[go: up one dir, main page]
Skip to main content

Advertisement

Springer Nature Link
Log in

Constitutive Overexpression of P-glycoprotein, Rather than Breast Cancer Resistance Protein or Organic Cation Transporter 1, Contributes to Acquisition of Imatinib-Resistance in K562 Cells

  • Research Paper
  • Published:
Pharmaceutical Research Aims and scope Submit manuscript

    We’re sorry, something doesn't seem to be working properly.

    Please try refreshing the page. If that doesn't work, please contact support so we can address the problem.

Abstract

Purpose

The purpose of this study was to investigate the contribution of drug transporters in acquired imatinib-resistance. Specifically, we focused on the efflux transporters, P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP), and an influx transporter, organic cation transporter 1 (OCT1).

Materials and methods

We established imatinib-resistant K562 cells (K562/IM). Real-time PCR or Western blot analyses were performed to examine the mRNA or protein levels. Alamar blue method was used in the cytotoxicity assay. The transport activities and intracellular imatinib levels were measured by flow cytometry and HPLC, respectively.

Results

K562/IM displayed a 47-fold increase in resistance to imatinib over the parent K562 cells. Both P-gp and BCRP were overexpressed in K562/IM relative to K562. Furthermore, the intracellular imatinib level was markedly reduced in K562/IM. Interestingly, cyclosporin A, a P-gp inhibitor, but not fumitremorgin C, a BCRP inhibitor, restored both imatinib-sensitivity and the intracellular imatinib level. By contrast, no significant difference in the expression and function of OCT1 was observed between K562/IM and K562.

Conclusions

P-gp, rather than BCRP or OCT1, is partially responsible for the development of imatinib-resistance due to constitutive and functional overexpression, leading to reduced intracellular accumulation of imatinib in K562/IM.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Subscribe and save

Springer+ Basic
$34.99 /Month
  • Get 10 units per month
  • Download Article/Chapter or eBook
  • 1 Unit = 1 Article or 1 Chapter
  • Cancel anytime
Subscribe now

Buy Now

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3

Similar content being viewed by others

Abbreviations

BCRP:

breast cancer resistance protein

IM:

imatinib

OCT1:

organic cation transporter 1

P-gp:

P-glycoprotein

References

  1. M. Kalidas, H. Kantarjian, and M. Talpaz. Chronic myelogenous leukemia. JAMA 286:895–898 (2001).

    Article  PubMed  CAS  Google Scholar 

  2. R. Capdeville, E. Buchdunger, J. Zimmermann, and A. Matter. Glivec (STI571, imatinib), a rationally developed, targeted anticancer drug. Nat. Rev. Drug Discov. 1:493–502 (2002).

    Article  PubMed  CAS  Google Scholar 

  3. B. J. Druker, C. L. Sawyers, H. Kantarjian, D. J. Resta, S. F. Reese, J. M. Ford, R. Capdeville, and M. Talpaz. Activity of a specific inhibitor of the BCR-ABL tyrosine kinase in the blast crisis of chronic myeloid leukemia and acute lymphoblastic leukemia with the Philadelphia chromosome. N. Engl. J. Med. 344:1038–1042 (2001).

    Article  PubMed  CAS  Google Scholar 

  4. C. L. Sawyers, A. Hochhaus, E. Feldman, J. M. Goldman, C. B. Miller, O. G. Ottmann, C. A. Schiffer, M. Talpaz, F. Guilhot, M. W. N. Deininger, T. Fischer, S. G. O’Brien, R. M. Stone, C. B. Gambacorti-Passerini, N. H. Russell, J. J. Reiffers, T. C. Shea, B. Chapuis, S. Coutre, S. Tura, E. Morra, R. A. Larson, A. Saven, C. Peschel, A. Gratwohl, F. Mandelli, M. Ben-Am, I. Gathmann, R. Capdeville, R. L. Paquette, and B. J. Druker. Imatinib induces hematologic and cytogenetic responses in patients with chronic myelogenous leukemia in myeloid blast crisis: results of a phase II study. Blood 99:3530–3539 (2002).

    Article  PubMed  CAS  Google Scholar 

  5. O. G. Ottmann, B. J. Druker, C. L. Sawyers, J. M. Goldman, J. Reiffers, R. T. Silver, S. Tura, T. Fischer, M. W. Deininger, C. A. Schiffer, M. Baccarani, A. Gratwohl, A. Hochhaus, D. Hoelzer, S. Fernandes-Reese, I. Gathmann, R. Capdeville, and S. G. O’Brien. A phase 2 study of imatinib in patients with relapsed or refractory Philadelphia chromosome-positive acute lymphoid leukemias. Blood 100:1965–1971 (2002).

    Article  PubMed  CAS  Google Scholar 

  6. M. E. Gorre, M. Mohammed, K. Ellwood, N. Hsu, R. Paquette, P. N. Rao, and C. L. Sawyers. Clinical resistance to STI-571 cancer therapy caused by BCR-ABL gene mutation or amplification. Science 293:876–880 (2001).

    Article  PubMed  CAS  Google Scholar 

  7. M. E. O’Dwyer, M. J. Mauro, G. Kurilik, M. Mori, S. Balleisen, S. Olson, E. Magenis, R. Capdeville, and B. J. Druker. The impact of clonal evolution on response to imatinib mesylate (STI571) in accelerated phase CML. Blood 100:1628–1633 (2002).

    Article  PubMed  CAS  Google Scholar 

  8. J. E. Cortes, M. Talpaz, F. Giles, S. O’Brien, M. B. Rios, J. Shan, G. Garcia-Manero, S. Faderl, D. A. Thomas, W. Wierda, A. Ferrajoli, S. Jeha, and H. M. Kantarjian. Prognostic significance of cytogenetic clonal evolution in patients with chronic myelogenous leukemia on imatinib mesylate therapy. Blood 101:3794–3800 (2003).

    Article  PubMed  CAS  Google Scholar 

  9. S. Branford, Z. Rudzki, S. Walsh, A. Grigg, C. Arthur, K. Taylor, R. Herrmann, K. P. Lynch, and T. P. Hughes. High frequency of point mutations clustered within the adenosine triphosphate-binding region of BCR/ABL in patients with chronic myeloid leukemia or Ph-positive acute lymphoblastic leukemia who develop imatinib (STI571) resistance. Blood 99:3472–3475 (2002).

    Article  PubMed  CAS  Google Scholar 

  10. C. Roche-Lestienne, V. Soenen-Cornu, N. Grardel-Duflos, J.-L. Lai, N. Philippe, T. Facon, P. Fenaux, and C. Preudhomme. Several types of mutations of the Abl gene can be found in chronic myeloid leukemia patients resistant to STI571, and they can pre-exist to the onset of treatment. Blood 100:1014–1018 (2002).

    Article  PubMed  CAS  Google Scholar 

  11. N. C. Wolff, D. R. Veach, W. P. Tong, W. G. Bornmann, B. Clarkson, and R. L. Ilaria, Jr. PD166326, a novel tyrosine kinase inhibitor, has greater antileukemic activity than imatinib mesylate in a murine model of chronic myeloid leukemia. Blood 105:3995–4003 (2005).

    Article  PubMed  CAS  Google Scholar 

  12. H. Kantarjian, F. Giles, L. Wunderle, K. Bhalla, S. O’Brien, B. Wassmann, C. Tanaka, P. Manley, P. Rae, W. Mietlowski, K. Bochinski, A. Hochhaus, J. D. Griffin, D. Hoelzer, M. Albitar, M. Dugan, J. Cortes, L. Alland, and O. G. Ottmann. Nilotinib in imatinib-resistant CML and Philadelphia chromosome-positive ALL. N. Engl. J. Med. 354:2542–2551 (2006).

    Article  PubMed  Google Scholar 

  13. M. Talpaz, N. P. Shah, H. Kantarjian, N. Donato, J. Nicoll, R. Paquette, J. Cortes, S. O’Brien, C. Nicaise, E. Bleickardt, M. A. Blackwood-Chirchir, V. Iyer, T.-T. Chen, F. Huang, A. P. Decillis, and C. L. Sawyers. Dasatinib in imatinib-resistant Philadelphia chromosome-positive leukemias. N. Engl. J. Med. 354:2531–2541 (2006).

    Article  PubMed  CAS  Google Scholar 

  14. F. X. Mahon, M. W. Deininger, B. Schultheis, J. Chabrol, J. Reiffers, J. M. Goldman, and J. V. Melo. Selection and characterization of BCR-ABL positive cell lines with differential sensitivity to the tyrosine kinase inhibitor STI571: diverse mechanisms of resistance. Blood 96:1070–1079 (2000).

    PubMed  CAS  Google Scholar 

  15. N. J. Donato, J. Y. Wu, J. Stapley, G. Gallick, H. Lin, R. Arlinghaus, and M. Talpaz. BCR-ABL independence and LYN kinase overexpression in chronic myelogenous leukemia cells selected for resistance to STI571. Blood 101:690–698 (2003).

    Article  PubMed  CAS  Google Scholar 

  16. Y. Dai, M. Rahmani, S. J. Corey, P. Dent, and S. Grant. A Bcr/Abl-independent, Lyn-dependent form of imatinib mesylate (STI-571) resistance is associated with altered expression of Bcl-2. J. Biol. Chem. 279:34227–34239 (2004).

    Article  PubMed  CAS  Google Scholar 

  17. F.-X. Mahon, F. Belloc, V. Lagarde, C. Chollet, F. Moreau-Gaudry, J. Reiffers, J. M. Goldman, and J. V. Melo. MDR1 gene overexpression confers resistance to imatinib mesylate in leukemia cell line models. Blood 101:2368–2373 (2003).

    Article  PubMed  CAS  Google Scholar 

  18. A. Hamada, H. Miyano, H. Watanabe, and H. Saito. Interaction of imatinib mesilate with human P-glycoprotein. J. Pharmacol. Exp. Ther. 307:824–828 (2003).

    Article  PubMed  CAS  Google Scholar 

  19. N. Widmer, S. Colombo, T. Buclin, and L. A. Decosterd. Functional consequence of MDR1 expression on imatinib intracellular concentrations. Blood 102:1142

    Article  PubMed  CAS  Google Scholar 

  20. T. Illmer, M. Schaich, U. Platzbecker, J. Freiberg-Richter, U. Oelschlagel, M. von Bonin, S. Pursche, T. Bergemann, G. Ehninger, and E. Schleyer. P-glycoprotein-mediated drug efflux is a resistance mechanism of chronic myelogenous leukemia cells to treatment with imatinib mesylate. Leukemia 18:401–408 (2004).

    Article  PubMed  CAS  Google Scholar 

  21. A. Radujkovic, M. Schad, J. Topaly, M. R. Veldwijk, S. Laufs, B. S. Schultheis, A. Jauch, J. V. Melo, S. Fruehauf, and W. J. Zeller. Synergistic activity of imatinib and 17-AAG in imatinib-resistant CML cells overexpressing BCR-ABL—inhibition of P-glycoprotein function by 17-AAG. Leukemia 19:1198–1206 (2005).

    Article  PubMed  CAS  Google Scholar 

  22. M. Mukai, X.-F. Che, T. Furukawa, T. Sumizawa, S. Aoki, X.-Q. Ren, M. Haraguchi, Y. Sugimoto, M. Kobayashi, H. Takamatsu, and S.-i. Akiyama. Reversal of the resistance to STI571 in human chronic myelogenous leukemia K562 cells. Cancer Sci. 94:557–563 (2003).

    Article  PubMed  CAS  Google Scholar 

  23. F. J. Giles, H. Kantarjian, J. Cortes, D. Thomas, M. Talpaz, T. Manshouri, and M. Albitar. Multidrug resistance protein expression in chronic myeloid leukemia: Associations and significance. Cancer 86:805–813 (1999).

    Article  PubMed  CAS  Google Scholar 

  24. H. Burger, H. van Tol, A. W. Boersma, M. Brok, E. A. Wiemer, G. Stoter, and K. Nooter. Imatinib mesylate (STI571) is a substrate for the breast cancer resistance protein (BCRP)/ABCG2 drug pump. Blood 104:2940–2942 (2004).

    Article  PubMed  CAS  Google Scholar 

  25. P. Breedveld, D. Pluim, G. Cipriani, P. Wielinga, O. van Tellingen, A. H. Schinkel, and J. H. M. Schellens. The Effect of Bcrp1 (Abcg2) on the in vivo pharmacokinetics and brain penetration of imatinib mesylate (Gleevec): implications for the use of breast cancer resistance protein and P-glycoprotein inhibitors to enable the brain penetration of imatinib in patients. Cancer Res. 65:2577–2582 (2005).

    Article  PubMed  CAS  Google Scholar 

  26. P. J. Houghton, G. S. Germain, F. C. Harwood, J. D. Schuetz, C. F. Stewart, E. Buchdunger, and P. Traxler. Imatinib mesylate is a potent inhibitor of the ABCG2 (BCRP) transporter and reverses resistance to topotecan and SN-38 in vitro. Cancer Res. 64:2333–2337 (2004).

    Article  PubMed  CAS  Google Scholar 

  27. N. E. Jordanides, H. G. Jorgensen, T. L. Holyoake, and J. C. Mountford. Functional ABCG2 is overexpressed on primary CML CD34+ cells and is inhibited by imatinib mesylate. Blood 108:1370–1373 (2006).

    Article  PubMed  CAS  Google Scholar 

  28. J. Thomas, L. Wang, R. E. Clark, and M. Pirmohamed. Active transport of imatinib into and out of cells: implications for drug resistance. Blood 104:3739–3745 (2004).

    Article  PubMed  CAS  Google Scholar 

  29. L. C. Crossman, B. J. Druker, M. W. N. Deininger, M. Pirmohamed, L. Wang, and R. E. Clark. hOCT 1 and resistance to imatinib. Blood 106:1133–1134 (2005).

    Article  PubMed  CAS  Google Scholar 

  30. D. L. White, V. A. Saunders, P. Dang, J. Engler, A. C. Zannettino, A. C. Cambareri, S. R. Quinn, P. W. Manley, and T. P. Hughes. OCT-1-mediated influx is a key determinant of the intracellular uptake of imatinib but not nilotinib (AMN107): reduced OCT-1 activity is the cause of low in vitro sensitivity to imatinib. Blood 108:697–704 (2006).

    Article  PubMed  CAS  Google Scholar 

  31. H. M. Kantarjian, M. Talpaz, S. O’Brien, F. Giles, G. Garcia-Manero, S. Faderl, D. Thomas, J. Shan, M. B. Rios, and J. Cortes. Dose escalation of imatinib mesylate can overcome resistance to standard-dose therapy in patients with chronic myelogenous leukemia. Blood 101:473–475 (2003).

    Article  PubMed  CAS  Google Scholar 

  32. J. O’Brien, I. Wilson, T. Orton, and F. Pognan. Investigation of the Alamar Blue (resazurin) fluorescent dye for the assessment of mammalian cell cytotoxicity. Eur. J. Biochem. 267:5421–5426 (2000).

    Article  PubMed  CAS  Google Scholar 

  33. G. W. Dewald, W. A. Wyatt, A. L. Juneau, R. O. Carlson, A. R. Zinsmeister, S. M. Jalal, J. L. Spurbeck, and R. T. Silver. Highly sensitive fluorescence in situ hybridization method to detect double BCR/ABL fusion and monitor response to therapy in chronic myeloid leukemia. Blood 91:3357–3365 (1998).

    PubMed  CAS  Google Scholar 

  34. R. W. Robey, K. Steadman, O. Polgar, K. Morisaki, M. Blayney, P. Mistry, and S. E. Bates. Pheophorbide a is a specific probe for ABCG2 function and inhibition. Cancer Res. 64:1242–1246 (2004).

    Article  PubMed  CAS  Google Scholar 

  35. C. Özvegy-Laczka, T. Hegedüs, G. Varady, O. Ujhelly, J. D. Schuetz, A. Varadi, G. Keri, L. Örfi, K. Nemet, and B. Sarkadi. High-affinity interaction of tyrosine kinase inhibitors with the ABCG2 multidrug transporter. Mol. Pharmacol. 65:1485–1495 (2004).

    Article  PubMed  Google Scholar 

  36. H. Burger, H. van Tol, M. Brok, E. A. Wiemer, E. A. de Bruijn, G. Guetens, G. de Boeck, A. Sparreboom, J. Verweij, and K. Nooter. Chronic imatinib mesylate exposure leads to reduced intracellular drug accumulation by induction of the ABCG2 (BCRP) and ABCB1 (MDR1) drug transport pumps. Cancer Biol. Ther. 4:747–752 (2005).

    Article  PubMed  CAS  Google Scholar 

  37. T. Nakanishi, K. Shiozawa, B. A. Hassel, and D. D. Ross. Complex interaction of BCRP/ABCG2 and imatinib in BCR-ABL-expressing cells: BCRP-mediated resistance to imatinib is attenuated by imatinib-induced reduction of BCRP expression. Blood 108:678–684 (2006).

    Article  PubMed  CAS  Google Scholar 

  38. E. R. Gardner, H. Burger, R. H. van Schaik, A. T. van Oosterom, E. A. de Bruijn, G. Guetens, H. Prenen, F. A. de Jong, S. D. Baker, S. E. Bates, W. D. Figg, J. Verweij, A. Sparreboom, and K. Nooter. Association of enzyme and transporter genotypes with the pharmacokinetics of imatinib. Clin. Pharmacol. Ther. 80:192–201 (2006).

    Article  PubMed  CAS  Google Scholar 

  39. M. Warmuth, N. Simon, O. Mitina, R. Mathes, D. Fabbro, P. W. Manley, E. Buchdunger, K. Forster, I. Moarefi, and M. Hallek. Dual-specific Src and Abl kinase inhibitors, PP1 and CGP76030, inhibit growth and survival of cells expressing imatinib mesylate-resistant Bcr-Abl kinases. Blood 101:664–672 (2003).

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

We thank Dr. Robert W. Robey and Dr. Susan E. Bate at National Cancer Institute (Betheda, MD) for providing FTC and PhA. We also acknowledge Novartis Pharma AG (Basel, Switzerland) for kindly providing imatinib and cyclosporin A.

Author information

Authors and Affiliations

  1. Department of Pharmacy, Kumamoto University Hospital, 1-1-1 Honjo, Kumamoto, 860-8556, Japan

    Chie Hirayama, Hiroshi Watanabe, Reiko Nakashima, Takeru Nanbu, Akinobu Hamada & Hideyuki Saito

  2. Department of Molecular Cell Function, Graduate School of Medical and Pharmaceutical Sciences, Kumamoto University, 5-1 Oe-honmachi, Kumamoto, 862-0973, Japan

    Akihiko Kuniyasu & Hitoshi Nakayama

  3. Department of Hematology and Infectious Diseases, Kumamoto University Hospital, 1-1-1 Honjo, Kumamoto, 860-8556, Japan

    Tatsuya Kawaguchi

Authors
  1. Chie Hirayama
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Hiroshi Watanabe
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Reiko Nakashima
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Takeru Nanbu
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Akinobu Hamada
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Akihiko Kuniyasu
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Hitoshi Nakayama
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Tatsuya Kawaguchi
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Hideyuki Saito
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Hideyuki Saito.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Hirayama, C., Watanabe, H., Nakashima, R. et al. Constitutive Overexpression of P-glycoprotein, Rather than Breast Cancer Resistance Protein or Organic Cation Transporter 1, Contributes to Acquisition of Imatinib-Resistance in K562 Cells. Pharm Res 25, 827–835 (2008). https://doi.org/10.1007/s11095-007-9376-3

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11095-007-9376-3

Key words