Hepatobiliary secretion of organic compounds; molecular mechanisms of membrane transport. Biochim Biophys Acta. 1995 Jul 17;1241(2):215–268. doi: 10.1016/0304-4157(95)00006-d. [DOI] [PubMed] [Google Scholar]
  26. Paulusma C. C., Bosma P. J., Zaman G. J., Bakker C. T., Otter M., Scheffer G. L., Scheper R. J., Borst P., Oude Elferink R. P. Congenital jaundice in rats with a mutation in a multidrug resistance-associated protein gene. Science. 1996 Feb 23;271(5252):1126–1128. doi: 10.1126/science.271.5252.1126. [DOI] [PubMed] [Google Scholar]
  27. Paulusma C. C., Kool M., Bosma P. J., Scheffer G. L., ter Borg F., Scheper R. J., Tytgat G. N., Borst P., Baas F., Oude Elferink R. P. A mutation in the human canalicular multispecific organic anion transporter gene causes the Dubin-Johnson syndrome. Hepatology. 1997 Jun;25(6):1539–1542. doi: 10.1002/hep.510250635. [DOI] [PubMed] [Google Scholar]
  28. Paulusma C. C., van Geer M. A., Evers R., Heijn M., Ottenhoff R., Borst P., Oude Elferink R. P. Canalicular multispecific organic anion transporter/multidrug resistance protein 2 mediates low-affinity transport of reduced glutathione. Biochem J. 1999 Mar 1;338(Pt 2):393–401. [PMC free article] [PubMed] [Google Scholar]
  29. Rappa G., Lorico A., Flavell R. A., Sartorelli A. C. Evidence that the multidrug resistance protein (MRP) functions as a co-transporter of glutathione and natural product toxins. Cancer Res. 1997 Dec 1;57(23):5232–5237. [PubMed] [Google Scholar]
  30. Taguchi Y., Yoshida A., Takada Y., Komano T., Ueda K. Anti-cancer drugs and glutathione stimulate vanadate-induced trapping of nucleotide in multidrug resistance-associated protein (MRP). FEBS Lett. 1997 Jan 13;401(1):11–14. doi: 10.1016/s0014-5793(96)01421-4. [DOI] [PubMed] [Google Scholar]
  31. Tew K. D. Glutathione-associated enzymes in anticancer drug resistance. Cancer Res. 1994 Aug 15;54(16):4313–4320. [PubMed] [Google Scholar]
  32. Tietze F. Enzymic method for quantitative determination of nanogram amounts of total and oxidized glutathione: applications to mammalian blood and other tissues. Anal Biochem. 1969 Mar;27(3):502–522. doi: 10.1016/0003-2697(69)90064-5. [DOI] [PubMed] [Google Scholar]
  33. Versantvoort C. H., Broxterman H. J., Bagrij T., Scheper R. J., Twentyman P. R. Regulation by glutathione of drug transport in multidrug-resistant human lung tumour cell lines overexpressing multidrug resistance-associated protein. Br J Cancer. 1995 Jul;72(1):82–89. doi: 10.1038/bjc.1995.281. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Wielinga P. R., de Waal E., Westerhoff H. V., Lankelma J. In vitro transepithelial drug transport by on-line measurement: cellular control of paracellular and transcellular transport. J Pharm Sci. 1999 Dec;88(12):1340–1347. doi: 10.1021/js980497z. [DOI] [PubMed] [Google Scholar]
  35. Zaman G. J., Flens M. J., van Leusden M. R., de Haas M., Mülder H. S., Lankelma J., Pinedo H. M., Scheper R. J., Baas F., Broxterman H. J. The human multidrug resistance-associated protein MRP is a plasma membrane drug-efflux pump. Proc Natl Acad Sci U S A. 1994 Sep 13;91(19):8822–8826. doi: 10.1073/pnas.91.19.8822. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Zaman G. J., Lankelma J., van Tellingen O., Beijnen J., Dekker H., Paulusma C., Oude Elferink R. P., Baas F., Borst P. Role of glutathione in the export of compounds from cells by the multidrug-resistance-associated protein. Proc Natl Acad Sci U S A. 1995 Aug 15;92(17):7690–7694. doi: 10.1073/pnas.92.17.7690. [DOI] [PMC free article] [PubMed] [Google Scholar]

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