Tumor Biology

, Volume 37, Issue 2, pp 1901–1908 | Cite as

Reversal of efflux of an anticancer drug in human drug-resistant breast cancer cells by inhibition of protein kinase Cα (PKCα) activity

  • Chan Woo Kim
  • Daisuke Asai
  • Jeong-Hun Kang
  • Akihiro Kishimura
  • Takeshi Mori
  • Yoshiki Katayama
Original Article


P-glycoprotein (Pgp) is a 170-kDa transmembrane protein that mediates the efflux of anticancer drugs from cells. Pgp overexpression has a distinct role in cells exhibiting multidrug resistance (MDR). We examined reversal of drug resistance in human MDR breast cancer cells by inhibition of protein kinase Cα (PKCα) activity, which is associated with Pgp-mediated efflux of anticancer drugs. PKCα activity was confirmed by measurement of phosphorylation levels of a PKCα-specific peptide substrate (FKKQGSFAKKK-NH2), showing relatively higher basal activity in drug-resistant MCF-7/ADR cells (84 %) than that in drug-sensitive MCF-7 cells (63 %). PKCα activity was effectively suppressed by the PKC inhibitor, Ro-31-7549, and reversal of intracellular accumulation of doxorubicin was observed by inhibition of PKCα activity in MCF-7/ADR cells compared with their intrinsic drug resistance. Importantly, increased accumulation of doxorubicin could enhance the therapeutic efficacy of doxorubicin in MDR cells significantly. These results suggest a potential for overcoming MDR via inhibition of PKCα activity with conventional anticancer drugs.


Drug resistance P-glycoprotein Protein kinase C α Protein kinase inhibitor Anticancer drug 



We thank Professor Masahiro Goto (Kyushu University) for assistance in CLSM studies and Dr. Ick Chan Kwon for the kind gift of the MCF-7/ADR cell line. We also thank Ms. Sigemi Terakubo and Ms. Ninyo Okamura (St. Marianna University School of Medicine) for technical assistance. This work was supported by a Grant-in-Aid for Scientific Research from the Ministry of Education, Culture, Sports, Science and Technology of Japan, and a grant from the KRIBB Research Initiative Program (Korean Biomedical Scientist Fellowship Program), Korea Research Institute of Bioscience and Biotechnology, Republic of Korea (C.W.K.).

Conflicts of interest


Supplementary material

13277_2015_3963_MOESM1_ESM.doc (118 kb)
ESM 1 (DOC 118 kb)


  1. 1.
    Zgurskaya HI, Nikaido H. Multidrug resistance mechanisms: drug efflux across two membranes. Mol Microbiol. 2000;37:219–25.CrossRefPubMedGoogle Scholar
  2. 2.
    Gottesman MM, Fojo T, Bates SE. Multidrug resistance in cancer: role of ATP-dependent transporters. Nat Rev Cancer. 2002;2:48–58.CrossRefPubMedGoogle Scholar
  3. 3.
    Thomas H, Coley HM. Overcoming multidrug resistance in cancer: an update on the clinical strategy of inhibiting P-glycoprotein. Cancer Control. 2003;10:159–65.PubMedGoogle Scholar
  4. 4.
    Jones PM, George AM. The ABC transporter structure and mechanism: perspectives on recent research. Cell Mol Life Sci. 2004;61:682–99.CrossRefPubMedGoogle Scholar
  5. 5.
    Deeley RG, Westlake C, Cole SPC. Transmembrane transport of endo- and xenobiotics by mammalian ATP-binding cassette multidrug resistance proteins. Physiol Rev. 2006;86:849–99.CrossRefPubMedGoogle Scholar
  6. 6.
    Fletcher JI, Haber M, Henderson MJ, Norris MD. ABC transporters in cancer: more than just drug efflux pumps. Nat Rev Cancer. 2010;10:147–56.CrossRefPubMedGoogle Scholar
  7. 7.
    Yu M, Ocana A, Tannock IF. Reversal of ATP-binding cassette drug transporter activity to modulate chemoresistance: why has it failed to provide clinical benefit? Cancer Metast Rev. 2013;32:211–27.CrossRefGoogle Scholar
  8. 8.
    Allen JD, van Loevezijn A, Lakhai JM, van der Valk M, van Tellingen O, Reid G, et al. Potent and specific inhibition of the breast cancer resistance protein multidrug transporter in vitro and in mouse intestine by a novel analogue of fumitremorgin C. Mol Cancer Ther. 2002;1:417–25.CrossRefPubMedGoogle Scholar
  9. 9.
    Doyle LA, Ross DD. Multidrug resistance mediated by the breast cancer resistance protein BCRP (ABCG2). Oncogene. 2003;22:7340–58.CrossRefPubMedGoogle Scholar
  10. 10.
    Dubikovskaya EA, Thorne SH, Pillow TH, Contag CH, Wender PA. Overcoming multidrug resistance of small-molecule therapeutics through conjugation with releasable octaarginine transporters. Proc Natl Acad Sci U S A. 2008;105:12128–33.CrossRefPubMedPubMedCentralGoogle Scholar
  11. 11.
    Abu Ajaj K, Graeser R, Kratz F. Zosuquidar and an albumin-binding prodrug of zosuquidar reverse multidrug resistance in breast cancer cells of doxorubicin and an albumin-binding prodrug of doxorubicin. Breast Cancer Res Treat. 2012;134:117–29.CrossRefPubMedGoogle Scholar
  12. 12.
    Goda K, Fenyvesi F, Bacso Z, Nagy H, Marian T, Megyeri A, et al. Complete inhibition of P-glycoprotein by simultaneous treatment with a distinct class of modulators and the UIC2 monoclonal antibody. J Pharmacol Exp Ther. 2007;320:81–8.CrossRefPubMedGoogle Scholar
  13. 13.
    Patel NR, Rathi A, Mongayt D, Torchilin VP. Reversal of multidrug resistance by co-delivery of tariquidar (XR9576) and paclitaxel using long-circulating liposomes. Int J Pharmaceut. 2011;416:296–9.CrossRefGoogle Scholar
  14. 14.
    Li XR, Li PZ, Zhang YH, Zhou YX, Chen XW, Huang YQ, et al. Novel mixed polymeric micelles for enhancing delivery of anticancer drug and overcoming multidrug resistance in tumor cell lines simultaneously. Pharm Res-Dordr. 2010;27:1498–511.CrossRefGoogle Scholar
  15. 15.
    Gottesman MM. Mechanisms of cancer drug resistance. Annu Rev Med. 2002;53:615–27.CrossRefPubMedGoogle Scholar
  16. 16.
    Malam Y, Loizidou M, Seifalian AM. Liposomes and nanoparticles: nanosized vehicles for drug delivery in cancer. Trends Pharmacol Sci. 2009;30:592–9.CrossRefPubMedGoogle Scholar
  17. 17.
    Hu CMJ, Zhang LF. Therapeutic nanoparticles to combat cancer drug resistance. Curr Drug Metab. 2009;10:836–41.CrossRefPubMedGoogle Scholar
  18. 18.
    Chambers TC, Mcavoy EM, Jacobs JW, Eilon G. Protein kinase C phosphorylates P-glycoprotein in multidrug resistant human KB carcinoma cells. J Biol Chem. 1990;265:7679–86.PubMedGoogle Scholar
  19. 19.
    Budworth J, Gant TW, Gescher A. Co-ordinate loss of protein kinase C and multidrug resistance gene expression in revertant MCF-7/Adr breast carcinoma cells. Br J Cancer. 1997;75:1330–5.CrossRefPubMedPubMedCentralGoogle Scholar
  20. 20.
    Idriss H, Urquidi V, Basavappa S. Selective modulation of P-glycoprotein’s ATPase and anion efflux regulation activities with PKC α and PKC ε in Sf9 cells. Cancer Chemother Pharmacol. 2000;46:287–92.CrossRefPubMedGoogle Scholar
  21. 21.
    Bates SE, Lee JS, Dickstein B, Spolyar M, Fojo AT. Differential modulation of P-glycoprotein transport by protein kinase inhibition. Biochemistry. 1993;32:9156–64.CrossRefPubMedGoogle Scholar
  22. 22.
    Chambers TC, Pohl J, Glass DB, Kuo JF. Phosphorylation by protein kinase C and cyclic AMP-dependent protein kinase of synthetic peptides derived from the linker region of human P-glycoprotein. Biochem J. 1994;299:309–15.CrossRefPubMedPubMedCentralGoogle Scholar
  23. 23.
    Ahmad S, Safa AR, Glazer RI. Modulation of P-glycoprotein by protein kinase C α in a baculovirus expression system. Biochemistry. 1994;33:10313–8.CrossRefPubMedGoogle Scholar
  24. 24.
    Idriss HT, Hannun YA, Boulpaep E, Basavappa S. Regulation of volume-activated chloride channels by P-glycoprotein: phosphorylation has the final say! J Physiol. 2000;524:629–36.CrossRefPubMedPubMedCentralGoogle Scholar
  25. 25.
    Fine RL, Chambers TC, Sachs CW. P-glycoprotein, multidrug resistance and protein kinase C. Oncologist. 1996;1:261–8.PubMedGoogle Scholar
  26. 26.
    Dean M, Fojo T, Bates S. Tumour stem cells and drug resistance. Nat Rev Cancer. 2005;5:275–84.CrossRefPubMedGoogle Scholar
  27. 27.
    Dean M. ABC transporters, drug resistance, and cancer stem cells. J Mammary Gland Biol. 2009;14:3–9.CrossRefGoogle Scholar
  28. 28.
    Tam WL, Lu HH, Buikhuisen J, Soh BS, Lim E, Reinhardt F, et al. Protein kinase C α is a central signaling node and therapeutic target for breast cancer stem cells. Cancer Cell. 2013;24:347–64.CrossRefPubMedPubMedCentralGoogle Scholar
  29. 29.
    Kang JH, Asai D, Toita R, Kitazaki H, Katayama Y. Plasma protein kinase C (PKC)α as a biomarker for the diagnosis of cancers. Carcinogenesis. 2009;30:1921–31.Google Scholar
  30. 30.
    Kang JH, Asai D, Yamada S, Toita R, Oishi J, Mori T, et al. A short peptide is a protein kinase C (PKC) α-specific substrate. Proteomics. 2008;8:2006–11.CrossRefPubMedGoogle Scholar
  31. 31.
    Wilkinson SE, Parker PJ, Nixon JS. Isoenzyme specificity of bisindolylmaleimides, selective inhibitors of protein kinase C. Biochem J. 1993;294:335–7.CrossRefPubMedPubMedCentralGoogle Scholar
  32. 32.
    Murphy CT, Westwick J. Selective inhibition of protein kinase C. Effect on platelet-activating-factor-induced platelet functional responses. Biochem J. 1992;283:159–64.CrossRefPubMedPubMedCentralGoogle Scholar
  33. 33.
    Scambia G, Ranelletti FO, Panici PB, Devincenzo R, Bonanno G, Ferrandina G, et al. Quercetin potentiates the effect of adriamycin in a multidrug-resistant MCF-7 human breast-cancer cell line: P-glycoprotein as a possible target. Cancer Chemother Pharmacol. 1994;34:459–64.CrossRefPubMedGoogle Scholar
  34. 34.
    Doyle LA, Yang WD, Abruzzo LV, Krogmann T, Gao YM, Rishi AK, et al. A multidrug resistance transporter from human MCF-7 breast cancer cells. Proc Natl Acad Sci U S A. 1998;95:15665–70.CrossRefPubMedPubMedCentralGoogle Scholar
  35. 35.
    Yu G, Ahmad S, Aquino A, Fairchild CR, Trepel JB, Ohno S, et al. Transfection with protein kinase C alpha confers increased multidrug resistance to MCF-7 cells expressing P-glycoprotein. Cancer Commun. 1991;3:181–9.PubMedGoogle Scholar
  36. 36.
    Blobe GC, Sachs CW, Khan WA, Fabbro D, Stabel S, Wetsel WC, et al. Selective regulation of expression of protein kinase C (PKC) isoenzymes in multidrug-resistant MCF-7 cells: functional significance of enhanced expression of PKC α. J Biol Chem. 1993;268:658–64.PubMedGoogle Scholar

Copyright information

© International Society of Oncology and BioMarkers (ISOBM) 2015

Authors and Affiliations

  • Chan Woo Kim
    • 1
    • 9
  • Daisuke Asai
    • 2
  • Jeong-Hun Kang
    • 3
  • Akihiro Kishimura
    • 1
    • 4
    • 5
  • Takeshi Mori
    • 1
    • 4
    • 5
  • Yoshiki Katayama
    • 1
    • 4
    • 5
    • 6
    • 7
    • 8
  1. 1.Department of Applied Chemistry, Faculty of EngineeringKyushu UniversityNishi-kuJapan
  2. 2.Department of MicrobiologySt. Marianna University School of MedicineMiyamae-kuJapan
  3. 3.Division of Biopharmaceutics and PharmacokineticsNational Cerebral and Cardiovascular Center Research InstituteSuitaJapan
  4. 4.Graduate School of Systems Life SciencesKyushu UniversityNishi-kuJapan
  5. 5.Center for Future ChemistryKyushu UniversityNishi-kuJapan
  6. 6.International Research Center for Molecular SystemsKyushu UniversityNishi-kuJapan
  7. 7.Center for Advanced Medical InnovationKyushu UniversityHigashi-kuJapan
  8. 8.Innovation Center for Medical Redox NavigationKyushu UniversityHigashi-kuJapan
  9. 9.Laboratory of Molecular Imaging and Nanomedicine, National Institute of Biomedical Imaging and BioengineeringNational Institutes of HealthBethesdaUSA

Personalised recommendations