Clinical & Experimental Metastasis

, Volume 22, Issue 4, pp 351–361 | Cite as

A New Model for Lymphatic Metastasis: Development of a Variant of the MDA-MB-468 Human Breast Cancer Cell Line that Aggressively Metastasizes to Lymph Nodes

  • Sharon A. Vantyghem
  • Alison L. Allan
  • Carl O. Postenka
  • Waleed Al-Katib
  • Michael Keeney
  • Alan B. Tuck
  • Ann F. Chambers


Breast cancer often spreads from the primary tumor to regional lymph nodes. Lymph node status provides clinically important information for making treatment decisions. Spread via lymphatics is also important for the biology of breast cancer, as tumor cells in lymph nodes may provide a reservoir of cells leading to distant, lethal metastases. Improved understanding of the biology of lymphatic spread thus is important for improved breast cancer survival. Advances towards understanding the interactions between tumors cells and lymphatic vessels have in part been limited by the lack of suitable cell lines and experimental models. We have addressed this need by developing a new model of lymphatic metastasis. Here we describe the establishment of 468LN cells, a variant of the MDA-MB-468 human breast adenocarcinoma cell line, which produces extensive lymph node metastasis following orthotopic injection of nude mice. 468LN cells are also more aggressive in vitro, produce more osteopontin and express different surface integrins compared to the parent line. The dramatic in vitro and in vivo phenotypic and molecular differences of 468LN and parental 468GFP cells make this pair of cell lines a unique model for the specific study of lymph node metastasis of breast cancer.


breast cancer lymph node metastasis orthotopic model 



extracellular matrix


epidermal growth factor receptor


green fluorescent protein


mammary fat pad




vascular cell adhesion molecule


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  1. 1.
    Fisher, B, Bauer, M, Wickerham, DL,  et al. 1983Relation of number of positive nodes to the prognosis of patients with primary breast cancer. An N.S.A.B.P. updateCancer52155157PubMedGoogle Scholar
  2. 2.
    Foster, RS,Jr 1996The biologic and clinical significance of lym- phatic metastases in breast cancerSurg Oncol Clin N Am579104PubMedGoogle Scholar
  3. 3.
    Carter, CL, Allen, C, Henson, DE 1989Relation of tumor size, lymph node status, and survival in 24,740 breast cancer casesCancer6318187PubMedGoogle Scholar
  4. 4.
    Ranaboldo, CJ, Mitchel, A, Royle, GT,  et al. 1993Axillary nodal status in women with screen-detected breast cancerEur J Surg Oncol1913033PubMedGoogle Scholar
  5. 5.
    Greenlee, RT, Hill-Harmon, MB, Murray, T, Thun, M 2001Cancer statistics, 2001CA Cancer J Clin511536PubMedGoogle Scholar
  6. 6.
    Fidler, IJ 2001Molecular biology of cancer: invasion and metastasisDevita, VT,Jr.Hellman, SRosenberg, SA eds. Cancer, Principles and Practice of Oncology6Lippincott Williams and WilkinsPhiladelphia13553Google Scholar
  7. 7.
    Trappen, PO, Pepper, MS 2002Lymphatic dissemination of tumor cells and the formation of micrometastasesLancet Oncol34452CrossRefPubMedGoogle Scholar
  8. 8.
    McGuire, WL 1987Prognostic factors for recurrence and survival in human breast cancerBreast Cancer Res1059CrossRefGoogle Scholar
  9. 9.
    Nemoto, T, Vana, J, Bedwani, RN,  et al. 1980Management and survival of female breast cancer: results of a national survey by the American College of SurgeonsCancer45291724PubMedGoogle Scholar
  10. 10.
    Chambers, AF, Groom, AC, MacDonald, IC 2002Dissemination and growth of cancer cells in metastatic sitesNat Rev Cancer856372CrossRefGoogle Scholar
  11. 11.
    Pantel, K, Brakenhoff, RH 2004Dissecting the metastatic cascadeNat Rev Cancer.444856CrossRefPubMedGoogle Scholar
  12. 12.
    Liotta, LA, Stetler-Stevenson, WG 1993Principles of molecular cell biology of cancer: Cancer metastasisDevita, VT,JrHellman, SRosenberg, SA eds. Cancer, Principles and Practice of Oncology4JB LippincottPhiladelphia13449Google Scholar
  13. 13.
    Cunnick, GH, Jiang, WG, Gomez, KF, Mansel, RE 2002Lymphangiogenesis and breast cancer metastasisHistol Histopathol1786370PubMedGoogle Scholar
  14. 14.
    Pickren, JW 1956Lymph node metastases in carcinoma of the female mammary glandBull Roswell Park Mem Inst17985Google Scholar
  15. 15.
    Stacker, SA, Baldwin, ME, Achen, MG 2002The role of tumor lym- phangiogenesis in metastatic spreadFASEB J1692234CrossRefPubMedGoogle Scholar
  16. 16.
    Oliver, G, Detmar, M 2002The rediscovery of the lymphatic system: old and new insights into the development and biological function of the lymphatic vasculatureGenes Dev1677383CrossRefPubMedGoogle Scholar
  17. 17.
    Carr, I 2004Lymphatic metastasisBr J Cancer9075255CrossRefPubMedGoogle Scholar
  18. 18.
    Sleeman, JP 2000The lymph node as a bridgehead in the metastatic dissemination of tumorsRecent Results Cancer Res1575581PubMedGoogle Scholar
  19. 19.
    McCarter, MD, Clarke, JH, Harken, AH 2004Lymphangiogenesis is pivotal to the trials of a successful cancer metastasisSurgery13512124CrossRefPubMedGoogle Scholar
  20. 20.
    Jeltsch, M, Tammela, T, Alitalo, K, Wilting, J 2003Genesis and pathogenesis of lymphatic vesselsCell Tissue Res3146984CrossRefPubMedGoogle Scholar
  21. 21.
    Price, JE, Polyzos, A, Zhang, RD, Daniels, LM 1990Tumorigenicity and metastasis of human breast carcinoma cell lines in nude miceCancer Res5071721PubMedGoogle Scholar
  22. 22.
    Price, JE, Zhang, RD 1990Studies of human breast cancer metastasis using nude miceCancer Metast Rev828597CrossRefGoogle Scholar
  23. 23.
    Zhang, RD, Fidler, IJ, Price, JE 1991Relative malignant potential of human breast carcinoma cell lines established from pleural effusions and a brain metastasisInvasion Metast1120415Google Scholar
  24. 24.
    Welch, DR 1997Technical considerations for studying cancer metastasis in vivoClin Exp Metast15272306CrossRefGoogle Scholar
  25. 25.
    Cailleau, R, Olive, M, Cruciger, QVJ 1978Long-term human breast carcinoma cell lines of metastatic origin: preliminary characterizationIn Vitro1491115PubMedGoogle Scholar
  26. 26.
    Naumov, GN, MacDonald, IC, Weinmeister, PM,  et al. 2002Persistence of solitary mammary carcinoma cells in a secondary site: A possible contributor to dormancyCancer Res62216268PubMedGoogle Scholar
  27. 27.
    Vantyghem, SA, Wilson, SM, Postenka, CO,  et al. 2005Dietary genistein reduces metastasis in a post-surgical orthotopic breast cancer modelCancer Res653396403PubMedGoogle Scholar
  28. 28.
    Bautista, DS, Xuan, JW, Hota, C,  et al. 1994Inhibition of Arg–Gly–Asp (RGD)-mediated cell adhesion to osteopontin by a monoclonal antibody against osteopontinJ Biol Chem2692328085PubMedGoogle Scholar
  29. 29.
    Tuck, AB, Arsenault, DM, O’Malley, FP,  et al. 1999Osteopontin induces increased invasiveness and plasminogen activator expression of human mammary epithelial cellsOncogene18423746CrossRefPubMedGoogle Scholar
  30. 30.
    Stacker, SA, Achen, MG, Jussila, L,  et al. 2002Lymphangiogenesis and cancer metastasisNat Rev Cancer257383CrossRefPubMedGoogle Scholar
  31. 31.
    Fidler, IJ 1973Selection of successive tumor lines for metastasisNature24214849Google Scholar
  32. 32.
    Poste, G 1982Experimental systems for analysis of the malignant phenotypeCancer Metast Rev114199CrossRefGoogle Scholar
  33. 33.
    Clark, EA, Golub, TR, Lander, ES, Hynes, RO 2000Genomic analysis of metastasis reveals an essential role for RhoCNature40653235CrossRefPubMedGoogle Scholar
  34. 34.
    Weiss, L 1983Random and nonrandom processes in metastasis, and metastatic inefficiencyInvasion Metast3193207Google Scholar
  35. 35.
    Hill, RP, Chambers, AF, Ling, V, Harris, JF 1984Dynamic heterogeneity: rapid generation of metastatic variants in mouse B16 melanoma cellsScience2249981001PubMedGoogle Scholar
  36. 36.
    Yu, JL, Rak, JW, Carmeliet, P,  et al. 2001Heterogeneous vascular dependence of tumor cell populationsAm J Pathol158132534PubMedGoogle Scholar
  37. 37.
    Boyer, B, Valles, AM, Edme, N 2000Induction and regulation of epithelial-mesenchymal transitionsBiochem Pharmacol60109199CrossRefPubMedGoogle Scholar
  38. 38.
    Savagner, P 2001Leaving the neighborhood. Molecular mechanisms involved during epithelial-mesenchymal transitionBioessays2391223CrossRefPubMedGoogle Scholar
  39. 39.
    Nicolson, GL 1988Cancer metastasis: tumor cell and host organ properties important in metastasis to specific secondary sitesBiochim Biophys Acta948175224PubMedGoogle Scholar
  40. 40.
    Fidler, IJ 2002The organ microenvironment and cancer metastasisDifferentiation70498505CrossRefPubMedGoogle Scholar
  41. 41.
    Yeatman, TJ, Nicolson, GL 1993Molecular basis of tumor progression: mechanisms of organ-specific tumor metastasisSemin Surg Oncol925663PubMedGoogle Scholar
  42. 42.
    Liotta, LA, Kohn, EC 2001The microenvironment of the tumour-host interfaceNature41137579CrossRefPubMedGoogle Scholar
  43. 43.
    Hood, JD, Cheresh, DA 2002Role of integrins in cell invasion and migrationNat Rev Cancer291100CrossRefPubMedGoogle Scholar
  44. 44.
    Jin, H, Varner, J 2004Integrins: Roles in cancer development and as treatment targetsBr J Cancer9056165CrossRefPubMedGoogle Scholar
  45. 45.
    Jones, JL, Walker, RA 1999Integrins: a role as cell signalling moleculesMol Pathol5220813PubMedGoogle Scholar
  46. 46.
    Coppolino, MG, Dedhar, S 2000Bi-directional signal transduction by integrin receptorsInt J Biochem Cell Biol,3217188Google Scholar
  47. 47.
    Tuck, AB, Chambers, AF 2001The role of osteopontin in breast cancer: clinical and experimental studiesJ Mammary Gland Biol Neoplasia441929CrossRefGoogle Scholar
  48. 48.
    Rittling, SR, Chambers, AF 2004Role of osteopontin in tumour progressionBr J Cancer90187781CrossRefPubMedGoogle Scholar
  49. 49.
    Denhardt, DV, Guo, X 1993Osteopontin. A protein with diverse functionsFASEB J7147582PubMedGoogle Scholar
  50. 50.
    Bellahcene, A, Castronovo, V 1995Increased expression of osteonectin and osteopontin, two bone matrix proteins, in human breast cancerAm J Pathol14695100PubMedGoogle Scholar
  51. 51.
    O’Regan, A, Berman, JS 2000Osteopontin: A key cytokine in cell-mediated and granulomatous inflammationInt J Exp Pathol8137390CrossRefPubMedGoogle Scholar
  52. 52.
    Tuck, AB, Elliott, BE, Hota, C,  et al. 2000Osteopontin-induced, integrin-dependent migration of human mammary epithelial cells involves activation of the hepatocyte growth factor receptor (Met)J Cell Biochem7846575CrossRefPubMedGoogle Scholar
  53. 53.
    Furger, KA, Allan, AL, Wilson, SM,  et al. 2003Beta(3) integrin expression increases breast carcinoma cell responsiveness to the malignancy-enhancing effects of osteopontinMol Cancer Res181019PubMedGoogle Scholar
  54. 54.
    Smith, LL, Cheung, HK, Ling, LE,  et al. 1996Osteopontin N-terminal domain contacts a cryptic adhesive sequence recognized by α9β1 integrinJ Biol Chem2712848591CrossRefPubMedGoogle Scholar
  55. 55.
    Yokosaki, Y, Matsuura, N, Sasaki, T,  et al. 1999The integrin α9β1 binds to a novel recognition sequence (SVVYGLR) in the thrombin-cleaved amino-terminal fragment of osteopontinJ Biol Chem2743632834CrossRefPubMedGoogle Scholar
  56. 56.
    Yokosaki, Y, Palmer, EL, Prieto, AL,  et al. 1994The integrin α9β1 mediates cell attachment to a non-RGD site in the third fibronectin type III repeat of tenascinJ Biol Chem2692669196PubMedGoogle Scholar
  57. 57.
    Taooka, Y, Chen, J, Yednock, T, Sheppard, D,  et al. 1999The integrin α9β1 mediates adhesion to activated endothelial cells and transendothelial neutrophil migration through interaction with vascular cell adhesion molecule-1J Cell Biol14541320CrossRefPubMedGoogle Scholar
  58. 58.
    Vlahakis, NE, Young, BA, Atakilit, A, Sheppard, D 2005The lymphangiogenic vascular endothelial growth factors VEGF-C and -D are ligands for the integrin α9β1J Biol Chem280454452CrossRefPubMedGoogle Scholar

Copyright information

© Springer 2005

Authors and Affiliations

  • Sharon A. Vantyghem
    • 1
    • 2
    • 3
  • Alison L. Allan
    • 1
    • 3
  • Carl O. Postenka
    • 1
  • Waleed Al-Katib
    • 1
  • Michael Keeney
    • 4
    • 5
  • Alan B. Tuck
    • 1
    • 2
    • 3
    • 4
  • Ann F. Chambers
    • 1
    • 2
    • 3
    • 5
  1. 1.London Regional Cancer ProgramOntarioCanada
  2. 2.Department of PathologyUniversity of Western OntarioOntarioCanada
  3. 3.Department of OncologyUniversity of Western OntarioOntarioCanada
  4. 4.London Health Sciences CentreOntarioCanada
  5. 5.Lawson Health Research InstituteOntarioCanada

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