Cellular Oncology

, Volume 40, Issue 3, pp 209–218 | Cite as

Inhibition of CDK4 sensitizes multidrug resistant ovarian cancer cells to paclitaxel by increasing apoptosiss

  • Yan Gao
  • Jacson Shen
  • Edwin Choy
  • Henry Mankin
  • Francis Hornicek
  • Zhenfeng DuanEmail author
Original Paper



Overexpression of cyclin-dependent kinase (CDK) 4 has been observed in a variety of cancers and has been found to contribute to tumor cell growth and proliferation. However, the effect of inhibition of CDK4 in ovarian cancer is unknown. We investigated the therapeutic effect of the CDK4 inhibitor palbociclib in combination with paclitaxel in ovarian cancer cells.


Cell viabilities were determined by MTT assay after exposure to different dosages of palbociclib and/or paclitaxel. Western blot, immunofluorescence, and Calcein AM assays were conducted to determine the mechanisms underlying the cytotoxic effects of palbociclib in combination with paclitaxel. CDK4 siRNA was used to validate the outcome of targeting CDK4 by palbociclib in ovarian cancer cells.


We found that combinations of palbociclib and paclitaxel significantly enhanced drug sensitivity in both Rb-positive (SKOV3TR) and Rb-negative (OVCAR8TR) ovarian cancer-derived cells. When combined with paclitaxel, palbociclib induced apoptosis in both SKOV3TR and OVCAR8TR cells. We also found that palbociclib inhibited the activity of P-glycoprotein (Pgp), and that siRNA-mediated CDK4 knockdown sensitized multidrug resistant (MDR) SKOV3TR and OVCAR8TR cells to paclitaxel.


Inhibition of CDK4 by palbociclib can enhance paclitaxel sensitivity in both Rb-positive and Rb-negative MDR ovarian cancer cells by increasing apoptosis. CDK4 may serve as a promising target in the treatment of ovarian cancer.


Ovarian cancer CDK4 Palbociclib Rb Paclitaxel Apoptosis MDR 



This work was supported in part by grants from the Gattegno and Wechsler funds. Dr. Duan is supported, in part, through a grant from the Sarcoma Foundation of America (SFA), a pilot grant from Sarcoma SPORE/NIH, and a grant from the National Cancer Institute (NCI)/National Institutes of Health (NIH), UO1, CA 151452.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no competing interests.

Supplementary material

13402_2017_316_MOESM1_ESM.pdf (113 kb)
Fig S1 (PDF 113 kb)
13402_2017_316_MOESM2_ESM.pdf (109 kb)
Fig S2 (PDF 109 kb)
13402_2017_316_MOESM3_ESM.pdf (115 kb)
Fig S3 (PDF 114 kb)
13402_2017_316_MOESM4_ESM.pdf (147 kb)
Fig S4 (PDF 146 kb)


  1. 1.
    R. Siegel, J. Ma, Z. Zou, A. Jemal, Cancer statistics, 2014. CA Cancer J. Clin. 64, 9–29 (2014)CrossRefPubMedGoogle Scholar
  2. 2.
    M. Zou, X. Zhang, C. Xu, IL6-induced metastasis modulators p-STAT3, MMP-2 and MMP-9 are targets of 3,3-diindolylmethane in ovarian cancer cells. Cell. Oncol. 39, 47–57 (2016)CrossRefGoogle Scholar
  3. 3.
    M.A. Bookman, M.F. Brady, W.P. McGuire, P.G. Harper, D.S. Alberts, M. Friedlander, N. Colombo, J.M. Fowler, P.A. Argenta, K. De Geest, D.G. Mutch, R.A. Burger, A.M. Swart, E.L. Trimble, C. Accario-Winslow, L.M. Roth, Evaluation of new platinum-based treatment regimens in advanced-stage ovarian cancer: a phase III trial of the gynecologic cancer intergroup. J. Clin. Oncol. 27, 1419–1425 (2009)CrossRefPubMedPubMedCentralGoogle Scholar
  4. 4.
    S. Vaughan, J.I. Coward, R.C. Bast Jr., A. Berchuck, J.S. Berek, J.D. Brenton, G. Coukos, C.C. Crum, R. Drapkin, D. Etemadmoghadam, M. Friedlander, H. Gabra, S.B. Kaye, C.J. Lord, E. Lengyel, D.A. Levine, I.A. McNeish, U. Menon, G.B. Mills, K.P. Nephew, A.M. Oza, A.K. Sood, E.A. Stronach, H. Walczak, D.D. Bowtell, F.R. Balkwill, Rethinking ovarian cancer: recommendations for improving outcomes. Nat. Rev. Cancer 11, 719–725 (2011)CrossRefPubMedPubMedCentralGoogle Scholar
  5. 5.
    B.Y. Karlan, A.M. Oza, G.E. Richardson, D.M. Provencher, V.L. Hansen, M. Buck, S.K. Chambers, P. Ghatage, C.H. Pippitt Jr., J.V. Brown 3rd, A. Covens, R.V. Nagarkar, M. Davy, C.A. Leath 3rd, H. Nguyen, D.E. Stepan, D.M. Weinreich, M. Tassoudji, Y.N. Sun, I.B. Vergote, Randomized, double-blind, placebo-controlled phase II study of AMG 386 combined with weekly paclitaxel in patients with recurrent ovarian cancer. J. Clin. Oncol. 30, 362–371 (2012)CrossRefPubMedGoogle Scholar
  6. 6.
    M. Strauss, J. Lukas, J. Bartek, Unrestricted cell cycling and cancer. Nat. Med. 1, 1245–1246 (1995)CrossRefPubMedGoogle Scholar
  7. 7.
    G. D'Andrilli, C. Kumar, G. Scambia, A. Giordano, Cell cycle genes in ovarian cancer: steps toward earlier diagnosis and novel therapies. Clin. Cancer Res. 10, 8132–8141 (2004)CrossRefPubMedGoogle Scholar
  8. 8.
    M. Malumbres, M. Barbacid, Cell cycle, CDKs and cancer: a changing paradigm. Nat. Rev. Cancer 9, 153–166 (2009)CrossRefPubMedGoogle Scholar
  9. 9.
    E.A. Musgrove, C.E. Caldon, J. Barraclough, A. Stone, R.L. Sutherland, Cyclin D as a therapeutic target in cancer. Nat. Rev. Cancer 11, 558–572 (2011)CrossRefPubMedGoogle Scholar
  10. 10.
    Y. Liao, Y. Feng, J. Shen, F.J. Hornicek and Z. Duan, The roles and therapeutic potential of cyclin-dependent kinases (CDKs) in sarcoma. Cancer Metastasis Rev. 35, 151–163 (2015)Google Scholar
  11. 11.
    T. Zhang, L.B. Nanney, C. Luongo, L. Lamps, K.J. Heppner, R.N. DuBois, R.D. Beauchamp, Concurrent overexpression of cyclin D1 and cyclin-dependent kinase 4 (Cdk4) in intestinal adenomas from multiple intestinal neoplasia (Min) mice and human familial adenomatous polyposis patients. Cancer Res. 57, 169–175 (1997)PubMedGoogle Scholar
  12. 12.
    Y. Hashiguchi, H. Tsuda, T. Inoue, S. Nishimura, T. Suzuki, N. Kawamura, Alteration of cell cycle regulators correlates with survival in epithelial ovarian cancer patients. Hum. Pathol. 35, 165–175 (2004)CrossRefPubMedGoogle Scholar
  13. 13.
    Y. Yang, B. Ma, L. Li, Y. Jin, W. Ben, D. Zhang, K. Jiang, S. Feng, L. Huang, J. Zheng, CDK2 and CDK4 play important roles in promoting the proliferation of SKOV3 ovarian carcinoma cells induced by tumor-associated macrophages. Oncol. Rep. 31, 2759–2768 (2014)PubMedGoogle Scholar
  14. 14.
    H. Shinozaki, S. Ozawa, N. Ando, H. Tsuruta, M. Terada, M. Ueda, M. Kitajima, Cyclin D1 amplification as a new predictive classification for squamous cell carcinoma of the esophagus, adding gene information. Clin. Cancer Res. 2, 1155–1161 (1996)PubMedGoogle Scholar
  15. 15.
    A. DeMichele, A.S. Clark, K.S. Tan, D.F. Heitjan, K. Gramlich, M. Gallagher, P. Lal, M. Feldman, P. Zhang, C. Colameco, D. Lewis, M. Langer, N. Goodman, S. Domchek, K. Gogineni, M. Rosen, K. Fox, P. O'Dwyer, CDK 4/6 inhibitor palbociclib (PD0332991) in Rb + advanced breast cancer: phase II activity, safety, and predictive biomarker assessment. Clin. Cancer Res. 21, 995–1001 (2015)CrossRefPubMedGoogle Scholar
  16. 16.
    M.A. Dickson, W.D. Tap, M.L. Keohan, S.P. D'Angelo, M.M. Gounder, C.R. Antonescu, J. Landa, L.X. Qin, D.D. Rathbone, M.M. Condy, Y. Ustoyev, A.M. Crago, S. Singer, G.K. Schwartz, Phase II trial of the CDK4 inhibitor PD0332991 in patients with advanced CDK4-amplified well-differentiated or dedifferentiated liposarcoma. J. Clin. Oncol. 31, 2024–2028 (2013)CrossRefPubMedPubMedCentralGoogle Scholar
  17. 17.
    F. Morschhauser, K. Bouabdallah, S. Stilgenbauer, C. Thieblemont, M. Wolf, S. de Guibert, F. Zettl, M. Hahka-Kemppinen, D.X. Wang, P. Brueck, Clinical activity of abemaciclib (LY2835219), a cell cycle inhibitor selective for CDK4 and CDK6, in patients with relapsed or refractory mantle cell lymphoma. Blood 124, 3067–3067 (2014)Google Scholar
  18. 18.
    J.R. Infante, G. Shapiro, P. Witteveen, J.F. Gerecitano, V. Ribrag, R. Chugh, I. Issa, A. Chakraborty, A. Matano and X. Zhao, In ASCO Annual Meeting Proceedings, p. 2528 (2014)Google Scholar
  19. 19.
    R.S. Finn, J.P. Crown, I. Lang, K. Boer, I.M. Bondarenko, S.O. Kulyk, J. Ettl, R. Patel, T. Pinter, M. Schmidt, Y. Shparyk, A.R. Thummala, N.L. Voytko, C. Fowst, X. Huang, S.T. Kim, S. Randolph, D.J. Slamon, The cyclin-dependent kinase 4/6 inhibitor palbociclib in combination with letrozole versus letrozole alone as first-line treatment of oestrogen receptor-positive, HER2-negative, advanced breast cancer (PALOMA-1/TRIO-18): a randomised phase 2 study. Lancet Oncol. 16, 25–35 (2015)CrossRefPubMedGoogle Scholar
  20. 20.
    D.W. Fry, P.J. Harvey, P.R. Keller, W.L. Elliott, M. Meade, E. Trachet, M. Albassam, X. Zheng, W.R. Leopold, N.K. Pryer, P.L. Toogood, Specific inhibition of cyclin-dependent kinase 4/6 by PD 0332991 and associated antitumor activity in human tumor xenografts. Mol. Cancer Ther. 3, 1427–1438 (2004)PubMedGoogle Scholar
  21. 21.
    J. Halder, C.N. Landen Jr., S.K. Lutgendorf, Y. Li, N.B. Jennings, D. Fan, G.M. Nelkin, R. Schmandt, M.D. Schaller, A.K. Sood, Focal adhesion kinase silencing augments docetaxel-mediated apoptosis in ovarian cancer cells. Clin. Cancer Res. 11, 8829–8836 (2005)CrossRefPubMedPubMedCentralGoogle Scholar
  22. 22.
    X. Yang, J. Shen, Y. Gao, Y. Feng, Y. Guan, Z. Zhang, H. Mankin, F.J. Hornicek, Z. Duan, Nsc23925 prevents the development of paclitaxel resistance by inhibiting the introduction of P-glycoprotein and enhancing apoptosis. Int. J. Cancer 137, 2029–2039 (2015)CrossRefPubMedPubMedCentralGoogle Scholar
  23. 23.
    X. Yang, A.K. Iyer, A. Singh, L. Milane, E. Choy, F.J. Hornicek, M.M. Amiji, Z. Duan, Cluster of differentiation 44 targeted hyaluronic acid based nanoparticles for MDR1 siRNA delivery to overcome drug resistance in ovarian cancer. Pharm. Res. 32, 2097–2109 (2015)CrossRefPubMedGoogle Scholar
  24. 24.
    H. Devalapally, Z. Duan, M.V. Seiden, M.M. Amiji, Modulation of drug resistance in ovarian adenocarcinoma by enhancing intracellular ceramide using tamoxifen-loaded biodegradable polymeric nanoparticles. Clin. Cancer Res. 14, 3193–3203 (2008)CrossRefPubMedGoogle Scholar
  25. 25.
    Z. Duan, K.A. Brakora, M.V. Seiden, Inhibition of ABCB1 (MDR1) and ABCB4 (MDR3) expression by small interfering RNA and reversal of paclitaxel resistance in human ovarian cancer cells. Mol. Cancer Ther. 3, 833–838 (2004)PubMedGoogle Scholar
  26. 26.
    Z. Duan, A.J. Feller, R.T. Penson, B.A. Chabner, M.V. Seiden, Discovery of differentially expressed genes associated with paclitaxel resistance using cDNA array technology: analysis of interleukin (IL) 6, IL-8, and monocyte chemotactic protein 1 in the paclitaxel-resistant phenotype. Clin. Cancer Res. 5, 3445–3453 (1999)PubMedGoogle Scholar
  27. 27.
    W.R. Sellers, W.G. Kaelin Jr., Role of the retinoblastoma protein in the pathogenesis of human cancer. J. Clin. Oncol. 15, 3301–3312 (1997)CrossRefPubMedGoogle Scholar
  28. 28.
    T.U. Barbie, D.A. Barbie, D.T. MacLaughlin, S. Maheswaran, P.K. Donahoe, Mullerian inhibiting substance inhibits cervical cancer cell growth via a pathway involving p130 and p107. Proc. Natl. Acad. Sci. U. S. A. 100, 15601–15606 (2003)CrossRefPubMedPubMedCentralGoogle Scholar
  29. 29.
    T.U. Ha, D.L. Segev, D. Barbie, P.T. Masiakos, T.T. Tran, D. Dombkowski, M. Glander, T.R. Clarke, H.K. Lorenzo, P.K. Donahoe, S. Maheswaran, Mullerian inhibiting substance inhibits ovarian cell growth through an Rb-independent mechanism. J. Biol. Chem. 275, 37101–37109 (2000)CrossRefPubMedGoogle Scholar
  30. 30.
    K. Milde-Langosch, M. Hagen, A.M. Bamberger, T. Loning, Expression and prognostic value of the cell-cycle regulatory proteins, Rb, p16MTS1, p21WAF1, p27KIP1, cyclin E, and cyclin D2, in ovarian cancer. Int. J. Gynecol. Pathol. 22, 168–174 (2003)CrossRefPubMedGoogle Scholar
  31. 31.
    J.P. Leonard, A.S. LaCasce, M.R. Smith, A. Noy, L.R. Chirieac, S.J. Rodig, J.Q. Yu, S. Vallabhajosula, H. Schoder, P. English, D.S. Neuberg, P. Martin, M.M. Millenson, S.A. Ely, R. Courtney, N. Shaik, K.D. Wilner, S. Randolph, A.D. Van den Abbeele, S.Y. Chen-Kiang, J.T. Yap, G.I. Shapiro, Selective CDK4/6 inhibition with tumor responses by PD0332991 in patients with mantle cell lymphoma. Blood 119, 4597–4607 (2012)CrossRefPubMedGoogle Scholar
  32. 32.
    C.K. Ingemarsdotter, L.A. Tookman, A. Browne, K. Pirlo, R. Cutts, C. Chelela, K.F. Khurrum, E.Y. Leung, S. Dowson, L. Webber, I. Khan, D. Ennis, N. Syed, T.R. Crook, J.D. Brenton, M. Lockley, I.A. McNeish, Paclitaxel resistance increases oncolytic adenovirus efficacy via upregulated CAR expression and dysfunctional cell cycle control. Mol. Oncol. 9, 791–805 (2015)CrossRefPubMedGoogle Scholar
  33. 33.
    A.S. Clark, P.J. O'Dwyer, D. Heitjan, P. Lal, M.D. Feldman, M. Gallagher, C. Redlinger, C. Colameco, D. Lewis, K. Zafman, M. Langer, M.A. Rosen, K. Gogineni, A.R. Bradbury, S.M. Domchek, K.R. Fox, A. DeMichele, A phase I trial of palbociclib and paclitaxel in metastatic breast cancer. J. Clin. Oncol. 32, 5 (2014)Google Scholar
  34. 34.
    S. Patel, L. Kumar, N. Singh, Metformin and epithelial ovarian cancer therapeutics. Cell. Oncol. 38, 365–375 (2015)CrossRefGoogle Scholar
  35. 35.
    K. Goetze, C.G. Fabian, A. Siebers, L. Binz, D. Faber, S. Indraccolo, G. Nardo, U.G. Sattler, W. Mueller-Klieser, Manipulation of tumor metabolism for therapeutic approaches: ovarian cancer-derived cell lines as a model system. Cell. Oncol. 38, 377–385 (2015)CrossRefGoogle Scholar
  36. 36.
    R.I. Sanchez, S. Mesia-Vela, F.C. Kauffman, Challenges of cancer drug design: a drug metabolism perspective. Curr. Cancer Drug Targets 1, 1–32 (2001)CrossRefPubMedGoogle Scholar
  37. 37.
    A.M. Reed, M.L. Fishel, M.R. Kelley, Small-molecule inhibitors of proteins involved in base excision repair potentiate the anti-tumorigenic effect of existing chemotherapeutics and irradiation. Future Oncol. 5, 713–726 (2009)CrossRefPubMedPubMedCentralGoogle Scholar
  38. 38.
    D.C. Altieri, The case for survivin as a regulator of microtubule dynamics and cell-death decisions. Curr. Opin. Cell Biol. 18, 609–615 (2006)CrossRefPubMedGoogle Scholar
  39. 39.
    N. Bah, L. Maillet, J. Ryan, S. Dubreil, F. Gautier, A. Letai, P. Juin, S. Barille-Nion, Bcl-xL controls a switch between cell death modes during mitotic arrest. Cell Death Dis. 5, e1291 (2014)CrossRefPubMedPubMedCentralGoogle Scholar
  40. 40.
    S.N. Kolomeichuk, D.T. Terrano, C.S. Lyle, K. Sabapathy, T.C. Chambers, Distinct signaling pathways of microtubule inhibitors--vinblastine and Taxol induce JNK-dependent cell death but through AP-1-dependent and AP-1-independent mechanisms, respectively. FEBS J. 275, 1889–1899 (2008)CrossRefPubMedGoogle Scholar
  41. 41.
    S.J. Lim, M.K. Choi, M.J. Kim, J.K. Kim, Alpha-tocopheryl succinate potentiates the paclitaxel-induced apoptosis through enforced caspase 8 activation in human H460 lung cancer cells. Exp. Mol. Med. 41, 737–745 (2009)CrossRefPubMedPubMedCentralGoogle Scholar
  42. 42.
    X.H. Zhang, Y. Cheng, J.Y. Shin, J.O. Kim, J.E. Oh, J.H. Kang, A CDK4/6 inhibitor enhances cytotoxicity of paclitaxel in lung adenocarcinoma cells harboring mutant KRAS as well as wild-type KRAS. Cancer Biol. Ther. 14, 597–605 (2013)CrossRefPubMedPubMedCentralGoogle Scholar
  43. 43.
    D.W. Fry, D.C. Bedford, P.H. Harvey, A. Fritsch, P.R. Keller, Z. Wu, E. Dobrusin, W.R. Leopold, A. Fattaey, M.D. Garrett, Cell cycle and biochemical effects of PD 0183812. A potent inhibitor of the cyclin D-dependent kinases CDK4 and CDK6. J. Biol. Chem. 276, 16617–16623 (2001)CrossRefPubMedGoogle Scholar
  44. 44.
    B. Taylor-Harding, P.J. Aspuria, H. Agadjanian, D.J. Cheon, T. Mizuno, D. Greenberg, J.R. Allen, L. Spurka, V. Funari, E. Spiteri, Q. Wang, S. Orsulic, C. Walsh, B.Y. Karlan, W.R. Wiedemeyer, Cyclin E1 and RTK/RAS signaling drive CDK inhibitor resistance via activation of E2F and ETS. Oncotarget 6, 696–714 (2015)CrossRefPubMedGoogle Scholar
  45. 45.
    G.E. Konecny, B. Winterhoff, T. Kolarova, J. Qi, K. Manivong, J. Dering, G. Yang, M. Chalukya, H.J. Wang, L. Anderson, K.R. Kalli, R.S. Finn, C. Ginther, S. Jones, V.E. Velculescu, D. Riehle, W.A. Cliby, S. Randolph, M. Koehler, L.C. Hartmann, D.J. Slamon, Expression of p16 and retinoblastoma determines response to CDK4/6 inhibition in ovarian cancer. Clin. Cancer Res. 17, 1591–1602 (2011)CrossRefPubMedPubMedCentralGoogle Scholar
  46. 46.
    G. Szakacs, J.K. Paterson, J.A. Ludwig, C. Booth-Genthe, M.M. Gottesman, Targeting multidrug resistance in cancer. Nat. Rev. Drug Discov. 5, 219–234 (2006)CrossRefPubMedGoogle Scholar
  47. 47.
    Z. Wang, Y. Chen, H. Liang, A. Bender, R.C. Glen, A. Yan, P-glycoprotein substrate models using support vector machines based on a comprehensive data set. J. Chem. Inf. Model. 51, 1447–1456 (2011)CrossRefPubMedGoogle Scholar
  48. 48.
    T. Hegedus, L. Orfi, A. Seprodi, A. Varadi, B. Sarkadi, G. Keri, Interaction of tyrosine kinase inhibitors with the human multidrug transporter proteins, MDR1 and MRP1. Biochim. Biophys. Acta 1587, 318–325 (2002)CrossRefPubMedGoogle Scholar
  49. 49.
    X.K. Wang, L.W. Fu, Interaction of tyrosine kinase inhibitors with the MDR-related ABC transporter proteins. Curr. Drug Metab. 11, 618–628 (2010)CrossRefPubMedGoogle Scholar
  50. 50.
    Z. Duan, A.J. Feller, H.C. Toh, T. Makastorsis, M.V. Seiden, TRAG-3, a novel gene, isolated from a taxol-resistant ovarian carcinoma cell line. Gene 229, 75–81 (1999)CrossRefPubMedGoogle Scholar

Copyright information

© International Society for Cellular Oncology 2017

Authors and Affiliations

  • Yan Gao
    • 1
    • 2
  • Jacson Shen
    • 2
  • Edwin Choy
    • 2
  • Henry Mankin
    • 2
  • Francis Hornicek
    • 2
  • Zhenfeng Duan
    • 2
    Email author
  1. 1.Department of Clinical Laboratory Diagnostics, Beijing Friendship HospitalCapital Medical UniversityBeijingChina
  2. 2.Sarcoma Biology Laboratory, Center for Sarcoma and Connective Tissue OncologyMassachusetts General Hospital and Harvard Medical SchoolBostonUSA

Personalised recommendations