Skip to main content

Therapeutic metformin/AMPK activation promotes the angiogenic phenotype in the ERα negative MDA-MB-435 breast cancer model

Breast Cancer Research and Treatment volume 113, pages 101–111 (2009)Cite this article

Abstract

Metformin, a first line treatment for type 2 diabetes, has been implicated as a potential anti-neoplastic agent for breast cancers as well as other cancers. Metformin is known to work in part through the activation of AMP-dependent kinase (AMPK). AMPK is a key regulator of cellular energy homeostasis, especially under stress conditions where biosynthetic pathways are blocked by the phosphorylation of downstream AMPK substrates. Stimulation of AMPK by metformin resulted in a significant repression of cell proliferation and active MAPK1/2 in both estrogen receptor α (ERα) negative (MDA-MB-231, MDA-MB-435) and positive (MCF-7, T47D) human breast cancer cell lines. However, when ERα negative MDA-MB-435 cells were treated with metformin, they demonstrated increased expression of vascular endothelial growth factor (VEGF) in an AMPK dependent manner; while the ERα positive MCF-7 cells did not. Systemic therapy with metformin was tested for efficacy in an orthotopic model of ERα negative breast cancer performed in athymic nude mice. Surprisingly, metformin therapy significantly improved tumorigenic progression as compared to untreated controls. The metformin-treated group showed increased VEGF expression, intratumoral microvascular density and reduced necrosis. Metformin treatment was sufficient, however, to reduce systemic IGF-1 and the proliferation rate of tumor cells in vascularized regions. The data presented here suggests that, although metformin significantly represses breast cancer cell growth in vitro, the efficacy with respect to its therapeutic application for ERα negative breast cancer lesions in vivo may result in promotion of the angiogenic phenotype and increased tumorigenic progression.

This is a preview of subscription content, access via your institution.

Access options

Buy single article

Instant access to the full article PDF.

43,34 €

Price includes VAT (Finland)
Tax calculation will be finalised during checkout.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

References

  1. Kirpichnikov D, McFarlane SI, Sowers JR (2002) Metformin: an update. Ann Intern Med 137:25–33

    PubMed  CAS  Google Scholar 

  2. DeFronzo RA, Goodman AM (1995) Efficacy of metformin in patients with non-insulin-dependent diabetes mellitus. The Multicenter Metformin Study Group. N Engl J Med 333:541–549

    PubMed  Article  CAS  Google Scholar 

  3. Hawley SA, Gadalla AE, Olsen GS, Hardie DG (2002) The antidiabetic drug metformin activates the AMP-activated protein kinase cascade via an adenine nucleotide-independent mechanism. Diabetes 51:2420–2425

    PubMed  Article  CAS  Google Scholar 

  4. Towler MC, Hardie DG (2007) AMP-activated protein kinase in metabolic control and insulin signaling. Circ Res 100:328–341

    PubMed  Article  CAS  Google Scholar 

  5. Zhou G, Myers R, Li Y, Chen Y, Shen X, Fenyk-Melody J, Wu M, Ventre J, Doebber T, Fujii N, Musi N, Hirshman MF, Goodyear LJ, Moller DE (2001) Role of AMP-activated protein kinase in mechanism of metformin action. J Clin Invest 108:1167–1174

    PubMed  CAS  Google Scholar 

  6. Zou MH, Kirkpatrick SS, Davis BJ, Nelson JS, Wiles WGT, Schlattner U, Neumann D, Brownlee M, Freeman MB, Goldman MH (2004) Activation of the AMP-activated protein kinase by the anti-diabetic drug metformin in vivo. Role of mitochondrial reactive nitrogen species. J Biol Chem 279:43940–43951

    PubMed  Article  CAS  Google Scholar 

  7. Bowker SL, Majumdar SR, Veugelers P, Johnson JA (2006) Increased cancer-related mortality for patients with type 2 diabetes who use sulfonylureas or insulin. Diabetes Care 29:254–258

    PubMed  Article  Google Scholar 

  8. Evans JM, Donnelly LA, Emslie-Smith AM, Alessi DR, Morris AD (2005) Metformin and reduced risk of cancer in diabetic patients. BMJ 330:1304–1305

    PubMed  Article  Google Scholar 

  9. Zakikhani M, Dowling R, Fantus IG, Sonenberg N, Pollak M (2006) Metformin is an AMP kinase-dependent growth inhibitor for breast cancer cells. Cancer Res 66:10269–10273

    PubMed  Article  CAS  Google Scholar 

  10. Isakovic A, Harhaji L, Stevanovic D, Markovic Z, Sumarac-Dumanovic M, Starcevic V, Micic D, Trajkovic V (2007) Dual antiglioma action of metformin: cell cycle arrest and mitochondria-dependent apoptosis. Cell Mol Life Sci 64:1290–1302

    PubMed  Article  CAS  Google Scholar 

  11. Baumann P, Mandl-Weber S, Emmerich B, Straka C, Schmidmaier R (2007) Activation of adenosine monophosphate activated protein kinase inhibits growth of multiple myeloma cells. Exp Cell Res 313:3592–3603

    PubMed  Article  CAS  Google Scholar 

  12. Saha AK, Persons K, Safer JD, Luo Z, Holick MF, Ruderman NB (2006) AMPK regulation of the growth of cultured human keratinocytes. Biochem Biophys Res Commun 349:519–524

    PubMed  Article  CAS  Google Scholar 

  13. Swinnen JV, Beckers A, Brusselmans K, Organe S, Segers J, Timmermans L, Vanderhoydonc F, Deboel L, Derua R, Waelkens E, De Schrijver E, Van de Sande T, Noel A, Foufelle F, Verhoeven G (2005) Mimicry of a cellular low energy status blocks tumor cell anabolism and suppresses the malignant phenotype. Cancer Res 65:2441–2448

    PubMed  Article  CAS  Google Scholar 

  14. Rattan R, Giri S, Singh AK, Singh I (2005) 5-Aminoimidazole-4-carboxamide-1-beta-D-ribofuranoside inhibits cancer cell proliferation in vitro and in vivo via AMP-activated protein kinase. J Biol Chem 280:39582–39593

    PubMed  Article  CAS  Google Scholar 

  15. Buzzai M, Jones RG, Amaravadi RK, Lum JJ, DeBerardinis RJ, Zhao F, Viollet B, Thompson CB Systemic treatment with the antidiabetic drug metformin selectively impairs p53-deficient tumor cell growth. Cancer Res 67:6745–6752

  16. Anisimov VN, Berstein LM, Egormin PA, Piskunova TS, Popovich IG, Zabezhinski MA, Kovalenko IG, Poroshina TE, Semenchenko AV, Provinciali M, Re F, Franceschi C (2005) Effect of metformin on life span and on the development of spontaneous mammary tumors in HER-2/neu transgenic mice. Exp Gerontol 40:685–693

    PubMed  Article  CAS  Google Scholar 

  17. Carling D (2004) The AMP-activated protein kinase cascade—a unifying system for energy control. Trends Biochem Sci 29:18–24

    PubMed  Article  CAS  Google Scholar 

  18. Salt IP, Johnson G, Ashcroft SJ, Hardie DG (1998) AMP-activated protein kinase is activated by low glucose in cell lines derived from pancreatic beta cells, and may regulate insulin release. Biochem J 335(Pt 3):533–539

    PubMed  CAS  Google Scholar 

  19. Hardie DG (1999) Roles of the AMP-activated/SNF1 protein kinase family in the response to cellular stress. Biochem Soc Symp 64:13–27

    PubMed  CAS  Google Scholar 

  20. Yun H, Lee M, Kim S-S, Ha J (2005) Glucose deprivation increases mRNA stability of vascular endothelial growth factor through activation of AMP-activated protein kinase in DU145 prostate carcinoma. J Biol Chem %R 10.1074/jbc.M412994200, 280:9963–9972

    Google Scholar 

  21. Neurath KM, Keough MP, Mikkelsen T, Claffey KP AMP-dependent protein kinase alpha 2 isoform promotes hypoxia-induced VEGF expression in human glioblastoma. Glia 53:733–743

  22. Lee M, Hwang JT, Lee HJ, Jung SN, Kang I, Chi SG, Kim SS, Ha J (2003) AMP-activated protein kinase activity is critical for hypoxia-inducible factor-1 transcriptional activity and its target gene expression under hypoxic conditions in DU145 cells. J Biol Chem 278:39653–39661

    PubMed  Article  CAS  Google Scholar 

  23. Ouchi N, Shibata R, Walsh K (2005) AMP-activated protein kinase signaling stimulates VEGF expression and angiogenesis in skeletal muscle. Circ Res 96:838–846

    PubMed  Article  CAS  Google Scholar 

  24. Nagata D, Mogi M, Walsh K (2003) AMP-activated protein kinase (AMPK) signaling in endothelial cells is essential for angiogenesis in response to hypoxic stress. J Biol Chem 278:31000–31006

    PubMed  Article  CAS  Google Scholar 

  25. Agarwal A, Munoz-Najar U, Klueh U, Shih SC, Claffey KP (2004) N-acetyl-cysteine promotes angiostatin production and vascular collapse in an orthotopic model of breast cancer. Am J Pathol 164:1683–1696

    PubMed  CAS  Google Scholar 

  26. Shih SC, Mullen A, Abrams K, Mukhopadhyay D, Claffey KP (1999) Role of protein kinase C isoforms in phorbol ester-induced vascular endothelial growth factor expression in human glioblastoma cells. J Biol Chem 274:15407–15414

    PubMed  Article  CAS  Google Scholar 

  27. Woods A, Vertommen D, Neumann D, Turk R, Bayliss J, Schlattner U, Wallimann T, Carling D, Rider MH (2003) Identification of phosphorylation sites in AMP-activated protein kinase (AMPK) for upstream AMPK kinases and study of their roles by site-directed mutagenesis. J Biol Chem 278:28434–28442

    PubMed  Article  CAS  Google Scholar 

  28. Berker B, Emral R, Demirel C, Corapcioglu D, Unlu C, Kose K (2004) Increased insulin-like growth factor-I levels in women with polycystic ovary syndrome, and beneficial effects of metformin therapy. Gynecol Endocrinol 19:125–133

    PubMed  Article  CAS  Google Scholar 

  29. Wykoff CC, Beasley NJP, Watson PH, Turner KJ, Pastorek J, Sibtain A, Wilson GD, Turley H, Talks KL, Maxwell PH, Pugh CW, Ratcliffe PJ, Harris AL (2000) Hypoxia-inducible expression of tumor-associated carbonic anhydrases. Cancer Res 60:7075–7083

    PubMed  CAS  Google Scholar 

  30. Shaw RJ, Lamia KA, Vasquez D, Koo SH, Bardeesy N, Depinho RA, Montminy M, Cantley LC (2005) The kinase LKB1 mediates glucose homeostasis in liver and therapeutic effects of metformin. Science 310:1642–1646

    PubMed  Article  CAS  Google Scholar 

  31. Shen Z, Wen XF, Lan F, Shen ZZ, Shao ZM (2002) The tumor suppressor gene LKB1 is associated with prognosis in human breast carcinoma. Clin Cancer Res 8:2085–2090

    PubMed  CAS  Google Scholar 

  32. Skobe M, Hawighorst T, Jackson DG, Prevo R, Janes L, Velasco P, Riccardi L, Alitalo K, Claffey K, Detmar M (2001) Induction of tumor lymphangiogenesis by VEGF-C promotes breast cancer metastasis. Nat Med 7:192–198

    PubMed  Article  CAS  Google Scholar 

  33. Foretz M, Carling D, Guichard C, Ferre P, Foufelle F (1998) AMP-activated protein kinase inhibits the glucose-activated expression of fatty acid synthase gene in rat hepatocytes. J Biol Chem 273:14767–14771

    PubMed  Article  CAS  Google Scholar 

  34. Leclerc I, Kahn A, Doiron B (1998) The 5′-AMP-activated protein kinase inhibits the transcriptional stimulation by glucose in liver cells, acting through the glucose response complex. FEBS Lett 431:180–184

    PubMed  Article  CAS  Google Scholar 

  35. Woods A, Azzout-Marniche D, Foretz M, Stein SC, Lemarchand P, Ferre P, Foufelle F, Carling D (2000) Characterization of the role of AMP-activated protein kinase in the regulation of glucose-activated gene expression using constitutively active and dominant negative forms of the kinase. Mol Cell Biol 20:6704–6711

    PubMed  Article  CAS  Google Scholar 

  36. Xiang X, Saha AK, Wen R, Ruderman NB, Luo Z (2004) AMP-activated protein kinase activators can inhibit the growth of prostate cancer cells by multiple mechanisms. Biochem Biophys Res Commun 321:161–167

    PubMed  Article  CAS  Google Scholar 

  37. Menendez JA, Vellon L, Oza BP, Lupu R (2005) Does endogenous fatty acid metabolism allow cancer cells to sense hypoxia and mediate hypoxic vasodilatation? Characterization of a novel molecular connection between fatty acid synthase (FAS) and hypoxia-inducible factor-1alpha (HIF-1alpha)-related expression of vascular endothelial growth factor (VEGF) in cancer cells overexpressing her-2/neu oncogene. J Cell Biochem 94:857–863

    PubMed  Article  CAS  Google Scholar 

  38. Dunnwald LK, Rossing MA, Li CI (2007) Hormone receptor status, tumor characteristics, and prognosis: a prospective cohort of breast cancer patients. Breast Cancer Res 9:R6

    PubMed  Article  Google Scholar 

  39. Tarone RE, Chu KC (2002) The greater impact of menopause on ER− than ER+ breast cancer incidence: a possible explanation (United States). Cancer Causes Control 13:7–14

    PubMed  Article  Google Scholar 

Download references

Acknowledgements

The authors would like to thank Nancy Ryan for all of her technical assistance. This work was supported by NIH:NCI CA064436, the Patrick and Catherine Weldon Donaghue Foundation, and the Connecticut Breast Health Initiative, Inc.

Author information

Affiliations

  1. Center for Vascular Biology, EM028, Department of Cell Biology-MC3501, University of Connecticut Health Center, 263 Farmington Ave., Farmington, CT, 06030-3501, USA

    Kathryn N. Phoenix, Frank Vumbaca & Kevin P. Claffey

Authors
  1. Kathryn N. Phoenix
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Frank Vumbaca
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Kevin P. Claffey
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Kevin P. Claffey.

Electronic supplementary material

Below is the link to the electronic supplementary material.

ESM1 (TIF 11899 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Phoenix, K.N., Vumbaca, F. & Claffey, K.P. Therapeutic metformin/AMPK activation promotes the angiogenic phenotype in the ERα negative MDA-MB-435 breast cancer model. Breast Cancer Res Treat 113, 101–111 (2009). https://doi.org/10.1007/s10549-008-9916-5

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10549-008-9916-5

Keywords

  • AMPK
  • Angiogenesis
  • Breast carcinoma
  • MAPK
  • Metformin
  • VEGF