Apigenin Sensitizes Huh-7 Human Hepatocellular Carcinoma Cells to TRAIL-induced Apoptosis

  • Kim, Eun-Young (College of Pharmacy, Sookmyung Women's University) ;
  • Kim, An-Keun (College of Pharmacy, Sookmyung Women's University)
  • Received : 2011.10.18
  • Accepted : 2011.11.29
  • Published : 2012.01.31


TNF-related apoptosis-inducing ligand (TRAIL) is a promising agent for management of cancer because of its selective cytotoxicity to cancer cells. However, some cancer cells have resistance to TRAIL. Accordingly, novel treatment strategies are required to overcome TRAIL resistance. Here, we examined the synergistic apoptotic effect of apigenin in combination with TRAIL in Huh-7 cells. We found that combined treatment of TRAIL and apigenin markedly inhibited Huh-7 cell growth compared to either agent alone by inducing apoptosis. Combined treatment with apigenin and TRAIL induced chromatin condensation and the cleavage of poly (ADP-ribose) polymerase (PARP). In addition, enhanced apoptosis by TRAIL/apigenin combination was quantified by annexin V/PI flow cytometry analysis. Western blot analysis suggested that apigenin sensitizes cells to TRAIL-induced apoptosis by activating both intrinsic and extrinsic apoptotic pathway-related caspases. The augmented apoptotic effect by TRAIL/apigenin combination was accompanied by triggering mitochondria-dependent signaling pathway, as indicated by Bax/Bcl-2 ratio up-regulation. Our results demonstrate that combination of TRAIL and apigenin facilitates apoptosis in Huh-7 cells.


  1. Adams, J. M. and Cory, S. (2007) The Bcl-2 apoptotic switch in cancer development and therapy. Oncogene. 26, 1324-1337.
  2. Andrzejewski, T., Deeb, D., Gao, X., Danyluk, A., Arbab, A. S., Dulchavsky, S. A. and Gautam, S. C. (2008) Therapeutic effi cacy of curcumin/TRAIL combination regimen for hormone-refractory prostate cancer. Oncol. Res. 17, 257-267.
  3. Armeanu, S., Lauer, U. M., Smirnow, I., Schenk, M., Weiss, T. S., Gregor, M. and Bitzer, M. (2003) Adenoviral gene transfer of tumor necrosis factor-related apoptosis-inducing ligand overcomes an impaired response of hepatoma cells but causes severe apoptosis in primary human hepatocytes. Cancer Res. 63, 2369-2372.
  4. Ashkenazi A, Pai RC, Fong S, Leung S, Lawrence DA, Marsters SA, Blackie C, Chang, L., McMurtrey, A. E., Hebert, A., DeForge, L., Koumenis, I. L., Lewis, D., Harris, L., Bussiere, J., Koeppen, H., Shahrokh, Z. and Schwall, R. H. (1999) Safety and antitumor activity of recombinant soluble Apo2 ligand. J. Clin. Invest. 104, 155-162.
  5. Bhojani, M. S., Rossú, B. D. and Rehemtulla, A. (2003) TRAIL and anti-tumor responses. Cancer Biol. Ther. 2 (4 Suppl 1), S71-S78.
  6. Birt, D. F., Mitchell, D., Gold, B., Pour, P. and Pinch, H. C. (1997) Inhibition of ultraviolet light induced skin carcinogenesis in SKH-1 mice by apigenin, a plant flavonoid. Anticancer. Res. 17, 85-91.
  7. Chiang, L. C., Ng, L. T., Lin, I. C., Kuo, P. L. and Lin, C. C. (2006) Antiproliferative effect of apigenin and its apoptotic induction in human Hep G2 cells. Cancer Lett. 237, 207-214.
  8. Cohen, G. M. (1997) Caspases: the executioners of apoptosis. Biochem. J. 326, 1-16.
  9. Deeb, D., Jiang, H., Gao, X., Hafner, M. S, Wong, H., Divine, G., Chapman, R. A., Dulchavsky, S. A. and Gautam, S. C. (2004) Curcumin sensitizes prostate cancer cells to tumor necrosis factor-related apoptosis-inducing ligand/Apo2L by inhibiting nuclear factor-${\kappa}B$through suppression of I${\kappa}B{\alpha}$ phosphorylation. Mol. Cancer Ther. 3, 803-812.
  10. Eggert, A., Grotzer, M. A., Zuzak, T. J., Wiewrodt, B. R., Ho, R., Ikegaki, N. and Brodeur, G. M. (2001) Resistance to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced apoptosis in neuroblastoma cells correlates with a loss of caspase-8 expression. Cancer Res. 61, 1314-1319.
  11. Gross, A., McDonnell, J. M. and Korsmeyer, S. J. (1999) BCL-2 family members and the mitochondria in apoptosis. Genes. Dev. 13, 1899-1911.
  12. Gupta, S., Afaq, F. and Mukhtar, H. (2001) Selective growth-inhibitory, cell-cycle deregulatory and apoptotic response of apigenin in normal versus human prostate carcinoma cells. Biochem. Biophys. Res. Commun. 287, 914-920.
  13. Hao, C., Song, J. H., Hsi, B., Lewis, J., Song, D. K., Petruk, K. C., Tyrrell, D. L. and Kneteman, N. M. (2004) TRAIL inhibits tumor growth but is nontoxic to human hepatocytes in chimeric mice. Cancer Res. 64, 8502-8506.
  14. Higashitsuji, H., Higashitsuji, H., Masuda, T., Liu, Y., Itoh, K. and Fujita, J. (2007) Enhanced deacetylation of p53 by the anti-apoptotic protein HSCO in association with histone deacetylase 1. J. Biol. Chem. 282, 13716-13725.
  15. Hinz, S., Trauzold, A., Boenicke, L., Sandberg, C., Beckmann, S., Bayer, E., Walczak, H., Kalthoff, H. and Ungefroren, H. (2000) Bcl-XL protects pancreatic adenocarcinoma cells against CD95- and TRAIL-receptor-mediated apoptosis. Oncogene. 19, 5477-5486.
  16. Jacquemin, G., Shirley, S. and Micheau, O. (2010) Combining naturally occurring polyphenols with TNF-related apoptosis-inducing ligand: a promising approach to kill resistant cancer cells? Cell Mol. Life Sci. 67, 3115-3130.
  17. Johnstone, R. W., Ruefl i, A. A. and Lowe, S. W. (2002) Apoptosis: a link between cancer genetics and chemotherapy. Cell. 108, 153-164.
  18. Jung, E. M., Lim, J. H., Lee, T. J., Park, J. W., Choi, K. S. and Kwon, T. K. (2005) Curcumin sensitizes tumor necrosis factor-related apoptosis- inducing ligand (TRAIL)-induced apoptosis through reactive oxygen species-mediated upregulation of death receptor 5 (DR5). Carcinogenesis. 26, 1905-1913.
  19. Kim, D. S., Jeong, Y. M., Moon, S. I., Kim, S. Y., Kwon, S. B., Park, E. S., Youn, S. W. and Park, K. C. (2006) Indole-3-carbinol enhances ultraviolet B-induced apoptosis by sensitizing human melanoma cells. Cell Mol. Life Sci. 63, 2661-2668.
  20. Koschny, R., Ganten, T. M., Sykora, J., Haas, T. L., Sprick, M. R., Kolb, A., Stremmel, W. and Walczak, H. (2007) TRAIL/bortezomib cotreatment is potentially hepatotoxic but induces cancer-specifi c apoptosis within a therapeutic window. Hepatology. 45, 649-658.
  21. Kruyt, F. A. (2008) TRAIL and cancer therapy. Cancer Lett. 263, 14-25.
  22. Lawrence, D., Shahrokh, Z., Marsters, S., Achilles K,, Shih, D., Mounho, B., Hillan, K., Totpal, K., DeForge, L., Schow, P., Hooley, J., Sherwood, S., Pai, R., Leung, S., Khan, L., Gliniak, B., Bussiere, J., Smith, C. A., Strom, S. S., Kelley, S., Fox, J. A., Thomas, D. and Ashkenazi, A. (2001) Differential hepatocyte toxicity of recombinant Apo2L/TRAIL versions. Nat. Med. 7, 383-385.
  23. Moll, U. M. and Zaika, A. (2001) Nuclear and mitochondrial apoptotic pathways of p53. FEBS. Lett. 493, 65-69.
  24. Nitsch, R., Bechmann, I., Deisz, R. A., Haas, D., Lehmann, T. N., Wendling, U., and Zipp, F. (2000) Human brain-cell death induced by tumour-necrosis-factor-related apoptosis-inducing ligand (TRAIL). Lancet. 356, 827-828.
  25. Roth, W., Isenmann, S., Naumann, U., Kugler, S., Bähr, M., Dichgans, J., Ashkenazi, A. and Weller, M. (1999) Locoregional Apo2L/TRAIL eradicates intracranial human malignant glioma xenografts in athymic mice in the absence of neurotoxicity. Biochem. Biophys. Res. Commun. 265, 479-483.
  26. Siegelin, M. D., Reuss, D. E., Habel, A., Herold-Mende, C. and von Deimling, A. (2008) The flavonoid kaempferol sensitizes human glioma cells to TRAIL-mediated apoptosis by proteasomal degradation of survivin. Mol. Cancer Ther. 7, 3566-3574.
  27. Sun, X. M., MacFarlane, M., Zhuang, J., Wolf, B. B., Green, D. R. and Cohen, G. M. (1999) Distinct caspase cascades are initiated in receptor-mediated and chemical-induced apoptosis. J. Biol. Chem. 274, 5053-5060.
  28. Van Geelen, C. M., de Vries, E. G. and de Jong, S. (2004) Lessons from TRAIL-resistance mechanisms in colorectal cancer cells: paving the road to patient-tailored therapy. Drug Resist. Updat. 7, 345-358.
  29. Walczak, H., Miller, R. E., Ariail, K., Gliniak, B., Griffi th, T.S., Kubin, M., Chin, W., Jones J, Woodward, A., Le, T., Smith, C., Smolak, P., Goodwin, R. G., Rauch, C. T., Schuh J, C. and Lynch, D. H. (1999) Tumoricidal activity of tumor necrosis factor-related apoptosis-inducing ligand in vivo. Nat. Med. 5, 157-163.
  30. Wang, W., Heideman, L., Chung, C. S., Pelling, J. C., Koehler, K. J. and Birt, D. F. (2000) Cell-cycle arrest at G2/M and growth inhibition by apigenin in human colon carcinoma cell lines. Mol. Carcinog. 28, 102-110.<102::AID-MC6>3.0.CO;2-2
  31. Way, T. D., Kao, M. C. and Lin, J. K. (2003) Apigenin induces apoptosis through proteasomal degradation of HER2/neu in HER2/neuoverexpressing breast cancer cells via the phosphatidylinositol 3-kinase/Akt-dependent pathway. J. Biol. Chem. 279, 4479-4489.
  32. Wei, H., Tye, L., Bresnick, E. and Birt, D. F. (1990) Inhibitory effect of apigenin, a plant fl avonoid, on epidermal ornithine decarboxylase and skin tumor promotion in mice. Cancer Res. 50, 499-502.
  33. Wolf, B. B. and Green, D. R. (1999) Suicidal tendencies: apoptotic cell death by caspase family proteinases. J. Biol. Chem. 274, 20049-20052.
  34. Yano, Y., Hayashi, Y., Nakaji, M., Nagano, H., Seo, Y., Ninomiya, T., Yoon, S., Wada, A., Hirai, M., Kim, S. R., Yokozaki, H. and Kasuga, M. (2003) Different apoptotic regulation of TRAIL-caspase pathway in HBV- and HCV-related hepatocellular carcinoma. Int. J. Mol. Med. 11, 499-504.
  35. Yoshida, T., Konishi, M., Horinaka, M., Yasuda, T., Goda, A. E., Taniguchi, H., Yano, K., Wakada, M. and Sakai, T. (2008) Kaempferol sensitizes colon cancer cells to TRAIL-induced apoptosis. Biochem. Biophys. Res. Commun. 375, 129-133.
  36. Zhang, L. and Fang, B. (2005) Mechanisms of resistance to TRAIL-induced apoptosis in cancer. Cancer Gene Ther. 12, 228-237.
  37. Zhang, Y. and Zhang, B. (2008) TRAIL resistance of breast cancer cells is associated with constitutive endocytosis of death receptors 4 and 5. Mol. Cancer Res. 6, 1861-1871.

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