DOI QR코드

DOI QR Code

Harmal Extract Induces Apoptosis of HCT116 Human Colon Cancer Cells, Mediated by Inhibition of Nuclear Factor-κB and Activator Protein-1 Signaling Pathways and Induction of Cytoprotective Genes

  • Elkady, Ayman I (Department of Biological Sciences, Faculty of Sciences, King Abdulaziz University) ;
  • Hussein, Rania A (Department of Clinical Nutrition, Faculty of Applied Medical Sciences, King Abdulaziz University) ;
  • El-Assouli, Sufian M (Department of Biological Sciences, Faculty of Sciences, King Abdulaziz University)
  • Published : 2016.06.01

Abstract

Background: Colorectal cancer (CRC) is a major cause of morbidity and mortality, being the second most common type of cancer worldwide in both men and women. It accounts yearly for approximately 9% of all new cases of cancers. Furthermore, the current chemotherapeutic regimens seem unsatisfactory, so that exploration of novel therapeutic modalities is needed. The present study was undertaken to investigate the inhibitory effects of a crude alkaloid extract (CAERS) of a medicinal herb, Rhazya stricta, on proliferation of CRC HCT116 cells and to elucidate mechanisms of action. To achieve these aims, we utilized MTT, comet, DNA laddering and gene reporter assays, along with Western blot and RT-PCR analyses. Results: We found that CAERS inhibited cell proliferation and induced apoptotic cell death in HCT116 cells. Hallmarks of morphological and biochemical signs of apoptosis were clearly evident. CAERS down-regulated DNA-binding and transcriptional activities of NF-${\kappa}B$ and AP-1 proteins, while up-regulating expression of the Nrf-2 protein. It also down-regulated expression levels of the ERK MAPK, Bcl-2, cyclin D1, CDK-4, survivin and VEGF and up-regulated levels of Bax, caspase-3/7 and -9, p53, p21, Nrf-2. Markedly, it promoted mRNA expression levels of cytoprotective genes including the hemeoxygenase-1, NAD(P)H quinine oxidoreductase 1 and UDP-glucuronyltransferase. Conclusions: These findings indicate that CAERS exerts antiproliferative action on CRC cells through induction of apoptotic mechanisms, and suggest CAERS could be a promising agent for studying and developing novel chemotherapeutic agents aimed at novel molecular targets for the treatment of CRC.

Keywords

Apoptosis;oncogenes;cytoprotective genes;transfection;DNA damage;caspase-3/7

Acknowledgement

Supported by : King Abdulaziz City for Science and Technology (KACST)

References

  1. Aggarwal BB, Ichikawa H (2005). Molecular targets and anticancer potential of indole-3-carbinol and its derivatives. Cell Cycle, 4, 1201-15. https://doi.org/10.4161/cc.4.9.1993
  2. Altieri DC (2003). Validating survivin as a cancer therapeutic target" Nat Rev Cancer, 3, 46-54. https://doi.org/10.1038/nrc968
  3. Anisz T (2007). Alkaloids: Secrets of Life Alkaloid Chemistry, Biological Significance, Applications and Ecological Role. Elsevier Amsterdam, The Netherlands
  4. Baeshen NA, Elkady AI, Abuzinadah OA, Mutwakil MH (2012). Potential anticancer activity of the medicinal herb, Rhazya stricta, against human breast cancer. African J Biotechnol, 11, 8960-72. https://doi.org/10.5897/AJB12.570
  5. Ban JO, Yuk DY, Woo KS, et al (2007). Inhibition of cell growth and induction of apoptosis via inactivation of NF-${\kappa}B$ by a sulfur compound isolated from garlic in human colon cancer cells. J Pharmacol Sci, 104, 374-83. https://doi.org/10.1254/jphs.FP0070789
  6. Bat-Chen W, Golan T, Peri I, Ludmer Z, Schwartz B (2010). Allicin purified from fresh garlic cloves induces apoptosis in colon cancer cells via Nrf2. Nutr Cancer, 62, 947-57. https://doi.org/10.1080/01635581.2010.509837
  7. Chidambara MKN, Jayaprakasha GK, Patil BS (2012). The natural alkaloid berberine targets multiple pathways to induce cell death in cultured human colon cancer cells. Eur J Pharmacol, 688, 14-21. https://doi.org/10.1016/j.ejphar.2012.05.004
  8. Chiou YS, Ma NJ, Sang S, Ho CT, Wang YJ, Pan MH (2012). Peracetylated (-)-epigallocatechin-3-gallate (AcEGCG) potently suppresses dextran sulfate sodium-induced colitis and colon tumorigenesis in mice. J Agric Food Chem, 60, 3441-51. https://doi.org/10.1021/jf300441p
  9. Debruyne PR, Bruyneel EA, Li X, et al (2001). The role of bile acids in carcinogenesis. Mutat Res, 480-481, 359-69. https://doi.org/10.1016/S0027-5107(01)00195-6
  10. Dunkern TR, Fritz G, Kaina B (2001). Ultraviolet light-induced DNA damage triggers apoptosis in nucleotide excision repair-deficient cells via Bcl-2 decline and caspase-3/-8 activation. Oncogene, 20, 6026-38. https://doi.org/10.1038/sj.onc.1204754
  11. El Gendy MA, Ali BH, Michail K, Siraki AG, El-Kadi AO (2012). Induction of quinone oxidoreductase 1 Enzyme by Rhazya stricta through Nrf2-dependent mechanism. J Ethnopharmacol, 144, 416-24. https://doi.org/10.1016/j.jep.2012.09.032
  12. Elkady AI (2013). Crude alkaloid extract of Rhazya stricta inhibits cell growth and sensitizes human lung cancer cells to cisplatin through induction of apoptosis. Genet Mol Biol, 36, 12-21. https://doi.org/10.1590/S1415-47572013005000009
  13. Elkady AI, Hussein RA, Abu-Zinadah AO (2014). Differential Control of Growth, Apoptotic Activity and Gene Expression in Human Colon Cancer Cells by Extracts Derived from Medicinal Herbs, Rhazya stricta and Zingiber officinale and Their Combination. World J Gastroenterol, 20, 15275-88. https://doi.org/10.3748/wjg.v20.i41.15275
  14. Elkady AI, El-Assouli SM, Hussein RAH (2015). Mechanism of Action of (Nigella sativa) on Human Colon Cancer Cells: the Suppression of AP-1 And NF-${\kappa}B$ Transcription Factors and the Induction of Cytoprotective Genes. Asian Pac J Cancer Prev, 16, 7943-57. https://doi.org/10.7314/APJCP.2015.16.17.7943
  15. Eng C (2010). The evolving role of monoclonal antibodies in colorectal cancer: early presumptions and impact on clinical trial development. Oncologist, 15, 73-84.
  16. Fang JY, Richardson BC (2005). The MAPK signaling pathways and colorectal cancer. Lancet Oncol, 6, 322-7. https://doi.org/10.1016/S1470-2045(05)70168-6
  17. Gilani SA, Kikuchi A, Shinwari ZK, Khattak ZI, Watanabe KN (2007). Phytochemical, pharmacological and ethnobotanical studies of Rhazya stricta Decne. Phytother Res, 21, 301-7. https://doi.org/10.1002/ptr.2064
  18. Gopalakrishnan A, Kong NT (2008). Anticarcinogenesis by dietary phytochemicals: Cytoprotection by Nrf2 in normal cells and cytotoxicity by modulation of transcription factors NF-${\kappa}B$ and AP-1 in abnormal cancer cells. Food Chemical Toxicol, 46, 1257-70. https://doi.org/10.1016/j.fct.2007.09.082
  19. Hanahan D, Weinberg RA (2011). Hallmarks of Cancer: The Next Generation. Cell, 144, 646-74. https://doi.org/10.1016/j.cell.2011.02.013
  20. Hardwick JC, van den Brink GR, Offerhaus GJ, van Deventer SJ, Peppelenbosch MP (2001). NF-${\kappa}B$, p38 MAPK and JNK are highly expressed and active in the stroma of human colonic adenomatous polyps. Oncogene, 20, 819-27. https://doi.org/10.1038/sj.onc.1204162
  21. Harun Z, Ghazali AR (2012). Potential chemoprevention activity of Pterostilbene by enhancing the detoxifying enzymes in the HT-29 Cell Line. Asian Pac J Cancer Prev, 13, 6403-7. https://doi.org/10.7314/APJCP.2012.13.12.6403
  22. Hayes JD, McMahon M, Chowdhry S, Dinkova-Kostova AT (2010). Cancer chemoprevention mechanisms mediated through the Keap1-Nrf2 pathway. Antioxid Redox Signal, 13, 1713-48. https://doi.org/10.1089/ars.2010.3221
  23. Hess J, Angel P, Schorpp-Kistner M (2004). AP-1 subunits: quarrel and harmony among siblings. J Cell Science, 117, 5965-73. https://doi.org/10.1242/jcs.01589
  24. Hu R, Chen N, Yao J, Zhao Q, Zhang F, Li ZY, You QD, Guo QL (2012). The role of Nrf2 and apoptotic signaling pathways in oroxylin A-mediated responses in HCT-116 colorectal adenocarcinoma cells and xenograft tumors. Anticancer Drugs, 23, 651-8. https://doi.org/10.1097/CAD.0b013e3283512703
  25. Ichikawa M, Sowa Y, Iizumi Y, Aono Y, Sakai T (2015). Resibufogenin Induces G1-Phase Arrest through the Proteasomal Degradation of Cyclin D1 in Human Malignant Tumor Cells. PLoS One, 10.
  26. Justin A Call, S Gail Eckhardt, D Ross Camidge (2008). Targeted manipulation of apoptosis in cancer treatment. Lancet Oncol, 9, 1002-11. https://doi.org/10.1016/S1470-2045(08)70209-2
  27. Khan N, Afaq F, Mukhtar H (2007). Apoptosis by dietary factors: the suicide solution for delaying cancer grow. Carcinogenesis, 28, 233-9.
  28. Kim MK, Kang YJ, Kim DH, et al (2014). A novel hydroxamic acid derivative, MHY218, induces apoptosis and cell cycle arrest through downregulation of NF-${\kappa}B$ in HCT116 human colon cancer cells. Int J Oncol, 44, 256-64. https://doi.org/10.3892/ijo.2013.2163
  29. Kim PJ, Plescia J, Clevers H, Fearon ER, Altieri DC (2003). Survivin and molecular pathogenesis of colorectal cancer. Lancet, 362, 205-9. https://doi.org/10.1016/S0140-6736(03)13910-4
  30. Kumar M1, Sharma VL, Sehgal A, Jain M (2012). Protective effects of green and white tea against benzo(a)pyrene induced oxidative stress and DNA damage in murine model. Nutr Cancer, 64, 300-6. https://doi.org/10.1080/01635581.2012.648300
  31. Kundu T1, Bhattacharya RK, Siddiqi M, Roy M (2005). Correlation of apoptosis with comet formation induced by tea polyphenols in human leukemia cells. J Environ Pathol Toxicol Oncol, 24, 115-28. https://doi.org/10.1615/JEnvPathToxOncol.v24.i2.50
  32. Kokileva L (1994). Multi-step chromatin degradation in apoptosis. DNA breakdown in apoptosis. Int Arch Allergy Immunol, 105, 339-43. https://doi.org/10.1159/000236779
  33. Lee J-M, Johnson JA (2004). An important role of Nrf2-ARE pathway in the cellular defense mechanism, J Biochem Molecular Biol, 37, 139-43. https://doi.org/10.5483/BMBRep.2004.37.2.139
  34. Levin B, Lieberman DA, McFarland B, et al (2008). Screening and surveillance for the early detection of colorectal cancer and adenomatous polyps, 2008: a joint guideline from the american cancer society, the us multi-society task force on colorectal cancer, and the American College of Radiology. Gastroenterol, 134, 1570-95. https://doi.org/10.1053/j.gastro.2008.02.002
  35. Lind DS, Hochwald SN, Malaty J, et al (2000). Nuclear factor-${\kappa}B$ is upregulated in colorectal cancer. Surgery, 130, 363-9.
  36. Liu B, Qu L, Tao H (2009) Cyclo-oxygenase 2 up-regulates the effect of multidrug resistance. Cell Biol Int, 34, 21-5.
  37. Lu JJ, Bao JL, Chen XP, Huang M, Wang YT (2012). Alkaloids isolated from natural herbs as the anticancer agents. Evidence-Based Complementary Alternative Med, 2012, 485042.
  38. Manna K, Khan A, Kr Das D, et al (2014). Protective effect of coconut water concentrate and its active component shikimic acid against hydroperoxide mediated oxidative stress through suppression of NF-${\kappa}B$ and activation of Nrf2 pathway. J Ethnopharmacol, 155, 132-46. https://doi.org/10.1016/j.jep.2014.04.046
  39. Marwat SK, Fazal-ur-Rehman, Usman K, Shah SS, Anwar N, Ullah I (2012). A review of phytochemistry, bioactivities and ethno medicinal uses of Rhazya stricta Decsne (Apocynaceae). African J Microbiol Res, 6, 1629-41.
  40. Meerana SM, Katiyara SK (2008). Cell cycle control as a basis for cancer chemoprevention through dietary agents. Front Biosci, 13, 2191-202. https://doi.org/10.2741/2834
  41. Nagata S (2000). Apoptotic DNA fragmentation. Exp Cell Res, 256, 12-1. https://doi.org/10.1006/excr.2000.4834
  42. Nagle AA, Reddy SA, Bertrand H, et al (2014). 3-(2-Oxoethylidene) indolin-2-one derivatives activate Nrf2 and inhibit NF-${\kappa}B$: potential candidates for chemoprevention. Chem Med Chem, 9, 1763-74.
  43. Nakano K, Vousden KH (2001). PUMA, a novel proapoptotic gene, isinduced by p53. Mol Cell, 7, 683-94. https://doi.org/10.1016/S1097-2765(01)00214-3
  44. O'Connor PM, Jackman J, Bae I, et al (1997). Characterization of the p53 tumor suppressor pathway in cell lines of the National Cancer Institute anticancer drug screen and correlations with the growth-inhibitory potency of 123 anticancer agents. Cancer Res, 57, 4285-300.
  45. Pandurangan, AK, Esa NM (2013). Dietary non-nutritive factors in targeting of regulatory molecules in colorectal cancer: an update. Asian Pac J Cancer Prev, 14, 5543-52. https://doi.org/10.7314/APJCP.2013.14.10.5543
  46. Pandurangan AK, Kumar SA, Dharmalingam P, Ganapasam S (2014). Inhibitory effect of Luteolin on Azoxymethaneinduced colon carcinogenesis: Involvement of iNOS and COX-2. Pharmacog Magazine, 10, 306-10. https://doi.org/10.4103/0973-1296.133285
  47. Pereira SG, Oakley F (2008). Nuclear factor-${\kappa}B1$: Regulation and function. Int J Biochemistry Cell Biol, 40, 1425-30. https://doi.org/10.1016/j.biocel.2007.05.004
  48. Rabik CA, Dolan ME (2007). Molecular mechanisms of resistance and toxicity associated with platinating agents. Cancer Treat Rev, 33, 9-23. https://doi.org/10.1016/j.ctrv.2006.09.006
  49. Ragnhammar P, Hafstrom L, Nygren P, Glimelius B (2001). A systematic overview of chemotherapy effects in colorectal cancer. Acta Oncol, 40, 282-308. https://doi.org/10.1080/02841860121543
  50. Reed JC (2000). Mechanisms of apoptosis. Am J Pathol, 157, 1415-30. https://doi.org/10.1016/S0002-9440(10)64779-7
  51. Saraste A, Pulkki K (2000). Morphologic and biochemical hallmarks of apoptosis. Cardiovascular Res, 45, 528-37. https://doi.org/10.1016/S0008-6363(99)00384-3
  52. Schulz WA, Molecular biology of human cancers: an advanced student's textbook. Springer Science Business Media, Springer Dordrecht, 2007.
  53. Seeram NP, Adams LS, Hardy ML, Herber D (2004). Total cranberry extract versus its phytochemical constituents: antiproliferative and synergistic effects against human tumor cell lines. J Agric Food Chem, 52, 2512-7. https://doi.org/10.1021/jf0352778
  54. Seeram NP, Adams LS, Henning SM, et al (2005). In vitro antiproliferative, apoptotic and antioxidant activities of punicalagin, ellagic acid and a total pomegranate tannin extract are enhanced in combination with other polyphenols as found in pomegranate juice. J Nutr. Biochem, 16, 360-7.
  55. Sethi G, Sung B, Aggarwal BB (2008). Nuclear Factor-${\kappa}B$ activation: from bench to bedside. ExpBiol Med, 233, 21-31.
  56. Shao J, Fujiwara T, Kadowaki Y, et al (2000). Overexpression of the wild-type p53 gene inhibits NF-kappaB activity and synergizes with aspirin to induce apoptosis in human colon cancer cells. Oncogene, 19, 726-36. https://doi.org/10.1038/sj.onc.1203383
  57. Shaulian E, Karin M (2002). AP-1 as a regulator of cell life and death. Nat Cell Biol, 4, 131-6. https://doi.org/10.1038/ncb0502-e131
  58. Surh YJ (2008). NF-${\kappa}B$ and Nrf2 as potential chemopreventive targets of some anti-inflammatory and antioxidative phytonutrients with anti-inflammatory and antioxidative activities. Asia Pac J Clin Nutr, 17, 269-72.
  59. Vaiopoulos G, Papachroni K, Papavassiliou AG (2010). Colon carcinogenesis: Learning from NF-${\kappa}B$ and AP-1. Int J Biochemistry Cell Biol, 42, 1061-5. https://doi.org/10.1016/j.biocel.2010.03.018
  60. Vermeulen K, Van Bockstaele DR, Berneman ZN (2003). The cell cycle: a review of regulation, deregulation and therapeutic targets in cancer. Cell Prolif, 36, 131-49. https://doi.org/10.1046/j.1365-2184.2003.00266.x
  61. Violette S, Poulain L, Dussaulx E, et al (2002). Resistance of colon cancer cells to long-term 5-fluorouracil exposure is correlated to the relative level of Bcl-2 and Bcl-XL in addition to Bax and p53 status. Int J Cancer, 98, 498-504. https://doi.org/10.1002/ijc.10146
  62. Vousden KH (2002). Activation of the p53 tumor suppressor protein. Biochim Biophys Acta, 1602, 47-59.
  63. Wang J, Liu L, Qiu H, et al (2013). Ursolic acid simultaneously targets multiple signaling pathways to suppress proliferation and induce apoptosis in colon cancer cells. PLoS One, 8, 63872. https://doi.org/10.1371/journal.pone.0063872
  64. Weller M (1998). Predicting response to cancer chemotherapy: the role of p53. Cell Tissue Res, 292, 435-45. https://doi.org/10.1007/s004410051072
  65. Wenge Li, Tin Oo Khor, Changjiang Xu, Guoxiang Shen, Woo-Sik Jeong, Siwang Yu, and Ah-Ng Kon (2008). Activation of Nrf2-antioxidant signaling attenuates NF-${\kappa}B$ inflammatory response and elicits apoptosis. Biochem Pharmacol, 76, 1485-9. https://doi.org/10.1016/j.bcp.2008.07.017
  66. Wilkins RC, Kutzner BC, Truong M, Sanchez-Dardon J, McLean JR (2002). Analysis of radiation-induced apoptosis in human lymphocytes: flow cytometry using Annexin V and propidium iodide versus the neutral comet assay. Cytometry, 48, 14-9. https://doi.org/10.1002/cyto.10098
  67. Wong RS (2011). Apoptosis in cancer: from pathogenesis to Treatment. J Experimental Clin Cancer Res, 30, 87. https://doi.org/10.1186/1756-9966-30-87
  68. Yang SY, Sales KM, Fuller B, Seifalian AM, Winslet MC (2009). Apoptosis and colorectal cancer: implications for therapy. Trends Mol Med, 15, 225-33. https://doi.org/10.1016/j.molmed.2009.03.003
  69. Yasuhara S, Zhu Y, Matsui T, et al (2003). Comparison of cometassay, electronmicroscopy, and flowcytometry for detection of apoptosis. J Histochem Cytochem, 51, 873-85. https://doi.org/10.1177/002215540305100703
  70. Yuan JH, Li YQ, Yang XY (2007). Inhibition of epigallocatechin gallate on orthotopic colon cancer by upregulating the Nrf2-UGT1A signal pathway in nude mice," Pharmacol, 80, 269-78. https://doi.org/10.1159/000106447
  71. Zimmermann KC, Bonzon C, R. Green DR (2001). The machinery of programmed cell death. Pharmacol Therapeutics, 92, 57-70. https://doi.org/10.1016/S0163-7258(01)00159-0
  72. Zhang ZM, Yang XY, Yuan JH, Sun ZY, Li YQ (2009). Modulation of NRF2 and UGT1A expression by epigallocatechin-3- gallate in colon cancer cells and BALB/c mice. Chin Med J, 122, 1660-5.

Cited by

  1. Mill. (Boraginaceae) pp.1724-5575, 2018, https://doi.org/10.1080/11263504.2018.1527790