Tumor Biology

, Volume 37, Issue 1, pp 685–698 | Cite as

Approach for chemosensitization of cisplatin-resistant ovarian cancer by cucurbitacin B

  • Fardous F. El-Senduny
  • Farid A. Badria
  • Ahmed M. EL-Waseef
  • Subhash C. Chauhan
  • Fathi HalaweishEmail author
Original Article


Ovarian cancer is the most deadly gynecological cancer. The first line in treatment is platinum-based drugs. However, most patients suffer from tumor recurrence, characterized by resistance to cisplatin. A plausible approach to address the tumor resistance is to co-administer the chemotherapeutic agents along with natural products to offer a synergistic effect and optimize the dosage regimen. Cucurbitacin B is a natural product and displays antitumor activity against a wide array of cancer cell lines. The aim of this work is to determine the antitumor activity against ovarian cancer cell line (A2780) and possible sensitization activity on cisplatin-resistant cell line (A2780CP) in 2D and 3D culture model. 3D spheroids were generated from A2780CP cell line. A2780, A2780CP, and the spheroids were treated with cucurbitacin B, cisplatin alone, or pretreated with cucurbitacin B followed by cisplatin. The viability, cell cycle, and apoptosis were analyzed. Level of ROS and total glutathione was measured. In this study, cucurbitacin B showed cytotoxicity against the ovarian cancer cell lines, and pretreatment of A2780CP cells leads to a significant increase in the cytotoxicity of cisplatin. The mechanism behind the sensitization effect was dependent in part on the depletion of the total glutathione, an increase in ROS through a decrease in the level of dual-specificity tyrosine-regulated kinase (Dyrk1B), decrease in pERK1/2 and pSTAT3 level. The viability of spheroids treated with a combination of cisplatin and cucurbitacin B were significantly decreased. The resulting data shows that cucurbitacin B is a promising chemosensitizer for the cisplatin-resistant ovarian cancer.


Ovarian cancer Chemotherapy resistance Cisplatin Cucurbitacin B Dyrk1B 



We would like to thank Lucas Kopel and Mahmoud Salama (Chemistry and Biochemistry Department, South Dakota State University, Brookings, SD, USA). This work was supported by the Egyptian government via the Egyptian Ministry of Higher Education and Scientific Research.

Conflict of interest


Authors’ contributions

FE helped in the design of the study, carried out the research, and drafted the manuscript. FB, AE, and FH participated in the design and conceive of the study. SC helped in the design and revision of the draft. All authors read and approved the final manuscript.

Supplementary material

13277_2015_3773_MOESM1_ESM.docx (2.3 mb)
ESM 1 (DOCX 2.30 MB)


  1. 1.
    Jemal A, Bray F, Center MM, Ferlay J, Ward E, Forman D. Global cancer statistics. CA Cancer J Clin. 2011;61(2):69–90. doi: 10.3322/caac.20107.CrossRefPubMedGoogle Scholar
  2. 2.
    Jemal A, Siegel R, Ward E, Hao Y, Xu J, Thun MJ. Cancer statistics, 2009. CA Cancer J Clin. 2009;59(4):225–49. doi: 10.3322/caac.20006.CrossRefPubMedGoogle Scholar
  3. 3.
    Iwanicki MP, Davidowitz RA, Ng MR, Besser A, Muranen T, Merritt M, et al. Ovarian cancer spheroids use myosin-generated force to clear the mesothelium. Cancer Discov. 2011;1(2):144–57. doi: 10.1158/ Scholar
  4. 4.
    Nederman T, Acker H, Carlsson J. Penetration of substances into tumor tissue: a methodological study with microelectrodes and cellular spheroids. In Vitro. 1983;19(6):479–88.CrossRefPubMedGoogle Scholar
  5. 5.
    Rebucci M, Michiels C. Molecular aspects of cancer cell resistance to chemotherapy. Biochem Pharmacol. 2013;85(9):1219–26. doi: 10.1016/j.bcp.2013.02.017.CrossRefPubMedGoogle Scholar
  6. 6.
    Stewart DJ. Mechanisms of resistance to cisplatin and carboplatin. Crit Rev Oncol Hematol. 2007;63(1):12–31. doi: 10.1016/j.critrevonc.2007.02.001.CrossRefPubMedGoogle Scholar
  7. 7.
    Mercurio F, Manning AM. Multiple signals converging on NF-kappaB. Curr Opin Cell Biol. 1999;11(2):226–32.CrossRefPubMedGoogle Scholar
  8. 8.
    Karin M, Lin A. NF-kappaB at the crossroads of life and death. Nat Immunol. 2002;3(3):221–7. doi: 10.1038/ni0302-221.CrossRefPubMedGoogle Scholar
  9. 9.
    Hua Y, Jove R. The STATs of cancer—new molecular targets come of age. Nat Rev Cancer. 2004;4(2):97–105. doi: 10.1038/nrc1275.CrossRefGoogle Scholar
  10. 10.
    Meinhold-Heerlein I, Bauerschlag D, Hilpert F, Dimitrov P, Sapinoso LM, Orlowska-Volk M, et al. Molecular and prognostic distinction between serous ovarian carcinomas of varying grade and malignant potential. Oncogene. 2005;24(6):1053–65. doi: 10.1038/sj.onc.1208298.CrossRefPubMedGoogle Scholar
  11. 11.
    Duan Z, Foster R, Bell DA, Mahoney J, Wolak K, Vaidya A, et al. Signal transducers and activators of transcription 3 pathway activation in drug-resistant ovarian cancer. Clin Cancer Res. 2006;12(17):5055–63. doi: 10.1158/1078-0432.ccr-06-0861.CrossRefPubMedGoogle Scholar
  12. 12.
    Han Z, Feng J, Hong Z, Chen L, Li W, Liao S, et al. Silencing of the STAT3 signaling pathway reverses the inherent and induced chemoresistance of human ovarian cancer cells. Biochem Biophys Res Commun. 2013;435(2):188–94. doi: 10.1016/j.bbrc.2013.04.087.CrossRefPubMedGoogle Scholar
  13. 13.
    Platanias LC. Map kinase signaling pathways and hematologic malignancies. Blood. 2003;101(12):4667–79. doi: 10.1182/blood-2002-12-3647.CrossRefPubMedGoogle Scholar
  14. 14.
    Lu Z, Xu S. ERK1/2 MAP kinases in cell survival and apoptosis. IUBMB Life. 2006;58(11):621–31. doi: 10.1080/15216540600957438.CrossRefPubMedGoogle Scholar
  15. 15.
    Kim KY, Choi KC, Park SH, Auersperg N, Leung PC. Extracellular signal-regulated protein kinase, but not c-Jun N-terminal kinase, is activated by type II gonadotropin-releasing hormone involved in the inhibition of ovarian cancer cell proliferation. J Clin Endocrinol Metab. 2005;90(3):1670–7. doi: 10.1210/jc.2004-1636.CrossRefPubMedGoogle Scholar
  16. 16.
    Hayakawa J, Ohmichi M, Kurachi H, Ikegami H, Kimura A, Matsuoka T, et al. Inhibition of extracellular signal-regulated protein kinase or c-Jun N-terminal protein kinase cascade, differentially activated by cisplatin, sensitizes human ovarian cancer cell line. J Biol Chem. 1999;274(44):31648–54.CrossRefPubMedGoogle Scholar
  17. 17.
    Wulfkuhle JD, Aquino JA, Calvert VS, Fishman DA, Coukos G, Liotta LA, et al. Signal pathway profiling of ovarian cancer from human tissue specimens using reverse-phase protein microarrays. Proteomics. 2003;3(11):2085–90. doi: 10.1002/pmic.200300591.CrossRefPubMedGoogle Scholar
  18. 18.
    Bast Jr RC. Molecular approaches to personalizing management of ovarian cancer. Ann Oncol. 2011;22 Suppl 8:viii5–viii15. doi: 10.1093/annonc/mdr516.CrossRefPubMedPubMedCentralGoogle Scholar
  19. 19.
    Hu J, Friedman E. Depleting mirk kinase increases cisplatin toxicity in ovarian cancer cells. Genes Cancer. 2010;1(8):803–11. doi: 10.1177/1947601910377644.CrossRefPubMedPubMedCentralGoogle Scholar
  20. 20.
    Lai GM, Ozols RF, Young RC, Hamilton TC. Effect of glutathione on DNA repair in cisplatin-resistant human ovarian cancer cell lines. J Natl Cancer Inst. 1989;81(7):535–9.CrossRefPubMedGoogle Scholar
  21. 21.
    Hockenbery DM, Oltvai ZN, Yin XM, Milliman CL, Korsmeyer SJ. Bcl-2 functions in an antioxidant pathway to prevent apoptosis. Cell. 1993;75(2):241–51.CrossRefPubMedGoogle Scholar
  22. 22.
    Chen X, Bao J, Guo J, Ding Q, Lu J, Huang M, et al. Biological activities and potential molecular targets of cucurbitacins: a focus on cancer. Anticancer Drugs. 2012;23(8):777–87. doi: 10.1097/CAD.0b013e3283541384.CrossRefPubMedGoogle Scholar
  23. 23.
    Alghasham AA. Cucurbitacins - a promising target for cancer therapy. Int J Health Sci. 2013;7(1):77–89.CrossRefGoogle Scholar
  24. 24.
    Chen JC, Chiu MH, Nie RL, Cordell GA, Qiu SX. Cucurbitacins and cucurbitane glycosides: structures and biological activities. Nat Prod Rep. 2005;22(3):386–99. doi: 10.1039/b418841c.CrossRefPubMedGoogle Scholar
  25. 25.
    Chen W, Leiter A, Yin D, Meiring M, Louw VJ, Koeffler HP. Cucurbitacin B inhibits growth, arrests the cell cycle, and potentiates antiproliferative efficacy of cisplatin in cutaneous squamous cell carcinoma cell lines. Int J Oncol. 2010;37(3):737–43.PubMedGoogle Scholar
  26. 26.
    Iwanski GB, Lee DH, En-Gal S, Doan NB, Castor B, Vogt M, et al. Cucurbitacin B, a novel in vivo potentiator of gemcitabine with low toxicity in the treatment of pancreatic cancer. Br J Pharmacol. 2010;160(4):998–1007. doi: 10.1111/j.1476-5381.2010.00741.x.CrossRefPubMedPubMedCentralGoogle Scholar
  27. 27.
    Lee DH, Thoennissen NH, Goff C, Iwanski GB, Forscher C, Doan NB, et al. Synergistic effect of low-dose cucurbitacin B and low-dose methotrexate for treatment of human osteosarcoma. Cancer Lett. 2011;306(2):161–70. doi: 10.1016/j.canlet.2011.03.001.CrossRefPubMedPubMedCentralGoogle Scholar
  28. 28.
    Friedrich J, Seidel C, Ebner R, Kunz-Schughart LA. Spheroid-based drug screen: considerations and practical approach. Nat Protoc. 2009;4(3):309–24. doi: 10.1038/nprot.2008.226.CrossRefPubMedGoogle Scholar
  29. 29.
    Tung YC, Hsiao AY, Allen SG, Torisawa YS, Ho M, Takayama S. High-throughput 3D spheroid culture and drug testing using a 384 hanging drop array. Analyst. 2011;136(3):473–8. doi: 10.1039/c0an00609b.CrossRefPubMedGoogle Scholar
  30. 30.
    Hamaguchi K, Godwin AK, Yakushiji M, O'Dwyer PJ, Ozols RF, Hamilton TC. Cross-resistance to diverse drugs is associated with primary cisplatin resistance in ovarian cancer cell lines. Cancer Res. 1993;53(21):5225–32.PubMedGoogle Scholar
  31. 31.
    Bartalis J, Halaweish FT. In vitro and QSAR studies of cucurbitacins on HepG2 and HSC-T6 liver cell lines. Bioorg Med Chem. 2011;19(8):2757–66. doi: 10.1016/j.bmc.2011.01.037.CrossRefPubMedGoogle Scholar
  32. 32.
    Alfadda AA, Sallam RM. Reactive oxygen species in health and disease. J Biomed Biotechnol. 2012;2012:936486. doi: 10.1155/2012/936486.CrossRefPubMedPubMedCentralGoogle Scholar
  33. 33.
    Chou TC, Martin N. CompuSyn for drug combinations: PC software and user’s guide: a computer program for quantitation of synergism and antagonism in drug combinations, and the determination of IC50 and ED50 and LD50 Values, ComboSyn Inc, Paramus, (NJ), 2005.Google Scholar
  34. 34.
    Tait SWG, Green DR. Mitochondria and cell death: outer membrane permeabilization and beyond. Nat Rev Mol Cell Biol. 2010;11(9):621–32. doi: 10.1038/nrm2952.CrossRefPubMedGoogle Scholar
  35. 35.
    Tan L, Kwok RP, Shukla A, Kshirsagar M, Zhao L, Opipari Jr AW, et al. Trichostatin A restores Apaf-1 function in chemoresistant ovarian cancer cells. Cancer. 2011;117(4):784–94. doi: 10.1002/cncr.25649.CrossRefPubMedGoogle Scholar
  36. 36.
    Shen M, Feng Y, Gao C, Tao D, Hu J, Reed E, et al. Detection of Cyclin B1 Expression in G1-phase cancer cell lines and cancer tissues by postsorting western blot analysis. Cancer Res. 2004;64(5):1607–10. doi: 10.1158/0008-5472.can-03-3321.CrossRefPubMedGoogle Scholar
  37. 37.
    Zheng H, Hu W, Deavers MT, Shen D-Y, Fu S, Li Y-F, et al. Nuclear cyclin B1 is overexpressed in low-malignant-potential ovarian tumors but not in epithelial ovarian cancer. Am J Obstet Gynecol. 2009;201(4):367.e1-.e6. doi: 10.1016/j.ajog.2009.05.021.CrossRefGoogle Scholar
  38. 38.
    Pahl HL. Activators and target genes of Rel/NF-kappaB transcription factors. Oncogene. 1999;18(49):6853–66. doi: 10.1038/sj.onc.1203239.CrossRefPubMedGoogle Scholar
  39. 39.
    Wang CY, Cusack Jr JC, Liu R, Baldwin Jr AS. Control of inducible chemoresistance: enhanced anti-tumor therapy through increased apoptosis by inhibition of NF-kappaB. Nat Med. 1999;5(4):412–7. doi: 10.1038/7410.CrossRefPubMedGoogle Scholar
  40. 40.
    Annunziata CM, Stavnes HT, Kleinberg L, Berner A, Hernandez LF, Birrer MJ, et al. Nuclear factor kappaB transcription factors are coexpressed and convey a poor outcome in ovarian cancer. Cancer. 2010;116(13):3276–84. doi: 10.1002/cncr.25190.CrossRefPubMedPubMedCentralGoogle Scholar
  41. 41.
    Jin HR, Jin X, Dat NT, Lee JJ. Cucurbitacin B suppresses the transactivation activity of RelA/p65. J Cell Biochem. 2011;112(6):1643–50. doi: 10.1002/jcb.23078.CrossRefPubMedGoogle Scholar
  42. 42.
    Chan KT, Li K, Liu SL, Chu KH, Toh M, Xie WD. Cucurbitacin B inhibits STAT3 and the Raf/MEK/ERK pathway in leukemia cell line K562. Cancer Lett. 2010;289(1):46–52. doi: 10.1016/j.canlet.2009.07.015.CrossRefPubMedGoogle Scholar
  43. 43.
    Chan KT, Meng FY, Li Q, Ho CY, Lam TS, To Y, et al. Cucurbitacin B induces apoptosis and S phase cell cycle arrest in BEL-7402 human hepatocellular carcinoma cells and is effective via oral administration. Cancer Lett. 2010;294(1):118–24. doi: 10.1016/j.canlet.2010.01.029.CrossRefPubMedGoogle Scholar
  44. 44.
    Duangmano S, Sae-Lim P, Suksamrarn A, Patmasiriwat P, Domann FE. Cucurbitacin B causes increased radiation sensitivity of human breast cancer Cells via G2/M Cell Cycle Arrest. J Oncol. 2012;2012:601682. doi: 10.1155/2012/601682.CrossRefPubMedPubMedCentralGoogle Scholar
  45. 45.
    Liu T, Peng H, Zhang M, Deng Y, Wu Z. Cucurbitacin B, a small molecule inhibitor of the Stat3 signaling pathway, enhances the chemosensitivity of laryngeal squamous cell carcinoma cells to cisplatin. Eur J Pharmacol. 2010;641(1):15–22. doi: 10.1016/j.ejphar.2010.04.062.CrossRefPubMedGoogle Scholar
  46. 46.
    Kaufmann SH, Desnoyers S, Ottaviano Y, Davidson NE, Poirier GG. Specific Proteolytic Cleavage of Poly(ADP-ribose) Polymerase: an early marker of chemotherapy-induced apoptosis. Cancer Res. 1993;53(17):3976–85.PubMedGoogle Scholar
  47. 47.
    Dantzer F, de la Rubia G, Ménissier-de Murcia J, Hostomsky Z, de Murcia G, Schreiber V. Base excision repair is impaired in mammalian cells lacking poly(adp-ribose) polymerase-1†. Biochemistry. 2000;39(25):7559–69. doi: 10.1021/bi0003442.CrossRefPubMedGoogle Scholar
  48. 48.
    Bowman KJ, Newell DR, Calvert AH, Curtin NJ. Differential effects of the poly (ADP-ribose) polymerase (PARP) inhibitor NU1025 on topoisomerase I and II inhibitor cytotoxicity in L1210 cells in vitro. Br J Cancer. 2001;84(1):106.CrossRefPubMedPubMedCentralGoogle Scholar
  49. 49.
    Godoy H, Mhawech-Fauceglia P, Beck A, Miller A, Lele S, Odunsi K. Expression of poly (adenosine diphosphate-ribose) polymerase and p53 in epithelial ovarian cancer and their role in prognosis and disease outcome. Int J Gynecol Pathol: Off J Int Soc Gynecol Pathol. 2011;30(2):139–44. doi: 10.1097/PGP.0b013e3181fa5a64.CrossRefGoogle Scholar
  50. 50.
    Michels J, Vitale I, Galluzzi L, Adam J, Olaussen KA, Kepp O, et al. Cisplatin resistance associated with PARP hyperactivation. Cancer Res. 2013;73(7):2271–80. doi: 10.1158/0008-5472.CAN-12-3000.CrossRefPubMedGoogle Scholar
  51. 51.
    Godwin AK, Meister A, O'Dwyer PJ, Huang CS, Hamilton TC, Anderson ME. High resistance to cisplatin in human ovarian cancer cell lines is associated with marked increase of glutathione synthesis. Proc Natl Acad Sci U S A. 1992;89(7):3070–4.CrossRefPubMedPubMedCentralGoogle Scholar
  52. 52.
    Gamcsik MP, Kasibhatla MS, Teeter SD, Colvin OM. Glutathione levels in human tumors. Biomarkers. 2012;17(8):671–91. doi: 10.3109/1354750X.2012.715672.CrossRefPubMedPubMedCentralGoogle Scholar
  53. 53.
    Andrews PA, Schiefer MA, Murphy MP, Howell SB. Enhanced potentiation of cisplatin cytotoxicity in human ovarian carcinoma cells by prolonged glutathione depletion. Chem Biol Interact. 1988;65(1):51–8.CrossRefPubMedGoogle Scholar
  54. 54.
    Kausar H, Munagala R, Bansal SS, Aqil F, Vadhanam MV, Gupta RC. Cucurbitacin B potently suppresses non-small-cell lung cancer growth: identification of intracellular thiols as critical targets. Cancer Lett. 2013;332(1):35–45. doi: 10.1016/j.canlet.2013.01.008.CrossRefPubMedGoogle Scholar
  55. 55.
    Burleson K, Hansen L, Skubitz A. Ovarian carcinoma spheroids disseminate on type I collagen and invade live human mesothelial cell monolayers. Clin Exp Metastasis. 2005;21:685-697.Google Scholar
  56. 56.
    Durand R, Sutherland R. Effects of intercellular contact on repair of radiation damage. Exp Cell Res. 1972;71:75–80.CrossRefPubMedGoogle Scholar
  57. 57.
    Filipovich I, Sorokina N, Robillard N, Chatal J. Radiation-induced apoptosis in human ovarian carcinoma cells growing as a monolayer and as multicell spheroids. Int J Cancer. 1997;72:851–9.CrossRefGoogle Scholar
  58. 58.
    Graham C, Kobayashi H, Stankiewicz K, Man S, Kapitain S, Kerbel R. Rapid acquisition of multicellular drug resistance after a single exposure of mammary tumor cells to antitumor alkylating agents. J Natl Cancer Inst. 1994;86:975–82.CrossRefPubMedGoogle Scholar
  59. 59.
    Makhija S, Taylor D, Gibb R, Gercel-Taylor. Taxol-induced Bcl-2 phosphorylation in ovarian cancer cell monolayer and spheroids. Int J Oncol. 1999;14:515–21.PubMedGoogle Scholar
  60. 60.
    Bardies M, Thedrez P, Gestin J, Marcille B, Guerreau D, Faivre-Chauvet A, et al. Use of multi-cell spheroids of ovarian carcinoma as an intraperitoneal radio-immunotherapy model: uptake, retention kinetics and dosimetric evaluation. Int J Cancer. 1992;50:984–91.CrossRefPubMedGoogle Scholar
  61. 61.
    Sutherland R, MacDonald H, Howell R. Multicellular spheroids: a new model target for in vitro studies of immunity to solid tumor allografts. J Natl Cancer Inst. 1977;58:1849–53.CrossRefPubMedGoogle Scholar
  62. 62.
    Sutherland R, McCredie J, Inch W. Growth of multicell spheroids in tissue culture as a model of nodular carcinoma. J Natl Cancer Inst. 1971;46:113–20.PubMedGoogle Scholar

Copyright information

© International Society of Oncology and BioMarkers (ISOBM) 2015

Authors and Affiliations

  • Fardous F. El-Senduny
    • 1
    • 3
  • Farid A. Badria
    • 2
  • Ahmed M. EL-Waseef
    • 3
  • Subhash C. Chauhan
    • 4
  • Fathi Halaweish
    • 1
    Email author
  1. 1.Department of Chemistry & BiochemistrySouth Dakota State UniversityBrookingsUSA
  2. 2.Pharmacognosy Department, Faculty of PharmacyMansoura UniversityMansouraEgypt
  3. 3.Chemistry Department, Faculty of ScienceMansoura UniversityMansouraEgypt
  4. 4.Department of Pharmaceutical Sciences and Center for Cancer ResearchUniversity of Tennessee Health Science CenterMemphisUSA

Personalised recommendations