Breast Cancer Research and Treatment

, Volume 130, Issue 1, pp 61–71 | Cite as

Oral administration of benzyl-isothiocyanate inhibits solid tumor growth and lung metastasis of 4T1 murine mammary carcinoma cells in BALB/c mice

  • Eun Ji Kim
  • Ji Eun Hong
  • Soon Ju Eom
  • Jae-Yong Lee
  • Jung Han Yoon ParkEmail author
Preclinical study


Benzyl-isothiocyanate (BITC) is a hydrolysis product of glucotropaeolin, a compound found in cruciferous vegetables, and has also been shown to have anti-tumor properties. To evaluate the effects of BITC administration on the tumor growth and metastasis of breast cancer, 4T1 murine mammary carcinoma cells were injected into the inguinal mammary fat pads of syngeneic female BALB/c mice. One day later, the mice were subjected to gavage for 4 weeks with BITC (0, 5, or 10 mg/kg body weight/day). Oral BITC treatment induced a significant reduction in the growth of solid tumors. BITC reduced hemoglobin contents and CD31 and vascular endothelial growth factor (VEGF) expression in the tumors, as well as circulating levels of VEGF. Reduced expressions of proliferating cell nuclear antigen and cyclin-dependent kinase 4 were noted in the tumors of BITC-treated mice. BITC markedly increased the numbers of apoptotic cells with increased Bax expression, cleaved caspase-3, and PARP levels, but reduced Bcl-2 expression in tumor tissues. In addition, BITC was shown to reduce the numbers of pulmonary tumor nodules and the total pulmonary metastatic volume. BITC induced a significant reduction in the levels of matrix metalloproteinase (MMP)-2, MMP-9, tissue inhibitor of metalloproteinase (TIMP)-1, and urokinase-type plasminogen activator in the sera and lungs of 4T1 cell-injected mice. However, the concentrations of TIMP-2 and plasminogen activator inhibitor-1 were increased in the sera and lungs of BITC-treated mice. The results of this study indicate that BITC has potential as a preventive agent for metastatic breast cancer.


Benzyl-isothiocyanate Apoptosis Metastasis Cancer Angiogenesis 



This study was supported by a research grant from the National Research Foundation of Korea (NRF) for the Biofoods Research Program, Ministry of Education, Science, and Technology (MEST), and the Basic Science Research Program through the NRF funded by the MEST (314-2008-1-F00069).


  1. 1.
    Jemal A, Siegel R, Ward E, Hao Y, Xu J, Thun MJ (2009) Cancer statistics, 2009. CA Cancer J Clin 59(4):225–249PubMedCrossRefGoogle Scholar
  2. 2.
    Ali SM, Harvey HA, Lipton A (2003) Metastatic breast cancer: overview of treatment. Clin Orthop Relat Res (415 Suppl):S132–S137Google Scholar
  3. 3.
    Moore S (2007) Managing treatment side effects in advanced breast cancer. Semin Oncol Nurs 23(4 Suppl 2):S23–S30PubMedCrossRefGoogle Scholar
  4. 4.
    Fowke JH, Chung FL, Jin F, Qi D, Cai Q, Conaway C, Cheng JR, Shu XO, Gao YT, Zheng W (2003) Urinary isothiocyanate levels, brassica, and human breast cancer. Cancer Res 63(14):3980–3986PubMedGoogle Scholar
  5. 5.
    Ambrosone CB, McCann SE, Freudenheim JL, Marshall JR, Zhang Y, Shields PG (2004) Breast cancer risk in premenopausal women is inversely associated with consumption of broccoli, a source of isothiocyanates, but is not modified by GST genotype. J Nutr 134(5):1134–1138PubMedGoogle Scholar
  6. 6.
    Talalay P, Fahey JW (2001) Phytochemicals from cruciferous plants protect against cancer by modulating carcinogen metabolism. J Nutr 131(11 Suppl):3027S–3033SPubMedGoogle Scholar
  7. 7.
    Higdon JV, Delage B, Williams DE, Dashwood RH (2007) Cruciferous vegetables and human cancer risk: epidemiologic evidence and mechanistic basis. Pharmacol Res 55(3):224–236PubMedCrossRefGoogle Scholar
  8. 8.
    Nakamura Y, Kawakami M, Yoshihiro A, Miyoshi N, Ohigashi H, Kawai K, Osawa T, Uchida K (2002) Involvement of the mitochondrial death pathway in chemopreventive benzyl isothiocyanate-induced apoptosis. J Biol Chem 277(10):8492–8499PubMedCrossRefGoogle Scholar
  9. 9.
    Lui VW, Wentzel AL, Xiao D, Lew KL, Singh SV, Grandis JR (2003) Requirement of a carbon spacer in benzyl isothiocyanate-mediated cytotoxicity and MAPK activation in head and neck squamous cell carcinoma. Carcinogenesis 24(10):1705–1712PubMedCrossRefGoogle Scholar
  10. 10.
    Srivastava SK, Singh SV (2004) Cell cycle arrest, apoptosis induction and inhibition of nuclear factor kappa B activation in anti-proliferative activity of benzyl isothiocyanate against human pancreatic cancer cells. Carcinogenesis 25(9):1701–1709PubMedCrossRefGoogle Scholar
  11. 11.
    Miyoshi N, Uchida K, Osawa T, Nakamura Y (2004) A link between benzyl isothiocyanate-induced cell cycle arrest and apoptosis: involvement of mitogen-activated protein kinases in the Bcl-2 phosphorylation. Cancer Res 64(6):2134–2142PubMedCrossRefGoogle Scholar
  12. 12.
    Tseng E, Scott-Ramsay EA, Morris ME (2004) Dietary organic isothiocyanates are cytotoxic in human breast cancer MCF-7 and mammary epithelial MCF-12A cell lines. Exp Biol Med (Maywood) 229(8):835–842Google Scholar
  13. 13.
    Xiao D, Vogel V, Singh SV (2006) Benzyl isothiocyanate-induced apoptosis in human breast cancer cells is initiated by reactive oxygen species and regulated by Bax and Bak. Mol Cancer Ther 5(11):2931–2945PubMedCrossRefGoogle Scholar
  14. 14.
    Xiao D, Powolny AA, Singh SV (2008) Benzyl isothiocyanate targets mitochondrial respiratory chain to trigger reactive oxygen species-dependent apoptosis in human breast cancer cells. J Biol Chem 283(44):30151–30163PubMedCrossRefGoogle Scholar
  15. 15.
    Hwang ES, Lee HJ (2008) Benzyl isothiocyanate inhibits metalloproteinase-2/-9 expression by suppressing the mitogen-activated protein kinase in SK-Hep1 human hepatoma cells. Food Chem Toxicol 46(7):2358–2364PubMedCrossRefGoogle Scholar
  16. 16.
    Lai KC, Huang AC, Hsu SC, Kuo CL, Yang JS, Wu SH, Chung JG (2010) Benzyl isothiocyanate (BITC) inhibits migration and invasion of human colon cancer HT29 cells by inhibiting matrix metalloproteinase-2/-9 and urokinase plasminogen (uPA) through PKC and MAPK signaling pathway. J Agric Food Chem 58(5):2935–2942PubMedCrossRefGoogle Scholar
  17. 17.
    Sugie S, Okumura A, Tanaka T, Mori H (1993) Inhibitory effects of benzyl isothiocyanate and benzyl thiocyanate on diethylnitrosamine-induced hepatocarcinogenesis in rats. Jpn J Cancer Res 84(8):865–870PubMedGoogle Scholar
  18. 18.
    Wattenberg LW (1981) Inhibition of carcinogen-induced neoplasia by sodium cyanate, tert-butyl isocyanate, and benzyl isothiocyanate administered subsequent to carcinogen exposure. Cancer Res 41(8):2991–2994PubMedGoogle Scholar
  19. 19.
    Lin JM, Amin S, Trushin N, Hecht SS (1993) Effects of isothiocyanates on tumorigenesis by benzo[a]pyrene in murine tumor models. Cancer Lett 74(3):151–159PubMedCrossRefGoogle Scholar
  20. 20.
    Warin R, Chambers WH, Potter DM, Singh SV (2009) Prevention of mammary carcinogenesis in MMTV-neu mice by cruciferous vegetable constituent benzyl isothiocyanate. Cancer Res 69(24):9473–9480PubMedCrossRefGoogle Scholar
  21. 21.
    Guy CT, Webster MA, Schaller M, Parsons TJ, Cardiff RD, Muller WJ (1992) Expression of the neu protooncogene in the mammary epithelium of transgenic mice induces metastatic disease. Proc Natl Acad Sci USA 89(22):10578–10582PubMedCrossRefGoogle Scholar
  22. 22.
    Heppner GH, Miller FR, Shekhar PM (2000) Nontransgenic models of breast cancer. Breast Cancer Res 2(5):331–334PubMedCrossRefGoogle Scholar
  23. 23.
    Sauter BV, Martinet O, Zhang WJ, Mandeli J, Woo SL (2000) Adenovirus-mediated gene transfer of endostatin in vivo results in high level of transgene expression and inhibition of tumor growth and metastases. Proc Natl Acad Sci USA 97(9):4802–4807PubMedCrossRefGoogle Scholar
  24. 24.
    Welch DR, Neri A, Nicolson GL (1983) Comparison of ‘spontaneous’ and ‘experimental’ metastasis using rat 13762 mammary adenocarcinoma metastatic cell clones. Invasion Metastasis 3(2):65–80PubMedGoogle Scholar
  25. 25.
    Rose DP, Connolly JM (1992) Influence of dietary fat intake on local recurrence and progression of metastases arising from MDA-MB-435 human breast cancer cells in nude mice after excision of the primary tumor. Nutr Cancer 18(2):113–122PubMedCrossRefGoogle Scholar
  26. 26.
    Kim EJ, Shin M, Park H, Hong JE, Shin HK, Kim J, Kwon DY, Park JH (2009) Oral administration of 3,3′-diindolylmethane inhibits lung metastasis of 4T1 murine mammary carcinoma cells in BALB/c mice. J Nutr 139(12):2373–2379PubMedCrossRefGoogle Scholar
  27. 27.
    Cho HJ, Kim WK, Kim EJ, Jung KC, Park S, Lee HS, Tyner AL, Park JH (2003) Conjugated linoleic acid inhibits cell proliferation and ErbB3 signaling in HT-29 human colon cell line. Am J Physiol Gastrointest Liver Physiol 284(6):G996–G1005PubMedGoogle Scholar
  28. 28.
    Bove K, Lincoln DW, Tsan MF (2002) Effect of resveratrol on growth of 4T1 breast cancer cells in vitro and in vivo. Biochem Biophys Res Commun 291(4):1001–1005PubMedCrossRefGoogle Scholar
  29. 29.
    Michigami T, Hiraga T, Williams PJ, Niewolna M, Nishimura R, Mundy GR, Yoneda T (2002) The effect of the bisphosphonate ibandronate on breast cancer metastasis to visceral organs. Breast Cancer Res Treat 75(3):249–258PubMedCrossRefGoogle Scholar
  30. 30.
    Hecht SS (1996) Chemoprevention of lung cancer by isothiocyanates. Adv Exp Med Biol 401:1–11PubMedCrossRefGoogle Scholar
  31. 31.
    Hecht SS, Kenney PM, Wang M, Trushin N, Upadhyaya P (2000) Effects of phenethyl isothiocyanate and benzyl isothiocyanate, individually and in combination, on lung tumorigenesis induced in A/J mice by benzo[a]pyrene and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone. Cancer Lett 150(1):49–56PubMedCrossRefGoogle Scholar
  32. 32.
    Yu R, Mandlekar S, Harvey KJ, Ucker DS, Kong AN (1998) Chemopreventive isothiocyanates induce apoptosis and caspase-3-like protease activity. Cancer Res 58(3):402–408PubMedGoogle Scholar
  33. 33.
    Speijers GJ, Danse LH, van Leeuwen FX, Loeber JG (1985) Four-week toxicity study of phenyl isothiocyanate in rats. Food Chem Toxicol 23(11):1015–1017PubMedCrossRefGoogle Scholar
  34. 34.
    Lewerenz HJ, Plass R, Macholz R (1988) Effect of allyl isothiocyanate on hepatic monooxygenases and serum transferases in rats. Toxicol Lett 44(1–2):65–70PubMedCrossRefGoogle Scholar
  35. 35.
    Lewerenz HJ, Plass R, Bleyl DW, Macholz R (1988) Short-term toxicity study of allyl isothiocyanate in rats. Die Nahr 32(8):723–728CrossRefGoogle Scholar
  36. 36.
    Lewerenz HJ, Bleyl DW, Plass R (1992) Subacute oral toxicity study of benzyl isothiocyanate in rats. Die Nahr 36(2):190–198CrossRefGoogle Scholar
  37. 37.
    Ozierenski B, Plass R, Lewerenz HJ (1993) Effects of glucosinolate breakdown products on the hepatic biotransformation system in male rats. Die Nahr 37(1):5–14CrossRefGoogle Scholar
  38. 38.
    Folkman J, Shing Y (1992) Angiogenesis. J Biol Chem 267(16):10931–10934PubMedGoogle Scholar
  39. 39.
    Coussens LM, Werb Z (1996) Matrix metalloproteinases and the development of cancer. Chem Biol 3(11):895–904PubMedCrossRefGoogle Scholar
  40. 40.
    Castellino FJ, Ploplis VA (2005) Structure and function of the plasminogen/plasmin system. Thromb Haemost 93(4):647–654PubMedGoogle Scholar
  41. 41.
    Kwaan HC (1992) The plasminogen-plasmin system in malignancy. Cancer Metastasis Rev 11(3–4):291–311PubMedCrossRefGoogle Scholar
  42. 42.
    Somiari SB, Somiari RI, Heckman CM, Olsen CH, Jordan RM, Russell SJ, Shriver CD (2006) Circulating MMP2 and MMP9 in breast cancer—potential role in classification of patients into low risk, high risk, benign disease and breast cancer categories. Int J Cancer 119(6):1403–1411PubMedCrossRefGoogle Scholar
  43. 43.
    Overall CM, Kleifeld O (2006) Tumour microenvironment—opinion: validating matrix metalloproteinases as drug targets and anti-targets for cancer therapy. Nat Rev Cancer 6(3):227–239PubMedCrossRefGoogle Scholar
  44. 44.
    Foekens JA, Peters HA, Look MP, Portengen H, Schmitt M, Kramer MD, Brunner N, Janicke F, Meijer-van Gelder ME, Henzen-Logmans SC et al (2000) The urokinase system of plasminogen activation and prognosis in 2780 breast cancer patients. Cancer Res 60(3):636–643PubMedGoogle Scholar
  45. 45.
    Giavazzi R, Taraboletti G (2001) Preclinical development of metalloproteasis inhibitors in cancer therapy. Crit Rev Oncol Hematol 37(1):53–60PubMedCrossRefGoogle Scholar
  46. 46.
    Brew K, Dinakarpandian D, Nagase H (2000) Tissue inhibitors of metalloproteinases: evolution, structure and function. Biochim Biophys Acta 1477(1–2):267–283PubMedCrossRefGoogle Scholar
  47. 47.
    Kuvaja P, Talvensaari-Mattila A, Turpeenniemi-Hujanen T (2008) High preoperative plasma TIMP-1 is prognostic for early relapse in primary breast carcinoma. Int J Cancer 123(4):846–851PubMedCrossRefGoogle Scholar
  48. 48.
    Chirco R, Liu XW, Jung KK, Kim HR (2006) Novel functions of TIMPs in cell signaling. Cancer Metastasis Rev 25(1):99–113PubMedCrossRefGoogle Scholar
  49. 49.
    Shirasuna K, Saka M, Hayashido Y, Yoshioka H, Sugiura T, Matsuya T (1993) Extracellular matrix production and degradation by adenoid cystic carcinoma cells: participation of plasminogen activator and its inhibitor in matrix degradation. Cancer Res 53(1):147–152PubMedGoogle Scholar
  50. 50.
    Stefansson S, Lawrence DA (1996) The serpin PAI-1 inhibits cell migration by blocking integrin alpha V beta 3 binding to vitronectin. Nature 383(6599):441–443PubMedCrossRefGoogle Scholar
  51. 51.
    Minisini AM, Fabbro D, Di Loreto C, Pestrin M, Russo S, Cardellino GG, Andreetta C, Damante G, Puglisi F (2007) Markers of the uPA system and common prognostic factors in breast cancer. Am J Clin Pathol 128(1):112–117PubMedCrossRefGoogle Scholar
  52. 52.
    Shariat SF, Roehrborn CG, McConnell JD, Park S, Alam N, Wheeler TM, Slawin KM (2007) Association of the circulating levels of the urokinase system of plasminogen activation with the presence of prostate cancer and invasion, progression, and metastasis. J Clin Oncol 25(4):349–355PubMedCrossRefGoogle Scholar
  53. 53.
    Yoshino Y, Kageshita T, Nakajima M, Funakubo M, Ihn H (2008) Clinical relevance of serum levels of matrix metallopeptidase-2, and tissue inhibitor of metalloproteinase-1 and -2 in patients with malignant melanoma. J Dermatol 35(4):206–214PubMedCrossRefGoogle Scholar
  54. 54.
    Steeg PS (2006) Tumor metastasis: mechanistic insights and clinical challenges. Nat Med 12(8):895–904PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC. 2010

Authors and Affiliations

  • Eun Ji Kim
    • 2
  • Ji Eun Hong
    • 1
  • Soon Ju Eom
    • 2
  • Jae-Yong Lee
    • 2
    • 3
  • Jung Han Yoon Park
    • 1
    • 2
    • 4
    Email author
  1. 1.Department of Food Science and NutritionHallym UniversityChuncheonKorea
  2. 2.Center for Efficacy Assessment and Development of Functional Foods and DrugsHallym UniversityChuncheonKorea
  3. 3.Department of Biochemistry, College of MedicineHallym UniversityChuncheonKorea
  4. 4.Medical and Bio-materials Research CenterHallym UniversityChuncheonKorea

Personalised recommendations