Advertisement

Journal of Neuro-Oncology

, Volume 81, Issue 1, pp 39–48 | Cite as

Differential Expression of Matrix Metalloproteinases in Brain- and Bone-Seeking Clones of Metastatic MDA-MB-231 Breast Cancer Cells

  • Andreas M. Stark
  • Bartosz Anuszkiewicz
  • Rolf Mentlein
  • Toshiyuki Yoneda
  • H. Maximilian Mehdorn
  • Janka Held-FeindtEmail author
Lab Investigation

Abstract

Matrix Metalloproteinases (MMPs) play a crucial role in breast cancer metastasis. We examined the mRNA and protein expression of several MMPs in brain- and bone-seeking clones of MDA-MB-231 breast cancer cells, their transcriptional regulation and their functional role in the metastatic process. MMP mRNA expression was examined using real-time reverse transcription polymerase chain reaction. Protein expression was examined using enzyme linked immunosorbent essay (ELISA). The inducibility of mRNA and protein expression was tested with TPA (phorbol 12-myristate 13-acetate; 50 µM); epidermal growth factor and transforming growth factor β (20 ng/ml both). Migration and invasion assays were performed with the QCM™ 96-Well Migration/Invasion Assay (8 µm; Chemicon) over 24 h with or without specific MMPs inhibitors (MMP Inhibitor I Mix (5 µM); MMP-2/MMP-9 Inhibitor III (50 µM); EMD Biosciences). We found significantly higher mRNA expression of MMP-1 and -9 in brain-seeking 231-clones in comparison to -bone and -parental cells. In contrast, the mRNA expression of MMP-3 and -14 was comparable in all cells lines examined and MMP-13 expression was lower in both selective metastatic lines. MMP-2 and -8 were not expressed. ELISA revealed a higher amount of total as well as active MMP-1 and -9 in brain-seeking cells. TPA stimulation showed that MMP-1 and -9 transcription was inducible on the mRNA and protein level in 231-parental but not in 231-brain or -bone. 231-brain showed the highest migration and invasive capacity which could be decreased by the application of MMP-1 and/or MMP-9 inhibitor. Our results indicate functional importance of MMP-1 and -9 overexpression in brain metastasis in an in vitro model.

Keywords

Breast Cancer Brain Metastases MMP Invasion Function 

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

Notes

Acknowledgements

We thank Brigitte Rehmke, Bärbel Hufnagel and Jörg Krause for their expert technical assistance. This work was supported by the “Hensel-Foundation”.

References

  1. 1.
    Arbit E, Wronski M (1996) Clinical decision making in brain metastases. Neurosurg Clin N Am 7:447–457PubMedGoogle Scholar
  2. 2.
    Schouten LJ, Rutten J, Huveneers HA, Twijnstra A (2002) Incidence of brain metastases in a cohort of patients with carcinoma of the breast, colon, kidney, and lung and melanoma. Cancer 94:2698–2707PubMedCrossRefGoogle Scholar
  3. 3.
    Sloan EK, Anderson RL (2002) Genes involved in breast cancer metastasis to bone. Cell Mol Life Sci 59:1491–1502PubMedCrossRefGoogle Scholar
  4. 4.
    Stark AM, Tscheslog H, Buhl R, Held-Feindt J, Mehdorn HM (2005) Surgical treatment for brain metastases: prognostic factors and survival in 177 patients. Neurosurg Rev 28:115–119PubMedCrossRefGoogle Scholar
  5. 5.
    Puduvalli VK (2001) Brain metastases: biology and the role of the brain microenvironment. Curr Oncol Rep 3:467–475PubMedGoogle Scholar
  6. 6.
    Weber GF, Ashkar S (2000) Molecular mechanisms of tumor dissemination in primary and metastatic brain cancers. Brain Res Bull 53:421–424PubMedCrossRefGoogle Scholar
  7. 7.
    Yoneda T, Williams PJ, Hiraga T, Niewolna M, Nishimura R (2001) A bone-seeking clone exhibits different biological properties from the MDA-MB-231 parental human breast cancer cells and a brain-seeking clone in vivo and in vitro. J Bone Mineral Res 16:1486–1495CrossRefGoogle Scholar
  8. 8.
    Jackson JG, Zhang X, Yoneda T, Yee D (2001) Regulation of breast cancer cell motility by insulin like receptor substrate-2 (IRS-2) in metastatic variants of human breast cancer cell lines. Oncogene 20:7318–7325PubMedCrossRefGoogle Scholar
  9. 9.
    Myoui A, Nishimura R, Williams PJ, Hiraga T, Tamura D, Michigami T, Mundy GR, Yoneda T (2003) C-SRC tyrosine kinase activity is associated with tumor colonization in bone and lung in an animal model of human breast cancer metastasis. Cancer Res 63:5028–5033PubMedGoogle Scholar
  10. 10.
    Weil RJ, Palmieri DC, Bronder JL, Stark AM, Steeg PS (2005) Breast cancer metastasis to the central nervous system. Am J Pathol 167:913–920PubMedGoogle Scholar
  11. 11.
    Visse R, Nagase H (2003) Matrix metalloproteinases and tissue inhibitors of metalloproteinases, structure, function and biochemistry. Circ Res 92:827–839PubMedCrossRefGoogle Scholar
  12. 12.
    Woessner JF (1998) The matrix metalloproteinase family. In: Parks WC, Mecham RP (eds) Matrix metalloproteinases. San Diego, California, Academic Press, pp 1–13Google Scholar
  13. 13.
    Hamacher S, Matern S, Roeb E (2004) Extracellular matrix – from basic research to clinical significance. An overview with special consideration of matrix metalloproteinases. Dtsch Med Wochenschr 129:1976–1980PubMedCrossRefGoogle Scholar
  14. 14.
    Brinckerhoff CE, Rutter JL, Benbow U (2000) Interstitial collagenases as markers of tumor progression. Clin Cancer Res 6:4823–4830PubMedGoogle Scholar
  15. 15.
    Morgan H, Hill PA (2005) Human breast cancer cell-mediated bone collagen degradation requires plasminogen activation and matrix metalloproteinase activity. Cancer Cell Int 5:1PubMedCrossRefGoogle Scholar
  16. 16.
    Wiesen JF, Werb Z (1996) The role of stromelysin-1 in stromal–epithelial interactions and cancer. Enzyme Protein 49:174–181PubMedGoogle Scholar
  17. 17.
    Rio MC, Lefebvre O, Santavicca M, Noel A, Chenard MP, Anglard P, Byrne JA, Okada A, Regnier CH, Masson R, Bellocq JP, Basset P (1996) Stromelysin-3 in the biology of the normal and neoplastic mammary gland. J Mam Gland Neoplasia 1:231–240CrossRefGoogle Scholar
  18. 18.
    Lochter A, Srebrow A, Sympson CJ, Terracio N, Werb Z, Bissell MJ (1997) Misregulation of stromelysin-1 expression in mouse mammary tumor cells accompanies acquisition of stromelysin-1-dependent invasive properties. J Biol Chem 272:5007–5015PubMedCrossRefGoogle Scholar
  19. 19.
    Davies B, Miles DW, Happerfield LC, Naylor MS, Bobrow LG, Rubens RD, Balkwill FR (1993) Activity of type IV collagenases in benign and malignant breast disease. Br J Cancer 67:1126–1131PubMedGoogle Scholar
  20. 20.
    Duffy MJ, Maguire TM, Hill A, McDermott E, O’Higgins N (2000) Metalloproteinases: role in breast carcinogenesis, invasion and metastasis. Breast Cancer Res 2:252–257PubMedCrossRefGoogle Scholar
  21. 21.
    Feindt J, Becker I, Blömer U, Hugo HH, Mehdorn HM, Krisch B, Mentlein R (1995) Expression of somatostatin receptor subtypes in cultured astrocytes and gliomas. J Neurochem 65:1997–2005PubMedCrossRefGoogle Scholar
  22. 22.
    Baker AH, Edwards DR, Murphy G (2002) Metalloproteinase inhibitors: biological actions and therapeutic opportunities. J Cell Sci 115:3719–3727PubMedCrossRefGoogle Scholar
  23. 23.
    Mendes O, Kim HT, Stoica G (2005) Expression of MMP2, MMP9 and MMP3 in breast cancer brain metastasis in a rat model. Clin Exp Metastasis 22:237–246PubMedCrossRefGoogle Scholar
  24. 24.
    Bachmeier BE, Nerlich AG, Lichtinghagen R, Sommerhofr CP (2001) Matrix metalloproteinases (MMPs) in breast cancer cell lines of different tumorigenicity. Anticancer Res 21:2821–2828Google Scholar
  25. 25.
    Barrett JM, Puglia MA, Singh G, Tozer RG (2002) Expression of Ets-related transcription factors and matrix metalloproteinase genes in human breast cancer cells. Breast Cancer Res Treat 72:227–232PubMedCrossRefGoogle Scholar
  26. 26.
    Behrens P, Rothe M, Wellmann A, Krischler J, Wernert T (2001) The Ets-1 transcription factor is up-regulated together with MMP1 and MMP9 in the stroma of pre-invasive breast cancer. J Pathol 194:43–50PubMedCrossRefGoogle Scholar
  27. 27.
    Mackay AR, Ballin M, Pelina MD, Farina AR, Nason AM, Hartzler JL, Thorgeirsson UP (1992) Effect of phorbol ester and cytokines on matrix metalloproteinase and tissue inhibitor of metalloproteinase expression in tumor and normal cell lines. Invasion Metastasis 12:168–184PubMedGoogle Scholar
  28. 28.
    Wang TN, Albo D, Tuszynski GP (2002) Fibroblasts promote breast cancer cell invasion by upregulating tumor matrix metalloproteinase-9 production. Surgery 132:220–225PubMedCrossRefGoogle Scholar
  29. 29.
    Duivenvoorden WC, Hirte HW, Singh G (1999) Transforming growth factor beta1 acts as an inducer of matrix metalloproteinase expression and activity in human bone-metastasizing cells. Clin Exp Metastasis 17:27–34PubMedCrossRefGoogle Scholar
  30. 30.
    Gadher SJ, Schmid TM, Heck LW, Woolley DE (1989) Cleavage of collagen type X by human synovial collagenase and neutrophil elastase. Matrix 9:109–115PubMedGoogle Scholar
  31. 31.
    Goldring MB (1993) Degradation of articular cartilage in culture: regulatory factors. In: Woessner JF, Howell DS (eds) Joint cartilage degradation. Marcel Dekker, New York, pp 281–345Google Scholar
  32. 32.
    Vincenti MP, Coon CI, White LA, Barchowsky A, Brinckerhoff CE (1996) src-Related tyrosine kinases regulate transcriptional activation of the intestinal collagenase gene, MMP-1, in interleukin-1 stimulated synovial fibroblasts. Arthritis Rheum 39:574–582PubMedGoogle Scholar
  33. 33.
    Nutt JE, Lunec J (1996) Induction of metalloproteinase (MMP1) expression by epidermal growth factor (EGF) receptor stimulation and serum deprivation in human breast tumor cells. Eur J Cancer 32:2127–2135CrossRefGoogle Scholar
  34. 34.
    Kousidou OC, Roussidis AE, Theocharis AD, Karamanos NK (2004) Expression of MMPs and TIMPs genes in human breast cancer epithelial cells depends on cell culture conditions and is associated with their invasive potential. Anticancer Res 24(6):4025–4030PubMedGoogle Scholar
  35. 35.
    Poola I, DeWitty RL, Marshalleck JJ, Bhatnagar R, Abraham J, Leffall LD (2005) Identification of MMP-1 as a piútative breast cancer predictive marker by global gene expression analysis. Nat Med 11:481–483PubMedCrossRefGoogle Scholar
  36. 36.
    Bachmeier BE, Rohrbach H, De Waal J, Jochum M, Nerlich AG (2005) Enhanced expression and activation of major matrix metalloproteinases in distinct topographic areas of invasive breast carcinomas. Int J Oncol 26:1203–1207PubMedGoogle Scholar
  37. 37.
    Proost P, Van Damme J, Opdenakker G (1993) Leukocyte gelatinase B cleavage releases encephalitogens from human myelin basic protein. Biochem Biophys Res Commun 192:1175–1181PubMedCrossRefGoogle Scholar
  38. 38.
    Witty JP, Foster SA, Stricklin GP, Matrisian LM, Stern PH (1996) Parathyroid hormone-induced resorption in fetal rat limb bones with production of the metalloproteinases collagenase and gelatinase B. J Bone Miner Res 11:72–78PubMedCrossRefGoogle Scholar
  39. 39.
    Kjeldsen L, Bjerrum OW, Askaa J, Borregaard N (1992) Subcellular localization and release of human neutrophil gelatinase, confirming the existence of separate gelatinase-containing granules. Biochem J 287:603–610PubMedGoogle Scholar
  40. 40.
    Welgus HG, Campbell EJ, Cury JD, Eisen AZ, Senior RM, Wilhelm SM, Goldberg GI (1991) Neutral metalloproteinases produced by human mononuclear phagocytes. Enzyme profile, regulation, and expression during cellular development. J Clin Invest 86:1496–1502CrossRefGoogle Scholar
  41. 41.
    Rosenberg GA (1995) Matrix metalloproteinases in brain injury. J Neurotrauma 12:833–842PubMedCrossRefGoogle Scholar
  42. 42.
    Arnold SM, Young AB, Munn RK, Patchell RA, Nanayakkara N, Markesbery WR (1999) Expression of p53, bcl-2, E-cadherin, matrix metalloproteinase-9, and tissue inhibitor of metalloproteinases-1 in paired primary tumors and brain metastasis. Clin Cancer Res 5:4028–4033PubMedGoogle Scholar
  43. 43.
    Rahko E, Jukkola A, Melkko J, Paavo P, Bloigu R, Talvensaari-Mattila A, Turpeenniemi-Hujanen T (2004) Matrix metalloproteinase-9 (MMP-9) immunoreactive protein has modest prognostic value in locally advanced breast carcinoma patients treated with an adjuvant antiestrogen therapy. Anticancer Res 24:4247–4253PubMedGoogle Scholar
  44. 44.
    Barsky SH, Togo S, Garbisa S, Liotta LA (1983) Type IV collagenase immunoreactivity in invasive breast carcinoma. Lancet 5:296–297CrossRefGoogle Scholar
  45. 45.
    Przybylowska K, Kluczna A, Zadrozny M, Krawczyk T, Kulig A, Rykala J, Kolacinska A, Morawiec Z, Drzewoski J, Blasiak J (2005) Polymorphisms of the promoter regions of matrix metalloproteinases genes MMP-1 and MMP-9 in breast cancer. Breast Cancer Res 95:65–72Google Scholar
  46. 46.
    Balduyck M, Zerimech F, Gouyer V, Lemaire R, Hemon B, Grard G, Thiebaut C, Lemaire V, Dacquembronne E, Duhem T, Lebrun A, Dejonghe MJ, Huet G (2000) Specific expression of matrix metalloproteinases 1, 3, 9 and 13 associated with invasiveness of breast cancer cells in vitro. Clin Exp Metastasis 18:171–178PubMedCrossRefGoogle Scholar
  47. 47.
    Saad S, Bendall LJ, James A, Gottlieb DJ, Bradstock KF (2000) Induction of matrix metalloproteinases MMP-1 and MMP-2 by coculture of breast cancer cells and bone marrow fibroblasts. Breast Cancer Res Treat 63:105–115PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2006

Authors and Affiliations

  • Andreas M. Stark
    • 1
  • Bartosz Anuszkiewicz
    • 1
  • Rolf Mentlein
    • 2
  • Toshiyuki Yoneda
    • 3
  • H. Maximilian Mehdorn
    • 1
  • Janka Held-Feindt
    • 1
    • 4
    Email author
  1. 1.Department of NeurosurgeryUniversity of KielKielGermany
  2. 2.Institute of Anatomy University of KielKielGermany
  3. 3.Division of Endocrinology and MetabolismUniversity of Texas Health Science CenterSan AntonioUSA
  4. 4.Department of Neurosurgery University of Schleswig-Holstein Medical CenterKielGermany

Personalised recommendations