Tumor Biology

, Volume 36, Issue 4, pp 2747–2761 | Cite as

(-)-Epigallocatechin-3-gallate inhibits nasopharyngeal cancer stem cell self-renewal and migration and reverses the epithelial–mesenchymal transition via NF-κB p65 inactivation

  • Ya-Jun Li
  • Shun-Long Wu
  • Song-Mei Lu
  • Fang Chen
  • Ying Guo
  • Sheng-Min Gan
  • Yan-Long Shi
  • Shuang Liu
  • Shao-Lin Li
Research Article


The cancer stem cell (CSC) theory states that many types of cancer, including nasopharyngeal cancer (NPC), are initiated from and maintained by CSCs, which may be responsible for tumor relapse and resistance to therapy. It is imperative that nasopharyngeal cancer stem cells (NPCSCs) be specifically targeted to eradicate NPC and prevent recurrence. Epigallocatechin-3-gallate (EGCG) inhibits cancer progression by attenuating NF-κB p65 activity, which is upregulated in CSCs and plays an important role in epithelial–mesenchymal transition (EMT). The purpose of this study is to confirm the self-renewal and migration inhibitory effects of EGCG toward NPCSCs and to clarify its mechanism of activity. We enriched and characterized NPCSCs by collecting spheroid-derived cells grown in serum-free medium (SFM) and examined the effects of EGCG on the characteristics of NPCSCs and studied the underlying mechanisms using soft agar colony assays, transwell migration assays, reverse transcriptase polymerase chain reaction (RT-PCR), Western blot analysis, immunofluorescence staining, and xenograft studies. NPC spheroids enriched from NPC cell lines acquired CSC traits and underwent EMT. EGCG inhibited the NPCSCs’ self-renewal and migration and reversed EMT, and combined treatment with EGCG and cisplatin reduced the growth of CSC tumor xenografts. Moreover, EGCG inhibited NF-κB p65 activity by modulating the cellular localization of p65 and decreasing the transcriptional regulation of NF-κB p65 on Twist1 expression. NF-κB p65 is a novel therapeutic target in NPCSCs, and the inhibition of activated NF-κB p65 in CSCs by EGCG may offer an effective treatment for NPC.


Nasopharyngeal cancer Cancer stem cells Epigallocatechin-3-gallate Epithelial–mesenchymal transition Self-renewal and migration NF-κB p65 



Cancer stem cell


Nasopharyngeal cancer


Nasopharyngeal cancer stem cells


Spheroid cell


Epithelial–mesenchymal transition




Serum-free medium


Nuclear factor κB


Quantitative reverse transcriptase polymerase chain reaction




Fluorescence-activated cell sorting


Fetal bovine serum


Epidermal growth factor


Human recombinant basic fibroblast growth factor


Dulbecco’s modified Eagle’s medium F12



This research is supported by the General Program of the National Natural Science Foundation of China (No. 81171365).

Conflicts of interest



  1. 1.
    Yu MC, Yuan JM. Epidemiology of nasopharyngeal carcinoma. Semin Cancer Biol. 2002;12:421–9.CrossRefPubMedGoogle Scholar
  2. 2.
    Lee AW, Yau TK, Wong DH, Chan EW, Yeung RM, Ng WT, et al. Treatment of stage IV(A-B)nasopharyngeal carcinoma by induction-concurrent chemoradiotherapy and accelerated fractionation. Int J Radiat Oncol Biol Phys. 2005;63:1331–8.CrossRefPubMedGoogle Scholar
  3. 3.
    Yeh SA, Tang Y, Lui CC, Huang YJ, Huang EY. Treatment outcomes and late complications of 849 patients with nasopharyngeal carcinoma treated with radiotherapy alone. Int J Radiat Oncol Biol Phys. 2005;62:672–9.CrossRefPubMedGoogle Scholar
  4. 4.
    Lo KW, To KF, Huang DP. Focus on nasopharyngeal carcinoma. Cancer Cell. 2004;5:423–8.CrossRefPubMedGoogle Scholar
  5. 5.
    Le QT, Tate D, Koong A, Gibbs IC, Chang SD, Adler JR, et al. Improved local control with stereotactic radiosurgical boost in patients with nasopharyngeal carcinoma. Int J Radiat Oncol Biol Phys. 2003;56:1046–54.CrossRefPubMedGoogle Scholar
  6. 6.
    Visvader JE, Lindeman GJ. Cancer stem cells in solid tumours: accumulating evidence and unresolved questions. Nat Rev Cancer. 2008;8:755–68.CrossRefPubMedGoogle Scholar
  7. 7.
    Er O. Cancer stem cells in solid tumors. Onkol. 2009;32:605–9.CrossRefGoogle Scholar
  8. 8.
    Lang SH, Anderson E, Fordham R, Collins AT. Modeling the prostate stem cell niche: an evaluation of stem cell survival and expansion in vitro. Stem Cells Dev. 2010;19:537–46.CrossRefPubMedPubMedCentralGoogle Scholar
  9. 9.
    Jones RJ, Matsui WH, Smith BD. Cancer stem cells: are we missing the target? J Natl Cancer Inst. 2004;96:583–5.CrossRefPubMedGoogle Scholar
  10. 10.
    Sakariassen PO, Immervoll H, Chekenya M. Cancer stem cells as mediators of treatment resistance in brain tumors: status and controversies. Neoplasia. 2007;9:882–92.CrossRefPubMedPubMedCentralGoogle Scholar
  11. 11.
    Su J, Xu XH, Huang Q, Lu MQ, Li DJ, Xue F, et al. Identification of cancer stem-like CD44+ cells in human nasopharyngeal carcinoma cell line. Arch Med Res. 2011;42:15–21.CrossRefPubMedGoogle Scholar
  12. 12.
    Wang J, Guo LP, Chen LZ, Zeng YX, Lu SH. Identification of cancer stem cell-like side population cells in human nasopharyngeal carcinoma cell line. Cancer Res. 2007;67:3716–24.CrossRefPubMedGoogle Scholar
  13. 13.
    Kong D, Banerjee S, Ahmad A, Li Y, Wang Z, Sethi S, et al. Epithelial to mesenchymal transition is mechanistically linked with stem cell signatures in prostate cancer cells. PLoS One. 2010;5:e12445.CrossRefPubMedPubMedCentralGoogle Scholar
  14. 14.
    Polyak K, Weinberg RA. Transitions between epithelial and mesenchymal states: acquisition of malignant and stem cell traits. Nat Rev Cancer. 2009;9:265–73.CrossRefPubMedGoogle Scholar
  15. 15.
    Shankar S, Ganapathy S, Srivastava RK. Green tea polyphenols: biology and therapeutic implications in cancer. Front Biosci. 2007;12:4881–99.CrossRefPubMedGoogle Scholar
  16. 16.
    Yang CS, Wang X, Lu G, Picinich SC. Cancer prevention by tea: animal studies, molecular mechanisms and human relevance. Nat Rev Cancer. 2009;9:429–39.CrossRefPubMedPubMedCentralGoogle Scholar
  17. 17.
    Ju J, Lu G, Lambert JD, Yang CS. Inhibition of carcinogenesis by tea constituents. Semin Cancer Biol. 2007;17:395–402.CrossRefPubMedPubMedCentralGoogle Scholar
  18. 18.
    Landis-Piwowar KR, Huo C, Chen D, Milacic V, Shi G, Chan TH, et al. A novel prodrug of the green tea polyphenol (-)-epigallocatechin-3-gallate as a potential anticancer agent. Cancer Res. 2007;67:4303–10.CrossRefPubMedGoogle Scholar
  19. 19.
    Shankar S, Ganapathy S, Hingorani SR, Srivastava RK. EGCG inhibits growth, invasion, angiogenesis and metastasis of pancreatic cancer. Front Biosci. 2008;13:440–52.CrossRefPubMedGoogle Scholar
  20. 20.
    Shirakami Y, Shimizu M, Adachi S, Sakai H, Nakagawa T, Yasuda Y, et al. (-)-Epigallocatechin gallate suppresses the growth of human hepatocellular carcinoma cells by inhibiting activation of the vascular endothelial growth factor-vascular endothelial growth factor receptor axis. Cancer Sci. 2009;100:1957–62.CrossRefPubMedGoogle Scholar
  21. 21.
    Takahashi H, Nomata K, Mori K, Matsuo M, Miyaguchi T, Noguchi M, et al. The preventive effect of green tea on the gap junction intercellular communication in renal epithelial cells treated with a renal carcinogen. Anticancer Res. 2004;24:3757–62.PubMedGoogle Scholar
  22. 22.
    Tang GQ, Yan TQ, Guo W, Ren TT, Peng CL, Zhao H, et al. (-)-Epigallocatechin-3-gallate induces apoptosis and suppresses proliferation by inhibiting the human Indian Hedgehog pathway in human chondrosarcoma cells. J Cancer Res Clin Oncol. 2010;136:1179–85.CrossRefPubMedGoogle Scholar
  23. 23.
    Zhu BH, Chen HY, Zhan WH, Wang CY, Cai SR, Wang Z, et al. (-)-Epigallocatechin-3-gallate inhibits VEGF expression induced by IL-6 via Stat3 in gastric cancer. World J Gastroenterol. 2011;17:2315–25.CrossRefPubMedPubMedCentralGoogle Scholar
  24. 24.
    Nishimura N, Hartomo TB, Pham TV, Lee MJ, Yamamoto T, Morikawa S, et al. Epigallocatechin gallate inhibits sphere formation of neuroblastoma BE(2)-C cells. Environ Health Prev Med. 2012;17:246–51.CrossRefPubMedGoogle Scholar
  25. 25.
    Chen D, Pamu S, Cui Q, Chan TH, Dou QP. Novel epigallocatechin gallate (EGCG) analogs activate AMP-activated protein kinase pathway and target cancer stem cells. Bioorg Med Chem. 2012;20:3031–7.CrossRefPubMedPubMedCentralGoogle Scholar
  26. 26.
    Tang SN, Fu J, Nall D, Rodova M, Shankar S, Srivastava RK. Inhibition of sonic Hedgehog pathway and pluripotency maintaining factors regulate human pancreatic cancer stem cell characteristics. Int J Cancer. 2012;131:30–40.CrossRefPubMedGoogle Scholar
  27. 27.
    Burnett J, Newman B, Sun D. Targeting cancer stem cells with natural products. Curr Drug Targets. 2012;13:1054–64.CrossRefPubMedGoogle Scholar
  28. 28.
    Hayden MS, Ghosh S. Shared principles in NF-kappa B signaling. Cell. 2008;132:344–62.CrossRefPubMedGoogle Scholar
  29. 29.
    Yan Z, Yong-Guang T, Fei-Jun L, Fa-Qing T, Min T, Ya C. Interference effect of epigallocatechin-3-gallate on targets of nuclear factor kB signal transduction pathways activated by EB virus encoded latent membrane protein 1. IntJ Biochem Cell Biol. 2004;36:1473–81.Google Scholar
  30. 30.
    Uchibori R, Tsukahara T, Mizuguchi H, Saga Y, Urabe M, Mizukami H, et al. NF-κB activity regulates mesenchymal stem cell accumulation at tumor sites. Cancer Res. 2013;73:364–72.CrossRefPubMedGoogle Scholar
  31. 31.
    Liu M, Sakamaki T, Casimiro MC, Willmarth NE, Quong AA, Ju X, et al. The canonical NF-kappaB pathway governs mammary tumorigenesis in transgenic mice and tumor stem cell expansion. Cancer Res. 2010;70:10464–73.CrossRefPubMedPubMedCentralGoogle Scholar
  32. 32.
    Afaq F, Adhami VM, Ahmad N, Mukhtar H. Inhibition of ultraviolet B-mediated activation of nuclear factor kappaB in normal human epidermal keratinocytes by green tea constituent (-)-epigallocatechin-3-gallate. Oncogene. 2003;22:1035–44.CrossRefPubMedGoogle Scholar
  33. 33.
    Gupta S, Hastak K, Afaq F, Ahmad N, Mukhtar H. Essential role of caspases in epigallocatechin-3-gallate-mediated inhibition of nuclear factor kappa B and induction of apoptosis. Oncogene. 2004;23:2507–22.CrossRefPubMedGoogle Scholar
  34. 34.
    Syed DN, Afaq F, Kweon MH, Hadi N, Bhatia N, Spiegelman VS, et al. Green tea polyphenol EGCG suppresses cigarette smoke condensate-induced NF-kappaB activation in normal human bronchial epithelial cells. Oncogene. 2007;26:673–82.CrossRefPubMedGoogle Scholar
  35. 35.
    Lee J, Kotliarova S, Kotliarov Y, Li A, Su Q, Donin NM, et al. Tumor stem cells derived from glioblastomas cultured in bFGF and EGF more closely mirror the phenotype and genotype of primary tumors than do serum-cultured cell lines. Cancer Cell. 2006;9:391–403.CrossRefPubMedGoogle Scholar
  36. 36.
    Li CW, Xia W, Huo L, Lim SO, Wu Y, Hsu JL, et al. Epithelial-mesenchymal transition induced by TNF-alpha requires NF-kappaB-mediated transcriptional upregulation of Twist1. Cancer Res. 2012;72:1290–300.CrossRefPubMedPubMedCentralGoogle Scholar
  37. 37.
    Pham CG, Bubici C, Zazzeroni F, Knabb JR, Papa S, Kuntzen C, et al. Upregulation of twist-1 by NF-kappaB blocks cytotoxicity induced by chemotherapeutic drugs. Mol Cell Biol. 2007;27:3920–35.CrossRefPubMedPubMedCentralGoogle Scholar
  38. 38.
    Frame FM, Maitland NJ. Cancer stem cells, models of study and implications of therapy resistance mechanisms. Adv Exp Med Biol. 2011;720:105–18.CrossRefPubMedGoogle Scholar
  39. 39.
    Mani SA, Guo W, Liao MJ, Eaton EN, Ayyanan A, Zhou AY, et al. The epithelial-mesenchymal transition generates cells with properties of stem cells. Cell. 2008;133:704–15.CrossRefPubMedPubMedCentralGoogle Scholar
  40. 40.
    Hu Y, Fu L. Targeting cancer stem cells: a new therapy to cure cancer patients. Am J Cancer Res. 2012;2:340–56.PubMedPubMedCentralGoogle Scholar
  41. 41.
    Na HK, Surh YJ. Intracellular signaling network as a prime chemopreventive target of (-)-epigallocatechin gallate. Mol Nutr Food Res. 2006;50:152–9.CrossRefPubMedGoogle Scholar
  42. 42.
    Jung YD, Ellis LM. Inhibition of tumour invasion and angiogenesis by epigallocatechin gallate (EGCG), a major component of green tea. Int J Exp Pathol. 2001;82:309–16.CrossRefPubMedPubMedCentralGoogle Scholar
  43. 43.
    Takebe N, Harris PJ, Warren RQ, Ivy SP. Targeting cancer stem cells by inhibiting Wnt, Notch, and Hedgehog pathways. Nat Rev Clin Oncol. 2011;8:97–106.CrossRefPubMedGoogle Scholar
  44. 44.
    Chen D, Pamu S, Cui Q, Chan TH, Dou QP. Novel epigallocatechin gallate (EGCG) analogs activate AMP-activated protein kinase pathway and target cancer stem cells. Bioorg Med Chem. 2012;20:3031–7.CrossRefPubMedPubMedCentralGoogle Scholar
  45. 45.
    Zhou J, Zhang H, Gu P, Bai J, Margolick JB, Zhang Y. NF-kappaB pathway inhibitors preferentially inhibit breast cancer stem-like cells. Breast Cancer Res Treat. 2008;111:419–27.CrossRefPubMedGoogle Scholar
  46. 46.
    Shimizu M, Deguchi A, Lim JT, Moriwaki H, Kopelovich L, Weinstein IB. (-)-Epigallocatechin gallate and polyphenon E inhibit growth and activation of the epidermal growth factor receptor and human epidermal growth factor receptor-2 signaling pathways in human colon cancer cells. Clin Cancer Res. 2005;11:2735–46.CrossRefPubMedGoogle Scholar
  47. 47.
    Sasaki CY, Barberi TJ, Ghosh P, Longo DL. Phosphorylation of RelA/p65 on serine 536 defines an I{kappa}B{alpha}-independent NF-{kappa}B pathway. J Biol Chem. 2005;280:34538–47.CrossRefPubMedGoogle Scholar
  48. 48.
    Huber MA, Beug H, Wirth T. Epithelial-mesenchymal transition: NF-kappaB takes center stage. Cell Cycle. 2004;3:1477–80.CrossRefPubMedGoogle Scholar
  49. 49.
    Huber MA, Azoitei N, Baumann B, Grünert S, Sommer A, Pehamberger H, et al. NF-Kb is essential for epithelial-mesenchymal transition and metastasis in a model of breast cancer progression. J Clin Invest. 2004;114:569–81.CrossRefPubMedPubMedCentralGoogle Scholar
  50. 50.
    Min C, Eddy SF, Sherr DH, Sonenshein GE. NF-kappaB and epithelial to mesenchymal transition of cancer. J Cell Biochem. 2008;104:733–44.CrossRefPubMedGoogle Scholar
  51. 51.
    Li S, Kendall SE, Raices R, Finlay J, Covarrubias M, Liu Z, et al. TWIST1 associates with NF-κB subunit RELA via carboxyl-terminal WR domain to promote cell autonomous invasion through IL8 production. BMC Biol. 2012;10:73.CrossRefPubMedPubMedCentralGoogle Scholar
  52. 52.
    Do SI, Kim JY, Kang SY, Lee JJ, Lee JE, Nam SJ, et al. Expression of TWIST1, Snail, Slug, and NF-κB and methylation of the TWIST1 promoter in mammary phyllodes tumor. Tumor Biol. 2013;34:445–53.CrossRefGoogle Scholar
  53. 53.
    Yang MH, Hsu DS, Wang HW, Wang HJ, Lan HY, Yang WH, et al. Bmi1 is essential in Twist1-induced epithelial–mesenchymal transition. Nat Cell Biol. 2010;12:982–92.CrossRefPubMedGoogle Scholar
  54. 54.
    Chiba T, Miyagi S, Saraya A, Aoki R, Seki A, Morita Y, et al. The polycomb gene product BMI1 contributes to the maintenance of tumor-initiating side population cells in hepatocellular carcinoma. Cancer Res. 2008;68:7742–9.CrossRefPubMedGoogle Scholar

Copyright information

© International Society of Oncology and BioMarkers (ISOBM) 2014

Authors and Affiliations

  • Ya-Jun Li
    • 1
  • Shun-Long Wu
    • 1
  • Song-Mei Lu
    • 1
  • Fang Chen
    • 2
  • Ying Guo
    • 3
  • Sheng-Min Gan
    • 1
  • Yan-Long Shi
    • 1
  • Shuang Liu
    • 1
  • Shao-Lin Li
    • 1
  1. 1.Department of Radiology, College of Basic MedicineChongqing Medical UniversityChongqingChina
  2. 2.Department of OncologyAffiliated Hospital of Zunyi Medical CollegeZunyiChina
  3. 3.Department of OncologyAffiliated Hospital of Luzhou Medical CollegeLuzhouChina

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