Pharmaceutical Research

, Volume 25, Issue 9, pp 1991–2001

Cyclooxygenase Inhibitors Down Regulate P-glycoprotein in Human Colorectal Caco-2 Cell Line

Research Paper



Elevated expression of the ABC transporters P-glycoprotein (P-gp), and breast cancer resistance protein (BCRP) seems to correlate with multidrug resistance of cancer cells. In this study we investigated the effect of COX inhibitors in modulating P-gp and BCRP expression and P-gp activity in Caco-2 cells.


mRNA and protein expression of MDR1 and BCRP were evaluated by real time PCR and western blot respectively. The activity of P-gp was measured by intracellular accumulation of rhodamine123 or 3H-Digoxin.


The chronic exposure of Caco-2 to indomethacin heptyl ester (indo HE) (0.4 μM) or nimesulide (10 μM) (selective COX-2 inhibitors) and naproxen (6 μM) (non selective inhibitor COX-1/COX-2) significantly decreased the expression and activity of P-gp. In contrast, the acute treatment by nimesulide and naproxen did not modify these parameters while indo HE treatment (48–72 h) caused a protein decrease and a functional inhibition of P-gp. Unexpectedly, the short-term treatment with naproxen induced an important increase of BCRP expression, but this induction was lost after long-term treatment. No modification of BCRP expression was observed after indo HE or nimesulide treatment.


Our observations suggest a possible down regulation of P-gp by COX inhibitors, which may enhance the accumulation of chemotherapy agents.


BCRP Caco-2 COX-2 inhibitor MDR1 P-gp 


  1. 1.
    V. Lingand, and L. H. Thompson. Reduced permeability in CHO cells as a mechanism of resistance to colchicine. J. Cell. Physiol. 83:103–116 (1974).CrossRefGoogle Scholar
  2. 2.
    F. Thiebaut, T. Tsuruo, H. Hamada, M. M. Gottesman, I. Pastan, and M. C. Willingham. Cellular localization of the multidrug-resistance gene product P-glycoprotein in normal human tissues. Proc. Natl. Acad. Sci. USA. 84:7735–7738 (1987).PubMedCrossRefGoogle Scholar
  3. 3.
    M. F. Fromm. P-glycoprotein: a defense mechanism limiting oral bioavailability and CNS accumulation of drugs. Int. J. Clin. Pharmacol. Ther. 38:69–74 (2000).PubMedGoogle Scholar
  4. 4.
    W. L. Smithand, and D. L. Dewitt. Prostaglandin endoperoxide H synthases-1 and -2. Adv. Immunol. 62:167–215 (1996).CrossRefGoogle Scholar
  5. 5.
    G. P. O'Neilland, and A. W. Ford-Hutchinson. Expression of mRNA for cyclooxygenase-1 and cyclooxygenase-2 in human tissues. FEBS Lett. 330:156–60 (1993).Google Scholar
  6. 6.
    C. C. Chan, S. Boyce, C. Brideau, A. W. Ford-Hutchinson, R. Gordon, D. Guay, R. G. Hill, C. S. Li, J. Mancini, M. Penneton et al. Pharmacology of a selective cyclooxygenase-2 inhibitor, L-745,337: a novel nonsteroidal anti-inflammatory agent with an ulcerogenic sparing effect in rat and nonhuman primate stomach. J. Pharmacol. Exp. Ther. 274:1531–1537 (1995).PubMedGoogle Scholar
  7. 7.
    T. Tanioka, Y. Nakatani, T. Kobayashi, M. Tsujimoto, S. Oh-ishi, M. Murakami, and I. Kudo. Regulation of cytosolic prostaglandin E2 synthase by 90-kDa heat shock protein. Biochem. Biophys. Res. Commun. 303:1018–1023 (2003).PubMedCrossRefGoogle Scholar
  8. 8.
    R. N. Dubois, S. B. Abramson, L. Crofford, R. A. Gupta, L. S. Simon, L. B. Van De Putte, and P. E. Lipsky. Cyclooxygenase in biology and disease. FASEB J. 12:1063–1073 (1998).PubMedGoogle Scholar
  9. 9.
    C. E. Trebino, J. L. Stock, C. P. Gibbons, B. M. Naiman, T. S. Wachtmann, J. P. Umland, K. Pandher, J. M. Lapointe, S. Saha, M. L. Roach, D. Carter, N. A. Thomas, B. A. Durtschi, J. D. McNeish, J. E. Hambor, P. J. Jakobsson, T. J. Carty, J. R. Perez, and L. P. Audoly. Impaired inflammatory and pain responses in mice lacking an inducible prostaglandin E synthase. Proc. Natl. Acad. Sci. USA. 100:9044–9049 (2003).PubMedCrossRefGoogle Scholar
  10. 10.
    C. Patrono, P. Patrignani, and L. A. Garcia Rodriguez. Cyclooxygenase-selective inhibition of prostanoid formation: transducing biochemical selectivity into clinical read-outs. J. Clin. Invest. 108:7–13 (2001).PubMedGoogle Scholar
  11. 11.
    J. A. Mitchelland, and T. D. Warner. Cyclo-oxygenase-2: pharmacology, physiology, biochemistry and relevance to NSAID therapy. Br. J. Pharmacol. 128:1121–32 (1999).CrossRefGoogle Scholar
  12. 12.
    I. I. Singer II, D. W. Kawka, S. Schloemann, T. Tessner, T. Riehl, and W. F. Stenson. Cyclooxygenase 2 is induced in colonic epithelial cells in inflammatory bowel disease. Gastroenterology 115:297–306 (1998).Google Scholar
  13. 13.
    C. E. Eberhart, R. J. Coffey, A. Radhika, F. M. Giardiello, S. Ferrenbach, and R. N. DuBois. Up-regulation of cyclooxygenase 2 gene expression in human colorectal adenomas and adenocarcinomas. Gastroenterology. 107:1183–8 (1994).PubMedGoogle Scholar
  14. 14.
    H. Sheng, J. Shao, J. D. Morrow, R. D. Beauchamp, and R. N. DuBois. Modulation of apoptosis and Bcl-2 expression by prostaglandin E2 in human colon cancer cells. Cancer Res. 58:362–366 (1998).PubMedGoogle Scholar
  15. 15.
    M. Tsujii, S. Kawano, S. Tsuji, H. Sawaoka, M. Hori, and R. N. DuBois. Cyclooxygenase regulates angiogenesis induced by colon cancer cells. Cell. 93:705–716 (1998).PubMedCrossRefGoogle Scholar
  16. 16.
    N. Arber, C. J. Eagle, J. Spicak, I. Racz, P. Dite, J. Hajer, M. Zavoral, M. J. Lechuga, P. Gerletti, J. Tang, R. B. Rosenstein, K. Macdonald, P. Bhadra, R. Fowler, J. Wittes, A. G. Zauber, S. D. Solomon, and B. Levin. Celecoxib for the prevention of colorectal adenomatous polyps. N. Engl. J. Med. 355:885–95 (2006).PubMedCrossRefGoogle Scholar
  17. 17.
    Y. Goldberg, Nassif, II, A. Pittas, L. L. Tsai, B. D. Dynlacht, B. Rigas, and S. J. Shiff. The anti-proliferative effect of sulindac and sulindac sulfide on HT-29 colon cancer cells: alterations in tumor suppressor and cell cycle-regulatory proteins. Oncogene 12:893–901 (1996).Google Scholar
  18. 18.
    J. Shao, T. Fujiwara, Y. Kadowaki, T. Fukazawa, T. Waku, T. Itoshima, T. Yamatsuji, M. Nishizaki, J. A. Roth, and N. Tanaka. Overexpression of the wild-type p53 gene inhibits NF-kappaB activity and synergizes with aspirin to induce apoptosis in human colon cancer cells. Oncogene. 19:726–36 (2000).PubMedCrossRefGoogle Scholar
  19. 19.
    S. Hashitani, M. Urade, N. Nishimura, T. Maeda, K. Takaoka, K. Noguchi, and K. Sakurai. Apoptosis induction and enhancement of cytotoxicity of anticancer drugs by celecoxib, a selective cyclooxygenase-2 inhibitor, in human head and neck carcinoma cell lines. Int. J. Oncol. 23:665–672 (2003).PubMedGoogle Scholar
  20. 20.
    J. L. Masferrer, K. M. Leahy, A. T. Koki, B. S. Zweifel, S. L. Settle, B. M. Woerner, D. A. Edwards, A. G. Flickinger, R. J. Moore, and K. Seibert. Antiangiogenic and antitumor activities of cyclooxygenase-2 inhibitors. Cancer Res. 60:1306–1311 (2000).PubMedGoogle Scholar
  21. 21.
    M. K. Jones, H. Wang, B. M. Peskar, E. Levin, R. M. Itani, I. J. Sarfeh, and A. S. Tarnawski. Inhibition of angiogenesis by nonsteroidal anti-inflammatory drugs: insight into mechanisms and implications for cancer growth and ulcer healing. Nat. Med. 5:1418–1423 (1999).PubMedCrossRefGoogle Scholar
  22. 22.
    V. A. Patel, M. J. Dunn, and A. Sorokin. Regulation of MDR-1 (P-glycoprotein) by cyclooxygenase-2. J. Biol. Chem. 277:38915–38920 (2002).PubMedCrossRefGoogle Scholar
  23. 23.
    M. C. Zatelli, A. Luchin, D. Piccin, F. Tagliati, A. Bottoni, C. Vignali, M. Bondanelli, and E. C. degli Uberti. Cyclooxygenase-2 inhibitors reverse chemoresistance phenotype in medullary thyroid carcinoma by a permeability glycoprotein-mediated mechanism. J. Clin. Endocrinol. Metab. 90:5754–5760 (2005).PubMedCrossRefGoogle Scholar
  24. 24.
    A. S. Kalgutkar, A. B. Marnett, B. C. Crews, R. P. Remmel, and L. J. Marnett. Ester and amide derivatives of the nonsteroidal antiinflammatory drug, indomethacin, as selective cyclooxygenase-2 inhibitors. J. Med. Chem. 43:2860–2870 (2000).PubMedCrossRefGoogle Scholar
  25. 25.
    T. Mosmann. Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays. J. Immunol. Methods. 65:55–63 (1983).PubMedCrossRefGoogle Scholar
  26. 26.
    M. Thamotharan, S. Z. Bawani, X. Zhou, and S. A. Adibi. Hormonal regulation of oligopeptide transporter pept-1 in a human intestinal cell line. Am. J. Physiol. 276:C821–C826 (1999).PubMedGoogle Scholar
  27. 27.
    S. Siissalo, L. Laitinen, M. Koljonen, K. S. Vellonen, H. Kortejarvi, A. Urtti, J. Hirvonen, and A. M. Kaukonen. Effect of cell differentiation and passage number on the expression of efflux proteins in wild type and vinblastine-induced Caco-2 cell lines. Eur. J. Pharm. Biopharm. 67:548–54 (2007).PubMedCrossRefGoogle Scholar
  28. 28.
    A. Pfrunder, H. Gutmann, C. Beglinger, and J. Drewe. Gene expression of CYP3A4, ABC-transporters (MDR1 and MRP1-MRP5) and hPXR in three different human colon carcinoma cell lines. J. Pharm. Pharmacol. 55:59–66 (2003).PubMedCrossRefGoogle Scholar
  29. 29.
    A. Geick, M. Eichelbaum, and O. Burk. Nuclear receptor response elements mediate induction of intestinal MDR1 by rifampin. J. Biol. Chem. 276:14581–14587 (2001).PubMedCrossRefGoogle Scholar
  30. 30.
    D. Ratnasinghe, P. J. Daschner, M. R. Anver, B. H. Kasprzak, P. R. Taylor, G. C. Yeh, and J. A. Tangrea. Cyclooxygenase-2, P-glycoprotein-170 and drug resistance; is chemoprevention against multidrug resistance possible? Anticancer Res. 21:2141–2147 (2001).PubMedGoogle Scholar
  31. 31.
    D. Kessel, W. T. Beck, D. Kukuruga, and V. Schulz. Characterization of multidrug resistance by fluorescent dyes. Cancer Res. 51:4665–4670 (1991).PubMedGoogle Scholar
  32. 32.
    M. Fontaine, W. F. Elmquist, and D. W. Miller. Use of rhodamine 123 to examine the functional activity of P-glycoprotein in primary cultured brain microvessel endothelial cell monolayers. Life Sci. 59:1521–1531 (1996).PubMedCrossRefGoogle Scholar
  33. 33.
    Y. Honjo, C. A. Hrycyna, Q. W. Yan, W. Y. Medina-Perez, R. W. Robey, A. van de Laar, T. Litman, M. Dean, and S. E. Bates. Acquired mutations in the MXR/BCRP/ABCP gene alter substrate specificity in MXR/BCRP/ABCP-overexpressing cells. Cancer Res. 61:6635–6639 (2001).PubMedGoogle Scholar
  34. 34.
    J. D. Allen, S. C. Jackson, and A. H. Schinkel. A mutation hot spot in the Bcrp1 (Abcg2) multidrug transporter in mouse cell lines selected for Doxorubicin resistance. Cancer Res. 62:2294–2299 (2002).PubMedGoogle Scholar
  35. 35.
    O. Alqawi, S. Bates, and E. Georges. Arginine482 to threonine mutation in the breast cancer resistance protein ABCG2 inhibits rhodamine 123 transport while increasing binding. Biochem. J. 382:711–716 (2004).PubMedCrossRefGoogle Scholar
  36. 36.
    R. W. Robey, Y. Honjo, A. van de Laar, K. Miyake, J. T. Regis, T. Litman, and S. E. Bates. A functional assay for detection of the mitoxantrone resistance protein, MXR (ABCG2). Biochim. Biophys. Acta. 1512:171–182 (2001).PubMedCrossRefGoogle Scholar
  37. 37.
    K. E. Pedersen, A. Dorph-Pedersen, S. Hvidt, N. A. Klitgaard, and K. K. Pedersen. The long-term effect of verapamil on plasma digoxin concentration and renal digoxin clearance in healthy subjects. Eur. J. Clin. Pharmacol. 22:123–127 (1982).PubMedCrossRefGoogle Scholar
  38. 38.
    U. Puhlmann, C. Ziemann, G. Ruedell, H. Vorwerk, D. Schaefer, C. Langebrake, P. Schuermann, U. Creutzig, and D. Reinhardt. Impact of the cyclooxygenase system on doxorubicin-induced functional multidrug resistance 1 overexpression and doxorubicin sensitivity in acute myeloid leukemic HL-60 cells. J. Pharmacol. Exp. Ther. 312:346–354 (2005).PubMedCrossRefGoogle Scholar
  39. 39.
    I. Tegeder, J. Pfeilschifter, and G. Geisslinger. Cyclooxygenase-independent actions of cyclooxygenase inhibitors. FASEB J. 15:2057–2072 (2001).PubMedCrossRefGoogle Scholar
  40. 40.
    M. Bentires-Alj, V. Barbu, M. Fillet, A. Chariot, B. Relic, N. Jacobs, J. Gielen, M. P. Merville, and V. Bours. NF-kappaB transcription factor induces drug resistance through MDR1 expression in cancer cells. Oncogene. 22:90–97 (2003).PubMedCrossRefGoogle Scholar
  41. 41.
    M. L. Smith, G. Hawcroft, and M. A. Hull. The effect of non-steroidal anti-inflammatory drugs on human colorectal cancer cells: evidence of different mechanisms of action. Eur. J. Cancer. 36:664–74 (2000).PubMedCrossRefGoogle Scholar
  42. 42.
    M. Goto, S. Masuda, H. Saito, and K. Inui. Decreased expression of P-glycoprotein during differentiation in the human intestinal cell line Caco-2. Biochem. Pharmacol. 66:163–170 (2003).PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2008

Authors and Affiliations

  • Afraa Zrieki
    • 1
    • 2
  • Robert Farinotti
    • 1
    • 2
    • 3
  • Marion Buyse
    • 1
    • 2
  1. 1.Université Paris-sud XI, Faculté de PharmacieLaboratoire de Pharmacie CliniqueChâtenay-MalabryFrance
  2. 2.IFR 141Université Paris-sud XIChâtenay-MalabryFrance
  3. 3.Hôpital Pitié SalpetrièreService de Pharmacie, Assistance Publique-Hopitaux de ParisParisFrance

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