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Development and characterization of a recombinant madin-darby canine kidney cell line that expresses rat multidrug resistance-associated protein 1 (rMRP1)

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Abstract

Multidrug resistance-associated protein 1 (MRP1) is one of the major proteins shown to mediate efflux transport of a broad range of antitumor drugs, glucuronide conjugates, and glutathione, in addition to endogenous substrates. Significant differences in substrate selectivity were reported for murine and human MRP1. As preclinical drug disposition and pharmacokinetics studies are often conducted in rats, we have recently cloned the rat MRP1 (rMRP1) and demonstrated that rMRP1 expressed in transfected cells effluxes calcein, a commonly used fluorescence substrate for human MRP1. To further characterize the rat ortholog of MRP1, we isolated a cell line stably expressing recombinant rMRP1. These cells were tested for their ability to transport calcein and a range of chemotherapeutic drugs. Our results showed that cells expressing rMRP1 consistently efflux calcein at a rate 5-fold greater than control cells. The rMRP1 transfected cells, like their human ortholog, can confer drug resistance to vinca alkaloid (vinblastine and vincristine) and anthracycline drugs (daunorubcin and doxorubicin), and the resistance conferred by the MRP1 can be partially abolished by the MRP-specific inhibitors. The transepithelial permeability due to rMRP1 expression in differentiated Madin-Darby canine kidney cells (MDCK) cells was also investigated. The MRP1 transport activity is directional, as demonstrated by directional vinblastine transport. Collectively, our results demonstrate that the cellular expression of rMRP1, like its human ortholog, could confer resistance to anticancer drugs.

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References

  1. Higgins CF. ABC transporters: from microorganisms to man. Annu Rev Cell Biol. 1992;8:67–113.

    Article  CAS  PubMed  Google Scholar 

  2. Wijnholds J. Drug resistance caused by multidrug resistance-associated proteins. Novartis Found Symp. 2002;243:69–79; discussion 80–62, 180–185.

    Article  CAS  PubMed  Google Scholar 

  3. Goldstein LJ, Pastan I, Gottesman MM. Multidrug resistance in human cancer. Crit Rev Oncol Hematol. 1992;12:243–253.

    Article  CAS  PubMed  Google Scholar 

  4. Gottesman MM, Pastan I. Biochemistry of multidrug resistance mediated by the multidrug transporter. Annu Rev Biochem. 1993;62:385–427.

    Article  CAS  PubMed  Google Scholar 

  5. Cole SP, Bhardwaj G, Gerlach JH, et al. Overexpression of a transporter gene in a multidrug-resistant human lung cancer cell line. Science. 1992;258:1650–1654.

    Article  CAS  PubMed  Google Scholar 

  6. Kruh GD, Zeng H, Rea PA, et al. MRP subfamily transporters and resistance to anticancer agents. J Bioenerg Biomembr. 2001;33:493–501.

    Article  CAS  PubMed  Google Scholar 

  7. Shen H, Paul S, Breuninger LM, et al. Cellular and in vitro transport of glutathione conjugates by MRP. Biochemistry. 1996;35:5719–5725.

    Article  CAS  PubMed  Google Scholar 

  8. Rappa G, Lorico A, Flavell RA, Sartorelli AC. Evidence that the multidrug resistance protein (MRP) functions as a co-transporter of glutathione and natural product toxins. Cancer Res. 1997;57:5232–5237.

    CAS  PubMed  Google Scholar 

  9. Loe DW, Deeley RG, Cole SP. Characterization of vincristine transport by the M(r) 190,000 multidrug resistance protein (MRP): evidence for cotransport with reduced glutathione. Cancer Res. 1998;58:5130–5136.

    CAS  PubMed  Google Scholar 

  10. Paul S, Belinsky MG, Shen H, Kruh GD. Structure and in vitro substrate specificity of the murine multidrug resistance-associated protein. Biochemistry. 1996;35:13647–13655.

    Article  CAS  PubMed  Google Scholar 

  11. Stride BD, Grant CE, Loe DW, Hipfner DR, Cole SP, Deeley RG. Pharmacological characterization of the murine and human orthologs of multidrug-resistance protein in transfected human embryonic kidney cells. Mol Pharmacol. 1997;52:344–353.

    CAS  PubMed  Google Scholar 

  12. Zhang DW, Cole SP, Deeley RG. Identification of an amino acid residue in multidrug resistance protein 1 critical for conferring resistance to anthracyclines. J Biol Chem. 2001;276:13231–13239.

    Article  CAS  PubMed  Google Scholar 

  13. Lin ZP, Johnson DR, Finch RA, Belinsky MG, Kruh GD, Sartorelli AC. Comparative study of the importance of multidrug resistance-associated protein 1 and P-glycoprotein to drug sensitivity in immortalized mouse embryonic fibroblasts. Mol Cancer Ther. 2002;1:1105–1114.

    CAS  PubMed  Google Scholar 

  14. Cherrington NJ, Hartley DP, Li N, Johnson DR, Klaassen CD. Organ distribution of multidrug resistance proteins 1, 2, and 3 (Mrpl, 2, and 3) mRNA and hepatic induction of Mrp3 by constitutive androstane receptor activators in rats. J Pharmacol Exp Ther. 2002;300:97–104.

    Article  CAS  PubMed  Google Scholar 

  15. Regina A, Koman A, Piciotti M, El Hafny B, Center MS, Bergmann R, Couraud PO and Roux F. Mrp1 multidrug resistance-associated protein and P-glycoprotein expression in rat brain microvessel endothelial cells. J Neurochem. 1998;71:705–715.

    Article  CAS  PubMed  Google Scholar 

  16. Decleves X, Regina A, Laplanche JL, et al. Functional expression of P-glycoprotein and multidrug resistance-associated protein (Mrp1) in primary cultures of rat astrocytes. J Neurosci Res. 2000;60:594–601.

    Article  CAS  PubMed  Google Scholar 

  17. Hirrlinger J, Konig J, Dringen R. Expression of mRNAs of multidrug resistance proteins (Mrps) in cultured rat astrocytes, oligodendrocytes, microglial cells and neurones. J Neurochem. 2002;82:716–719.

    Article  CAS  PubMed  Google Scholar 

  18. Leazer TM, Klaassen CD. The presence of xenobiotic transporters in rat placenta. Drug Metab Dispos. 2003;31:153–167.

    Article  CAS  PubMed  Google Scholar 

  19. Soroka CJ, Lee JM, Azzaroli F, Boyer JL. Cellular localization and up-regulation of multidrug resistance-associated protein 3 in hepatocytes and cholangiocytes during obstructive cholestasis in rat liver. Hepatology. 2001;33:783–791.

    Article  CAS  PubMed  Google Scholar 

  20. Yang Z, Li CS, Shen DD, Ho RJ. Cloning and characterization of the rat multidrug resistance-associated protein 1. AAPS Pharm Sci. 2002;4(3): Article 15.

    Article  CAS  Google Scholar 

  21. Nunoya K, Grant CE, Zhang D, Cole SP, Deeley RG. Molecular cloning and pharmacological characterization of rat multidrug resistance protein 1 (mrp1). Drug Metab Dispos. 2003;31:1016–1026.

    Article  CAS  PubMed  Google Scholar 

  22. Makhey VD, Guo A, Norris DA, Hu P, Yan J, Sinko PJ. Characterization of the regional intestinal kinetics of drug efflux in rat and human intestine and in Caco-2 cells. Pharm Res. 1998;15:1160–1167.

    Article  CAS  PubMed  Google Scholar 

  23. Cole SP, Sparks KE, Fraser K, et al. Pharmacological characterization of multidrug resistant MRP-transfected human tumor cells. Cancer Res. 1994;54:5902–5910.

    CAS  PubMed  Google Scholar 

  24. Grant CE, Valdimarsson G, Hipfner DR, Almquist KC, Cole SP, Deeley RG. Overexpression of multidrug resistance-associated protein (MRP) increases resistance to natural product drugs. Cancer Res. 1994;54:357–361.

    CAS  PubMed  Google Scholar 

  25. Stride BD, Cole SP, Deeley RG. Localization of a substrate specificity domain in the multidrug resistance protein. J Biol Chem. 1999;274:22877–22883.

    Article  CAS  PubMed  Google Scholar 

  26. Hipfner DR, Gao M, Scheffer G, Scheper RJ, Deeley RG, Cole SP. Epitope mapping of monoclonal antibodies specific for the 190-kDa multidrug resistance protein (MRP). Br J Cancer. 1998;78:1134–1140.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Graham FL, Smiley J, Russell WC, Nairn R. Characteristics of a human cell line transformed by DNA from human adenovirus type 5. J Gen Virol. 1977;36:59–74.

    Article  CAS  PubMed  Google Scholar 

  28. Goh LB, Spears KJ, Yao D, et al. Endogenous drug transporters in vitro and in vivo models for the prediction of drug disposition in man. Biochem Pharmacol. 2002;64:1569–1578.

    Article  CAS  PubMed  Google Scholar 

  29. Lin JH. Drug-drug interaction mediated by inhibition and induction of P-glycoprotein. Adv Drug Deliv Rev. 2003;55:53–81.

    Article  CAS  PubMed  Google Scholar 

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Authors and Affiliations

  1. Departments of Pharmaceutics and Pharmacy, University of Washington, 98195-7610, Seattle, WA

    Ziping Yang, Micha Horn, Joanne Wang, Danny D Shen & Rodney JY Ho

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  1. Ziping Yang
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  2. Micha Horn
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  3. Joanne Wang
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  4. Danny D Shen
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  5. Rodney JY Ho
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Correspondence to Rodney JY Ho.

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Published: March 9, 2004

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Yang, Z., Horn, M., Wang, J. et al. Development and characterization of a recombinant madin-darby canine kidney cell line that expresses rat multidrug resistance-associated protein 1 (rMRP1). AAPS J 6, 8 (2004). https://doi.org/10.1208/ps060108

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  • DOI: https://doi.org/10.1208/ps060108

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