Archives of Pharmacal Research

, Volume 41, Issue 2, pp 229–242 | Cite as

Nafamostat mesilate negatively regulates the metastasis of triple-negative breast cancer cells

  • Sunam Mander
  • Dong-Joo You
  • Sumi Park
  • Dong Hwi Kim
  • Hyo Jeong Yong
  • Dong-Sik Kim
  • Curie Ahn
  • Yun-Hee Kim
  • Jae Young Seong
  • Jong-Ik Hwang
Research Article


Triple-negative breast cancer (TNBC) lacking of oestrogen receptor, progesterone receptor, and epidermal growth factor receptor type 2 is a highly malignant disease which results in a poor prognosis and rare treatment options. Despite the use of conventional chemotherapy for TNBC tumours, resistance and short duration responses limit the treatment efficacy. Therefore, a need exists to develop a new chemotherapy for TNBC. The aim of this study was to examine the anti-cancer effects of nafamostat mesilate (NM), a previously known serine protease inhibitor and highly safe drug on breast cancer cells. Here, we showed that NM significantly inhibits proliferation, migration, and invasion in MDA-MB231 cells, induces G2/M phase cell-cycle arrest, and inhibits the expression of cyclin-dependent kinase 1 (CDK1). Exposure of MDA-MB231 cells to NM also resulted in decreased transcription factor activities accompanied by the regulated phosphorylation of signalling molecules and a decrease in metalloproteinases, the principal modulators of the extracellular environment during cancer progression. Especially, inhibition of TGFβ-stimulated Smad2 phosphorylation and subsequent metastasis-related gene expression, and downregulation of ERK activity may be pivotal mechanisms underlying inhibitory effects of NM on NM inhibits lung metastasis of breast cancer cells and growth of colonized tumours in mice. Taken together, our data revealed that NM inhibits cell growth and metastasis of TNBC cells and indicated that NM is a multi-targeted drug that could be an adjunct therapy for TNBC treatment.


Nafamostat mesilate Triple negative breast cancer Cell cycle Metastasis 



This work was supported by a Korea Research Foundation Grant funded by the Ministry of Science, ICT, and Future Planning (MSIP; No. NRF-2016R1A2B1010036).

Compliance with ethical standards

Conflict of interest

The authors have no conflict of interest to disclose.


  1. Akizawa T, Koshikawa S, Ota K, Kazama M, Mimura N, Hirasawa Y (1993) Nafamostat mesilate: a regional anticoagulant for hemodialysis in patients at high risk for bleeding. Nephron 64:376–381CrossRefPubMedGoogle Scholar
  2. Andre F, Zielinski CC (2012) Optimal strategies for the treatment of metastatic triple-negative breast cancer with currently approved agents. Ann Oncol 23(Suppl 6):vi46–vi51CrossRefPubMedGoogle Scholar
  3. Bartholomeusz C, Gonzalez-Angulo AM, Liu P, Hayashi N, Lluch A, Ferrer-Lozano J, Hortobagyi GN (2012) High ERK protein expression levels correlate with shorter survival in triple-negative breast cancer patients. Oncologist 17:766–774CrossRefPubMedPubMedCentralGoogle Scholar
  4. Benson JR (2004) Role of transforming growth factor beta in breast carcinogenesis. Lancet Oncol 5:229–239CrossRefPubMedGoogle Scholar
  5. Bertucci F, Finetti P, Cervera N, Esterni B, Hermitte F, Viens P, Birnbaum D (2008) How basal are triple-negative breast cancers? Int J Cancer 123:236–240CrossRefPubMedGoogle Scholar
  6. Bose S, Chandran S, Mirocha JM, Bose N (2006) The Akt pathway in human breast cancer: a tissue-array-based analysis. Mod Pathol 19:238–245CrossRefPubMedGoogle Scholar
  7. Boyle P (2012) Triple-negative breast cancer: epidemiological considerations and recommendations. Ann Oncol 23(Suppl 6):vi7–vi12CrossRefPubMedGoogle Scholar
  8. Brandi G, Tavolari S, De Rosa F, Di Girolamo S, Agostini V, Barbera MA, Frega G, Biasco G (2012) Antitumoral efficacy of the protease inhibitor gabexate mesilate in colon cancer cells harbouring KRAS, BRAF and PIK3CA mutations. PLoS ONE 7:e41347CrossRefPubMedPubMedCentralGoogle Scholar
  9. Carey LA, Dees EC, Sawyer L, Gatti L, Moore DT, Collichio F, Ollila DW, Sartor CI, Graham ML, Perou CM (2007) The triple negative paradox: primary tumor chemosensitivity of breast cancer subtypes. Clin Cancer Res 13:2329–2334CrossRefPubMedGoogle Scholar
  10. Cho EY, Choi SC, Lee SH, Ahn JY, Im LR, Kim JH, Xin M, Kwon SU, Kim DK, Lee YM (2011) Nafamostat mesilate attenuates colonic inflammation and mast cell infiltration in the experimental colitis. Int Immunopharmacol 11:412–417CrossRefPubMedGoogle Scholar
  11. Crown J, O’shaughnessy J, Gullo G (2012) Emerging targeted therapies in triple-negative breast cancer. Ann Oncol 23(Suppl 6):vi56–vi65CrossRefPubMedGoogle Scholar
  12. Dent R, Trudeau M, Pritchard KI, Hanna WM, Kahn HK, Sawka CA, Lickley LA, Rawlinson E, Sun P, Narod SA (2007) Triple-negative breast cancer: clinical features and patterns of recurrence. Clin Cancer Res 13:4429–4434CrossRefPubMedGoogle Scholar
  13. Egeblad M, Werb Z (2002) New functions for the matrix metalloproteinases in cancer progression. Nat Rev Cancer 2:161–174CrossRefPubMedGoogle Scholar
  14. Foulkes WD, Smith IE, Reis-Filho JS (2010) Triple-negative breast cancer. N Engl J Med 363:1938–1948CrossRefPubMedGoogle Scholar
  15. Fujii S, Hitomi Y (1981) New synthetic inhibitors of C1r, C1 esterase, thrombin, plasmin, kallikrein and trypsin. Biochim Biophys Acta 661:342–345CrossRefPubMedGoogle Scholar
  16. Fujiwara Y, Furukawa K, Haruki K, Shimada Y, Iida T, Shiba H, Uwagawa T, Ohashi T, Yanaga K (2011) Nafamostat mesilate can prevent adhesion, invasion and peritoneal dissemination of pancreatic cancer thorough nuclear factor kappa-B inhibition. J Hepatobil Pancreat Sci 18:731–739CrossRefGoogle Scholar
  17. Furukawa K, Iida T, Shiba H, Fujiwara Y, Uwagawa T, Shimada Y, Misawa T, Ohashi T, Yanaga K (2010) Anti-tumor effect by inhibition of NF-kappaB activation using nafamostat mesilate for pancreatic cancer in a mouse model. Oncol Rep 24:843–850CrossRefPubMedGoogle Scholar
  18. Furukawa K, Uwagawa T, Iwase R, Haruki K, Fujiwara Y, Gocho T, Shiba H, Misawa T, Yanaga K (2012) Prognostic factors of unresectable pancreatic cancer treated with nafamostat mesilate combined with gemcitabine chemotherapy. Anticancer Res 32:5121–5126PubMedGoogle Scholar
  19. Gajria D, Chandarlapaty S (2011) HER2-amplified breast cancer: mechanisms of trastuzumab resistance and novel targeted therapies. Expert Rev Anticancer Ther 11:263–275CrossRefPubMedPubMedCentralGoogle Scholar
  20. Giltnane JM, Balko JM (2014) Rationale for targeting the Ras/MAPK pathway in triple-negative breast cancer. Discov Med 17:275–283PubMedGoogle Scholar
  21. Gocho T, Uwagawa T, Furukawa K, Haruki K, Fujiwara Y, Iwase R, Misawa T, Ohashi T, Yanaga K (2013) Combination chemotherapy of serine protease inhibitor nafamostat mesilate with oxaliplatin targeting NF-kappaB activation for pancreatic cancer. Cancer Lett 333:89–95CrossRefPubMedGoogle Scholar
  22. Hahn SA, Schutte M, Shamsul Hoque ATM, Moskaluk CA, Da Costa LT, Rozenblum E, Weinstein CL, Fischer A, Yeo CJ, Hruban RH, Kern SE (1996) DPC4, a candidate tumor suppressor gene at human chromosome 18q21.1. Science 271:350–353CrossRefPubMedGoogle Scholar
  23. Hartman ZC, Poage GM, Den Hollander P, Tsimelzon A, Hill J, Panupinthu N, Zhang Y, Mazumdar A, Hilsenbeck SG, Mills GB, Brown PH (2013) Growth of triple-negative breast cancer cells relies upon coordinate autocrine expression of the proinflammatory cytokines IL-6 and IL-8. Cancer Res 73:3470–3480CrossRefPubMedGoogle Scholar
  24. Hortobagyi GN (2005) Trastuzumab in the treatment of breast cancer. N Engl J Med 353:1734–1736CrossRefPubMedGoogle Scholar
  25. Iwaki M, Ino Y, Motoyoshi A, Ozeki M, Sato T, Kurumi M, Aoyama T (1986) Pharmacological studies of FUT-175, nafamostat mesilate. V. Effects on the pancreatic enzymes and experimental acute pancreatitis in rats. Jpn J Pharmacol 41:155–162CrossRefPubMedGoogle Scholar
  26. Jakowlew SB (2006) Transforming growth factor-beta in cancer and metastasis. Cancer Metastasis Rev 25:435–457CrossRefPubMedGoogle Scholar
  27. Johnston SR, Maclennan KA, Sacks NP, Salter J, Smith IE, Dowsett M (1994) Modulation of Bcl-2 and Ki-67 expression in oestrogen receptor-positive human breast cancer by tamoxifen. Eur J Cancer 30a:1663–1669CrossRefPubMedGoogle Scholar
  28. Kalluri R, Neilson EG (2003) Epithelial-mesenchymal transition and its implications for fibrosis. J Clin Investig 112:1776–1784CrossRefPubMedPubMedCentralGoogle Scholar
  29. Kalluri R, Weinberg RA (2009) The basics of epithelial-mesenchymal transition. J Clin Investig 119:1420–1428CrossRefPubMedPubMedCentralGoogle Scholar
  30. Kassam F, Enright K, Dent R, Dranitsaris G, Myers J, Flynn C, Fralick M, Kumar R, Clemons M (2009) Survival outcomes for patients with metastatic triple-negative breast cancer: implications for clinical practice and trial design. Clin Breast Cancer 9:29–33CrossRefPubMedGoogle Scholar
  31. Kessenbrock K, Plaks V, Werb Z (2010) Matrix metalloproteinases: regulators of the tumor microenvironment. Cell 141:52–67CrossRefPubMedPubMedCentralGoogle Scholar
  32. Kimura T, Fuchimoto S, Iwagaki H, Hizuta A, Orita K (1992) Inhibitory effect of nafamostat mesilate on metastasis into the livers of mice and on invasion of the extracellular matrix by cancer cells. J Int Med Res 20:343–352CrossRefPubMedGoogle Scholar
  33. Lee HO, Sheen YY (1997) Estrogen modulation of human breast cancer cell growth. Arch Pharm Res 20:566–571CrossRefPubMedGoogle Scholar
  34. Liedtke C, Mazouni C, Hess KR, Andre F, Tordai A, Mejia JA, Symmans WF, Gonzalez-Angulo AM, Hennessy B, Green M, Cristofanilli M, Hortobagyi GN, Pusztai L (2008) Response to neoadjuvant therapy and long-term survival in patients with triple-negative breast cancer. J Clin Oncol 26:1275–1281CrossRefPubMedGoogle Scholar
  35. Lu YX, Ju HQ, Wang F, Chen LZ, Wu QN, Sheng H, Mo HY, Pan ZZ, Xie D, Kang TB, Chen G, Yun JP, Zeng ZL, Xu RH (2016) Inhibition of the NF-kappaB pathway by nafamostat mesilate suppresses colorectal cancer growth and metastasis. Cancer Lett 380:87–97CrossRefPubMedGoogle Scholar
  36. Lv ZD, Kong B, Li JG, Qu HL, Wang XG, Cao WH, Liu XY, Wang Y, Yang ZC, Xu HM, Wang HB (2013) Transforming growth factor-beta 1 enhances the invasiveness of breast cancer cells by inducing a Smad2-dependent epithelial-to-mesenchymal transition. Oncol Rep 29:219–225CrossRefPubMedGoogle Scholar
  37. Mukherjee D, Zhao J (2013) The Role of chemokine receptor CXCR4 in breast cancer metastasis. Am J Cancer Res 3:46–57PubMedPubMedCentralGoogle Scholar
  38. Naber HP, Wiercinska E, Pardali E, Van Laar T, Nirmala E, Sundqvist A, Van Dam H, Van Der Horst G, Van Der Pluijm G, Heckmann B, Danen EH, Ten Dijke P (2012) BMP-7 inhibits TGF-beta-induced invasion of breast cancer cells through inhibition of integrin beta(3) expression. Cell Oncol (Dordr) 35:19–28CrossRefGoogle Scholar
  39. Nakatsuka M, Asagiri K, Noguchi S, Habara T, Kudo T (2000) Nafamostat mesilate, a serine protease inhibitor, suppresses lipopolysaccharide-induced nitric oxide synthesis and apoptosis in cultured human trophoblasts. Life Sci 67:1243–1250CrossRefPubMedGoogle Scholar
  40. Noguchi S, Nakatsuka M, Konishi H, Kamada Y, Chekir C, Kudo T (2003) Nafamostat mesilate suppresses NF-kappaB activation and NO overproduction in LPS-treated macrophages. Int Immunopharmacol 3:1335–1344CrossRefPubMedGoogle Scholar
  41. Okamoto T, Mizoguchi S, Terasaki H, Morioka T (1994) Safety of high-dose of nafamostat mesilate: toxicological study in beagles. J Pharmacol Exp Ther 268:639–644PubMedGoogle Scholar
  42. Padua D, Massague J (2009) Roles of TGF[beta] in metastasis. Cell Res 19:89–102CrossRefPubMedGoogle Scholar
  43. Parkin DM, Fernandez LM (2006) Use of statistics to assess the global burden of breast cancer. Breast J 12(Suppl 1):S70–S80CrossRefPubMedGoogle Scholar
  44. Perou CM, Sorlie T, Eisen MB, Van De Rijn M, Jeffrey SS, Rees CA, Pollack JR, Ross DT, Johnsen H, Akslen LA, Fluge O, Pergamenschikov A, Williams C, Zhu SX, Lonning PE, Borresen-Dale AL, Brown PO, Botstein D (2000) Molecular portraits of human breast tumours. Nature 406:747–752CrossRefPubMedGoogle Scholar
  45. Prat A, Baselga J (2008) The role of hormonal therapy in the management of hormonal-receptor-positive breast cancer with co-expression of HER2. Nat Clin Pract Oncol 5:531–542CrossRefPubMedGoogle Scholar
  46. Singh JK, Simoes BM, Howell SJ, Farnie G, Clarke RB (2013) Recent advances reveal IL-8 signaling as a potential key to targeting breast cancer stem cells. Breast Cancer Res 15:210CrossRefPubMedPubMedCentralGoogle Scholar
  47. Slamon D, Clark G, Wong S, Levin W, Ullrich A, Mcguire W (1987) Human breast cancer: correlation of relapse and survival with amplification of the HER-2/neu oncogene. Science 235:177–182CrossRefPubMedGoogle Scholar
  48. Song G, Ouyang G, Bao S (2005) The activation of Akt/PKB signaling pathway and cell survival. J Cell Mol Med 9:59–71CrossRefPubMedGoogle Scholar
  49. Sorlie T, Tibshirani R, Parker J, Hastie T, Marron JS, Nobel A, Deng S, Johnsen H, Pesich R, Geisler S, Demeter J, Perou CM, Lonning PE, Brown PO, Borresen-Dale AL, Botstein D (2003) Repeated observation of breast tumor subtypes in independent gene expression data sets. Proc Natl Acad Sci USA 100:8418–8423CrossRefPubMedPubMedCentralGoogle Scholar
  50. Stingl J, Caldas C (2007) Molecular heterogeneity of breast carcinomas and the cancer stem cell hypothesis. Nat Rev Cancer 7:791–799CrossRefPubMedGoogle Scholar
  51. Tang CH, Lu DY, Yang RS, Tsai HY, Kao MC, Fu WM, Chen YF (2007) Leptin-induced IL-6 production is mediated by leptin receptor, insulin receptor substrate-1, phosphatidylinositol 3-kinase, Akt, NF-kappaB, and p300 pathway in microglia. J Immunol 179:1292–1302CrossRefPubMedGoogle Scholar
  52. Thiagalingam S, Lengauer C, Leach FS, Schutte M, Hahn SA, Overhauser J, Willson JK, Markowitz S, Hamilton SR, Kern SE, Kinzler KW, Vogelstein B (1996) Evaluation of candidate tumour suppressor genes on chromosome 18 in colorectal cancers. Nat Genet 13:343–346CrossRefPubMedGoogle Scholar
  53. Uchima Y, Sawada T, Nishihara T, Maeda K, Ohira M, Hirakawa K (2004) Inhibition and mechanism of action of a protease inhibitor in human pancreatic cancer cells. Pancreas 29:123–131CrossRefPubMedGoogle Scholar
  54. Uwagawa T, Li Z, Chang Z, Xia Q, Peng B, Sclabas GM, Ishiyama S, Hung MC, Evans DB, Abbruzzese JL, Chiao PJ (2007) Mechanisms of synthetic serine protease inhibitor (FUT-175)-mediated cell death. Cancer 109:2142–2153CrossRefPubMedGoogle Scholar
  55. Uwagawa T, Misawa T, Tsutsui N, Ito R, Gocho T, Hirohara S, Sadaoka S, Yanaga K (2013) Phase II study of gemcitabine in combination with regional arterial infusion of nafamostat mesilate for advanced pancreatic cancer. Am J Clin Oncol 36:44–48CrossRefPubMedGoogle Scholar
  56. Woolf DK, Padhani AR, Makris A (2015) Assessing response to treatment of bone metastases from breast cancer: what should be the standard of care? Ann Oncol 26:1048–1057CrossRefPubMedGoogle Scholar

Copyright information

© The Pharmaceutical Society of Korea 2017

Authors and Affiliations

  • Sunam Mander
    • 1
  • Dong-Joo You
    • 1
  • Sumi Park
    • 1
  • Dong Hwi Kim
    • 1
  • Hyo Jeong Yong
    • 1
  • Dong-Sik Kim
    • 2
  • Curie Ahn
    • 3
  • Yun-Hee Kim
    • 4
  • Jae Young Seong
    • 1
  • Jong-Ik Hwang
    • 1
  1. 1.Department of Biomedical Sciences, College of MedicineKorea UniversitySeoulRepublic of Korea
  2. 2.Department of HBP Surgery and Liver Transplantation, College of MedicineKorea UniversitySeoulRepublic of Korea
  3. 3.Transplantation Research Institute, Seoul National UniversitySeoulRepublic of Korea
  4. 4.Molecular Imaging & Therapy BranchResearch Institute of National Cancer CenterGoyangRepublic of Korea

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