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Synergistic effect of 5-fluorouracil and the flavanoid oroxylin A on HepG2 human hepatocellular carcinoma and on H22 transplanted mice

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Abstract

Aim

To investigate the synergistic inhibitory effects of the combination of 5-fluorouracil (5-FU) with the natural flavanoid oroxylin A on human hepatocellular carcinoma cells HepG2 in vitro and on transplanted murine hepatoma 22 (H22) tumors in vivo and the preliminary mechanisms.

Methods

The inhibitory effects of 5-FU combined with the natural flavanoid oroxylin A in vitro were detected by MTT assay and the effects in vivo were investigated by transplanted H22 mice model. DAPI staining and Annexin V/propidium iodide (PI) double staining were used to detect the cell morphological changes and apoptosis. The mRNA levels of thymidine synthetase (TS) and dihydropyrimidine dehydrogenase (DPD) in HepG2 cells after oroxylin A and 5-FU combination treatment were observed by quantitative real-time PCR. Western blotting assay was used to reveal the expressions of apoptotic-inducing proteins P53, cleaved PARP, COX-2, Bcl-2, and pro-caspase3.

Results

Oroxylin A in combination with 5-FU presented synergistic effect (CI < 1) on HepG2 cells in vitro when the inhibitory rate was higher than 7.5%. The inhibitory rate on H22 murine solid tumor in vivo in the combination group was higher than monotherapy. 5-FU combined with oroxylin A exerted stronger apoptotic induction in HepG2 cells than either single drug treatment. Quantitative real-time PCR discovered the downregulation of TS mRNA and DPD mRNA in HepG2 cells after combination treatment. Western blotting assay revealed oroxylin A enhanced 5-FU-induced apoptosis in HepG2 cells by elevating the expressions of apoptotic-inducing proteins P53 and cleaved PARP and decreasing the expression of apoptotic-inhibitory proteins COX-2, Bcl-2, and pro-caspase3.

Conclusion

The anti-hepatocellular carcinoma effects in vitro and in vivo of 5-FU and oroxylin A combinations were synergistic and oroxylin A increased the sensitivity of HepG2 cells to 5-FU by modulating the metabolic enzymes of 5-FU and apoptotic-related proteins.

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References

  1. Carrico CK, Glazer RI (1979) Effect of 5-fluorouracil on the synthesis and translation of polyadenylic acid-containing RNA from regenerating rat liver. Cancer Res 39:3694–3701

    CAS  PubMed  Google Scholar 

  2. Chen Y, Yang L, Lee TJ (2000) Oroxylin A inhibition of lipopolysaccharide-induced iNOS and COX-2 gene expression via suppression of nuclear factor-kappaB activation. Biochem Pharmacol 59:1445–1457

    Article  CAS  PubMed  Google Scholar 

  3. Cheung YC, Ko SF, Ng SH, Chan SC, Cheng YF (2005) Survival outcome of lobar or segmental transcatheter arterial embolization with ethanol–lipiodol mixture in treating hepatocellular carcinoma. World J Gastroenterol 11:2792–2795

    PubMed  Google Scholar 

  4. Chou TC (2006) Theoretical basis, experimental design, and computerized simulation of synergism and antagonism in drug combination studies. Pharmacol Rev 58:621–681

    Article  CAS  PubMed  Google Scholar 

  5. Chou TC, Motzer RJ, Tong Y, Bosl GJ (1994) Computerized quantitation of synergism and antagonism of taxol, topotecan, and cisplatin against human teratocarcinoma cell growth: a rational approach to clinical protocol design. J Natl Cancer Inst 86:1517–1524

    Article  CAS  PubMed  Google Scholar 

  6. Chou TC, Talalay P (1984) Quantitative analysis of dose–effect relationships: the combined effects of multiple drugs or enzyme inhibitors. Adv Enzyme Regul 22:27–55

    Article  CAS  PubMed  Google Scholar 

  7. Ebara M, Okabe S, Kita K, Sugiura N, Fukuda H, Yoshikawa M, Kondo F, Saisho H (2005) Percutaneous ethanol injection for small hepatocellular carcinoma: therapeutic efficacy based on 20-year observation. J Hepatol 43:458–464

    Article  CAS  PubMed  Google Scholar 

  8. Fischel JL, Etienne MC, Spector T, Formento P, Renee N, Milano G (1995) Dihydropyrimidine dehydrogenase: a tumoral target for fluorouracil modulation. Clin Cancer Res 1:991–996

    CAS  PubMed  Google Scholar 

  9. Hu Y, Yang Y, You QD, Liu W, Gu HY, Zhao L, Zhang K, Wang W, Wang XT, Guo QL (2006) Oroxylin A induced apoptosis of human hepatocellular carcinoma cell line HepG2 was involved in its antitumor activity. Biochem Biophys Res Commun 351:521–527

    Article  CAS  PubMed  Google Scholar 

  10. Huang WH, Lee AR, Yang CH (2006) Antioxidative and anti-inflammatory activities of polyhydroxyflavonoids of Scutellaria baicalensis GEORGI. Biosci Biotechnol Biochem 70:2371–2380

    Article  CAS  PubMed  Google Scholar 

  11. Kawamoto C, Ido K, Isoda N, Hozumi M, Nagamine N, Ono K, Sato Y, Kobayashi Y, Nagae G, Sugano K (2005) Long-term outcomes for patients with solitary hepatocellular carcinoma treated by laparoscopic microwave coagulation. Cancer 103:985–993

    Article  PubMed  Google Scholar 

  12. Kim DH, Jeon SJ, Son KH, Jung JW, Lee S, Yoon BH, Lee JJ, Cho YW, Cheong JH, Ko KH, Ryu JH (2007) The ameliorating effect of oroxylin A on scopolamine-induced memory impairment in mice. Neurobiol Learn Mem 87:536–546

    Article  CAS  PubMed  Google Scholar 

  13. Kim DH, Kim S, Jeon SJ, Son KH, Lee S, Yoon BH, Cheong JH, Ko KH, Ryu JH (2008) The effects of acute and repeated oroxylin A treatments on Abeta(25–35)-induced memory impairment in mice. Neuropharmacology 55:639–647

    Article  CAS  PubMed  Google Scholar 

  14. Kim JY, Lee S, Kim DH, Kim BR, Park R, Lee BM (2002) Effects of flavonoids isolated from Scutellariae radix on cytochrome P-450 activities in human liver microsomes. J Toxicol Environ Health A 65:373–381

    Article  CAS  PubMed  Google Scholar 

  15. Kitchens ME, Forsthoefel AM, Barbour KW, Spencer HT, Berger FG (1999) Mechanisms of acquired resistance to thymidylate synthase inhibitors: the role of enzyme stability. Mol Pharmacol 56:1063–1070

    CAS  PubMed  Google Scholar 

  16. Li HB, Chen F (2005) Isolation and purification of baicalein, wogonin and oroxylin A from the medicinal plant Scutellaria baicalensis by high-speed counter-current chromatography. J Chromatogr A 1074:107–110

    Article  CAS  PubMed  Google Scholar 

  17. Llovet JM (2005) Updated treatment approach to hepatocellular carcinoma. J Gastroenterol 40:225–235

    Article  PubMed  Google Scholar 

  18. Ma SC, Du J, But PP, Deng XL, Zhang YW, Ooi VE, Xu HX, Lee SH, Lee SF (2002) Antiviral Chinese medicinal herbs against respiratory syncytial virus. J Ethnopharmacol 79:205–211

    Article  CAS  PubMed  Google Scholar 

  19. Piper AA, Fox RM (1982) Biochemical basis for the differential sensitivity of human T- and B-lymphocyte lines to 5-fluorouracil. Cancer Res 42:3753–3760

    CAS  PubMed  Google Scholar 

  20. Salonga D, Danenberg KD, Johnson M, Metzger R, Groshen S, Tsao-Wei DD, Lenz HJ, Leichman CG, Leichman L, Diasio RB, Danenberg PV (2000) Colorectal tumors responding to 5-fluorouracil have low gene expression levels of dihydropyrimidine dehydrogenase, thymidylate synthase, and thymidine phosphorylase. Clin Cancer Res 6:1322–1327

    CAS  PubMed  Google Scholar 

  21. Sasaki E, Tominaga K, Kuwamura H, Watanabe T, Fujiwara Y, Oshitani N, Higuchi K, Arakawa T (2007) Synergistic antitumor effect of combined 5-fluorouracil (5-FU) with 5-chloro-2, 4-dihydroxypyridine on 5-FU-resistant gastric cancer cells: possible role of a dihydropyrimidine dehydrogenase-independent mechanism. J Gastroenterol 42:816–822

    Article  CAS  PubMed  Google Scholar 

  22. Sonoda M, Nishiyama T, Matsukawa Y, Moriyasu M (2004) Cytotoxic activities of flavonoids from two Scutellaria plants in Chinese medicine. J Ethnopharmacol 91:65–68

    Article  CAS  PubMed  Google Scholar 

  23. Spears CP, Shahinian AH, Moran RG, Heidelberger C, Corbett TH (1982) In vivo kinetics of thymidylate synthetase inhibition of 5-fluorouracil-sensitive and -resistant murine colon adenocarcinomas. Cancer Res 42:450–456

    CAS  PubMed  Google Scholar 

  24. Sun Y, Tang XM, Half E, Kuo MT, Sinicrope FA (2002) Cyclooxygenase-2 overexpression reduces apoptotic susceptibility by inhibiting the cytochrome c-dependent apoptotic pathway in human colon cancer cells. Cancer Res 62:6323–6328

    CAS  PubMed  Google Scholar 

  25. Tateishi R, Shiina S, Teratani T, Obi S, Sato S, Koike Y, Fujishima T, Yoshida H, Kawabe T, Omata M (2005) Percutaneous radiofrequency ablation for hepatocellular carcinoma. An analysis of 1000 cases. Cancer 103:1201–1209

    Article  PubMed  Google Scholar 

  26. van Kuilenburg AB, Haasjes J, Richel DJ, Zoetekouw L, Van Lenthe H, De Abreu RA, Maring JG, Vreken P, van Gennip AH (2000) Clinical implications of dihydropyrimidine dehydrogenase (DPD) deficiency in patients with severe 5-fluorouracil-associated toxicity: identification of new mutations in the DPD gene. Clin Cancer Res 6:4705–4712

    PubMed  Google Scholar 

  27. Van Triest B, Pinedo HM, Giaccone G, Peters GJ (2000) Downstream molecular determinants of response to 5-fluorouracil and antifolate thymidylate synthase inhibitors. Ann Oncol 11:385–391

    Article  PubMed  Google Scholar 

  28. Wada Y, Yoshida K, Suzuki T, Mizuiri H, Konishi K, Ukon K, Tanabe K, Sakata Y, Fukushima M (2006) Synergistic effects of docetaxel and S-1 by modulating the expression of metabolic enzymes of 5-fluorouracil in human gastric cancer cell lines. Int J Cancer 119:783–791

    Article  CAS  PubMed  Google Scholar 

  29. Wang X, Zhang J, Xu T (2008) Thioredoxin reductase inactivation as a pivotal mechanism of ifosfamide in cancer therapy. Eur J Pharmacol 579:66–73

    Article  CAS  PubMed  Google Scholar 

  30. Yang Y, Hu Y, Gu HY, Lu N, Liu W, Qi Q, Zhao L, Wang XT, You QD, Guo QL (2008) Oroxylin A induces G2/M phase cell-cycle arrest via inhibiting Cdk7-mediated expression of Cdc2/p34 in human gastric carcinoma BGC-823 cells. J Pharm Pharmacol 60:1459–1463

    Article  CAS  PubMed  Google Scholar 

  31. Yang Y, Yang L, You QD, Nie FF, Gu HY, Zhao L, Wang XT, Guo QL (2007) Differential apoptotic induction of gambogic acid, a novel anticancer natural product, on hepatoma cells and normal hepatocytes. Cancer Lett 256:259–266

    Article  CAS  PubMed  Google Scholar 

  32. Yee KS, Vousden KH (2005) Complicating the complexity of p53. Carcinogenesis 26:1317–1322

    Article  CAS  PubMed  Google Scholar 

  33. Yim EK, Lee KH, Bae JS, Namkoong SE, Um SJ, Park JS (2004) Proteomic analysis of antiproliferative effects by treatment of 5-fluorouracil in cervical cancer cells. DNA Cell Biol 23:769–776

    Article  CAS  PubMed  Google Scholar 

  34. Yu J, Guo QL, You QD, Zhao L, Gu HY, Yang Y, Zhang HW, Tan Z, Wang X (2007) Gambogic acid-induced G2/M phase cell-cycle arrest via disturbing CDK7-mediated phosphorylation of CDC2/p34 in human gastric carcinoma BGC-823 cells. Carcinogenesis 28:632–638

    Article  CAS  PubMed  Google Scholar 

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Acknowledgments

This work was supported by the National High Technology Research and Development Program of China (863 program, No. 2003AA2Z3267), National Institutes of Health (SCORE S06GM-008205) and Jiangsu Province Six Talent Pools in Education, Medical care and hygiene, Electronic information, Machinery and motor, Architecture and Agriculture Project.

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Authors and Affiliations

  1. Jiangsu Key Laboratory of Carcinogenesis and Intervention, China Pharmaceutical University, 24 Tongjiaxiang, 210009, Nanjing, China

    Li Zhao, Jun Wang, Li Yang, Qing Zhao, Jia Wang, Qi Qi, Rong Mu & Qing-Long Guo

  2. Department of Medicinal Chemistry, China Pharmaceutical University, 24 Tongjiaxiang, 210009, Nanjing, China

    Qi-Dong You

  3. Department of Pharmacology, China Pharmaceutical University, 24 Tongjiaxiang, 210009, Nanjing, China

    Zhen Chen

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  1. Li Zhao
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  2. Zhen Chen
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Correspondence to Qi-Dong You or Qing-Long Guo.

Additional information

Li Zhao, Zhen Chen contributed equally to this article.

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Zhao, L., Chen, Z., Wang, J. et al. Synergistic effect of 5-fluorouracil and the flavanoid oroxylin A on HepG2 human hepatocellular carcinoma and on H22 transplanted mice. Cancer Chemother Pharmacol 65, 481–489 (2010). https://doi.org/10.1007/s00280-009-1053-2

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