Hormones and Cancer

, Volume 5, Issue 6, pp 374–389 | Cite as

Metformin-Induced Killing of Triple-Negative Breast Cancer Cells Is Mediated by Reduction in Fatty Acid Synthase via miRNA-193b

  • Reema S. Wahdan-Alaswad
  • Dawn R. Cochrane
  • Nicole S. Spoelstra
  • Erin N. Howe
  • Susan M. Edgerton
  • Steven M. Anderson
  • Ann D. Thor
  • Jennifer K. RicherEmail author
Original Paper


The anti-diabetic drug metformin (1,1-dimethylbiguanide hydrochloride) reduces both the incidence and mortality of several types of cancer. Metformin has been shown to selectively kill cancer stem cells, and triple-negative breast cancer (TNBC) cell lines are more sensitive to the effects of metformin as compared to luminal breast cancer. However, the mechanism underlying the enhanced susceptibility of TNBC to metformin has not been elucidated. Expression profiling of metformin-treated TNBC lines revealed fatty acid synthase (FASN) as one of the genes most significantly downregulated following 24 h of treatment, and a decrease in FASN protein was also observed. Since FASN is critical for de novo fatty acid synthesis and is important for the survival of TNBC, we hypothesized that FASN downregulation facilitates metformin-induced apoptosis. Profiling studies also exposed a rapid metformin-induced increase in miR-193 family members, and miR-193b directly targets the FASN 3′UTR. Addition of exogenous miR-193b mimic to untreated TNBC cells decreased FASN protein expression and increased apoptosis of TNBC cells, while spontaneously immortalized, non-transformed breast epithelial cells remained unaffected. Conversely, antagonizing miR-193 activity impaired the ability of metformin to decrease FASN and cause cell death. Further, the metformin-stimulated increase in miR-193 resulted in reduced mammosphere formation by TNBC lines. These studies provide mechanistic insight into metformin-induced killing of TNBC.


Metformin Metformin Treatment TNBC Cell Mammosphere Formation TNBC Cell Line 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.



This work was covered in part by the S. G. Komen Foundation for the Cure Grant K100575 to ADT, SMA, JKR, NIH P01 PAR-10-245 to SMA, and the AMC Women’s Cancer Fund/Salah Foundation (JKR). We also acknowledge the following University of Colorado Cancer Center Shared Resource facilities supported by NIH/NCI P30CA046934: the DNA Sequencing and Analysis Shared Resource, the Microarray Shared Resource, the Flow Cytometry Shared Resource, and the Protein Production, Monoclonal antibody and Tissue Culture Shared Resource.

Conflicts of Interest

No potential conflicts of interest were disclosed.

Supplementary material

12672_2014_188_MOESM1_ESM.pdf (1.6 mb)
ESM 1 (PDF 1.57 mb)


  1. 1.
    Warburg O (1956) On respiratory impairment in cancer cells. Science 124(3215):269–270PubMedGoogle Scholar
  2. 2.
    Hilvo M et al (2011) Novel theranostic opportunities offered by characterization of altered membrane lipid metabolism in breast cancer progression. Cancer Res 71(9):3236–3245PubMedCrossRefGoogle Scholar
  3. 3.
    Medes G, Thomas A, Weinhouse S (1953) Metabolism of neoplastic tissue. IV. A study of lipid synthesis in neoplastic tissue slices in vitro. Cancer Res 13(1):27–29PubMedGoogle Scholar
  4. 4.
    Ookhtens M et al (1984) Liver and adipose tissue contributions to newly formed fatty acids in an ascites tumor. Am J Physiol 247(1 Pt 2):R146–R153PubMedGoogle Scholar
  5. 5.
    Sabine JR, Abraham S, Chaikoff IL (1967) Control of lipid metabolism in hepatomas: insensitivity of rate of fatty acid and cholesterol synthesis by mouse hepatoma BW7756 to fasting and to feedback control. Cancer Res 27(4):793–799PubMedGoogle Scholar
  6. 6.
    Menendez JA, Lupu R (2007) Fatty acid synthase and the lipogenic phenotype in cancer pathogenesis. Nat Rev Cancer 7(10):763–777PubMedCrossRefGoogle Scholar
  7. 7.
    Chajes V et al (2006) Acetyl-CoA carboxylase alpha is essential to breast cancer cell survival. Cancer Res 66(10):5287–5294PubMedCrossRefGoogle Scholar
  8. 8.
    Alo PL et al (1996) Expression of fatty acid synthase (FAS) as a predictor of recurrence in stage I breast carcinoma patients. Cancer 77(3):474–482PubMedCrossRefGoogle Scholar
  9. 9.
    Menendez JA, Lupu R (2006) Oncogenic properties of the endogenous fatty acid metabolism: molecular pathology of fatty acid synthase in cancer cells. Curr Opin Clin Nutr Metab Care 9(4):346–357PubMedCrossRefGoogle Scholar
  10. 10.
    Menendez JA, Lupu R, Colomer R (2004) Inhibition of tumor-associated fatty acid synthase hyperactivity induces synergistic chemosensitization of HER-2/neu-overexpressing human breast cancer cells to docetaxel (taxotere). Breast Cancer Res Treat 84(2):183–195PubMedCrossRefGoogle Scholar
  11. 11.
    Swinnen JV, Brusselmans K, Verhoeven G (2006) Increased lipogenesis in cancer cells: new players, novel targets. Curr Opin Clin Nutr Metab Care 9(4):358–365PubMedCrossRefGoogle Scholar
  12. 12.
    Turrado C et al (2012) New synthetic inhibitors of fatty acid synthase with anticancer activity. J Med Chem 55(11):5013–5023PubMedCrossRefGoogle Scholar
  13. 13.
    Knowles LM et al (2008) Inhibition of fatty-acid synthase induces caspase-8-mediated tumor cell apoptosis by up-regulating DDIT4. J Biol Chem 283(46):31378–31384PubMedCentralPubMedCrossRefGoogle Scholar
  14. 14.
    Pizer ES et al (1996) Fatty acid synthase (FAS): a target for cytotoxic antimetabolites in HL60 promyelocytic leukemia cells. Cancer Res 56(4):745–751PubMedGoogle Scholar
  15. 15.
    Samudio I et al (2010) Pharmacologic inhibition of fatty acid oxidation sensitizes human leukemia cells to apoptosis induction. J Clin Invest 120(1):142–156PubMedCentralPubMedCrossRefGoogle Scholar
  16. 16.
    Chuang HY, Chang YF, Hwang JJ (2011) Antitumor effect of orlistat, a fatty acid synthase inhibitor, is via activation of caspase-3 on human colorectal carcinoma-bearing animal. Biomed Pharmacother 65(4):286–292PubMedCrossRefGoogle Scholar
  17. 17.
    Kridel SJ et al (2004) Orlistat is a novel inhibitor of fatty acid synthase with antitumor activity. Cancer Res 64(6):2070–2075PubMedCrossRefGoogle Scholar
  18. 18.
    Deepa PR et al (2012) Therapeutic and toxicologic evaluation of anti-lipogenic agents in cancer cells compared with non-neoplastic cells. Basic Clin Pharmacol Toxicol 110(6):494–503PubMedCrossRefGoogle Scholar
  19. 19.
    McGowan MM et al (2013) A proof of principle clinical trial to determine whether conjugated linoleic acid modulates the lipogenic pathway in human breast cancer tissue. Breast Cancer Res Treat 138(1):175–183PubMedCentralPubMedCrossRefGoogle Scholar
  20. 20.
    Puig T et al (2011) A novel inhibitor of fatty acid synthase shows activity against HER2 + breast cancer xenografts and is active in anti-HER2 drug-resistant cell lines. Breast Cancer Res 13(6):R131PubMedCentralPubMedCrossRefGoogle Scholar
  21. 21.
    Puig T et al (2009) Novel inhibitors of fatty acid synthase with anticancer activity. Clin Cancer Res 15(24):7608–7615PubMedCrossRefGoogle Scholar
  22. 22.
    Romero IL et al (2012) Relationship of type II diabetes and metformin use to ovarian cancer progression, survival, and chemosensitivity. Obstet Gynecol 119(1):61–67PubMedCentralPubMedCrossRefGoogle Scholar
  23. 23.
    Gandini S et al (2013) Metformin and breast cancer risk. J Clin Oncol 31(7):973–974PubMedCrossRefGoogle Scholar
  24. 24.
    Dowling RJ et al (2012) Metformin in cancer: translational challenges. J Mol Endocrinol 48(3):R31–R43PubMedCrossRefGoogle Scholar
  25. 25.
    Goodwin PJ, Ligibel JA, Stambolic V (2009) Metformin in breast cancer: time for action. J Clin Oncol 27(20):3271–3273PubMedCrossRefGoogle Scholar
  26. 26.
    Niraula S et al (2012) Metformin in early breast cancer: a prospective window of opportunity neoadjuvant study. Breast Cancer Res Treat 135(3):821–830PubMedCrossRefGoogle Scholar
  27. 27.
    Algire C et al (2010) Metformin blocks the stimulative effect of a high-energy diet on colon carcinoma growth in vivo and is associated with reduced expression of fatty acid synthase. Endocr Relat Cancer 17(2):351–360PubMedCrossRefGoogle Scholar
  28. 28.
    Vazquez Martin A et al (2013) The mitochondrial H(+)-ATP synthase and the lipogenic switch: new core components of metabolic reprogramming in induced pluripotent stem (iPS) cells. Cell Cycle 12(2):207–218PubMedCentralPubMedCrossRefGoogle Scholar
  29. 29.
    Vazquez‐Martin A et al (2012) Metformin limits the tumourigenicity of iPS cells without affecting their pluripotency. Sci Rep 2:964PubMedCentralPubMedGoogle Scholar
  30. 30.
    Alimova IN et al (2009) Metformin inhibits breast cancer cell growth, colony formation and induces cell cycle arrest in vitro. Cell Cycle 8((6):909–915CrossRefGoogle Scholar
  31. 31.
    Giles ED et al (2012) Obesity and overfeeding affecting both tumor and systemic metabolism activates the progesterone receptor to contribute to postmenopausal breast cancer. Cancer Res 72(24):6490–6501PubMedCentralPubMedCrossRefGoogle Scholar
  32. 32.
    Liu B et al (2009) Metformin induces unique biological and molecular responses in triple negative breast cancer cells. Cell Cycle 8(13):2031–2040PubMedCrossRefGoogle Scholar
  33. 33.
    Wahdan‐Alaswad R et al (2013) Glucose promotes breast cancer aggression and reduces metformin efficacy. Cell Cycle 12(24):3759–3769PubMedCentralPubMedCrossRefGoogle Scholar
  34. 34.
    Deng XS et al (2012) Metformin targets Stat3 to inhibit cell growth and induce apoptosis in triple-negative breast cancers. Cell Cycle 11(2):367–376PubMedCrossRefGoogle Scholar
  35. 35.
    Hirsch HA et al (2009) Metformin selectively targets cancer stem cells, and acts together with chemotherapy to block tumor growth and prolong remission. Cancer Res 69(19):7507–7511PubMedCentralPubMedCrossRefGoogle Scholar
  36. 36.
    Zakikhani M et al (2006) Metformin is an AMP kinase-dependent growth inhibitor for breast cancer cells. Cancer Res 66(21):10269–10273PubMedCrossRefGoogle Scholar
  37. 37.
    Creighton CJ et al (2009) Residual breast cancers after conventional therapy display mesenchymal as well as tumor-initiating features. Proc Natl Acad Sci U S A 106(33):13820–13825PubMedCentralPubMedCrossRefGoogle Scholar
  38. 38.
    Honeth G et al (2008) The CD44+/CD24− phenotype is enriched in basal-like breast tumors. Breast Cancer Res 10(3):R53PubMedCentralPubMedCrossRefGoogle Scholar
  39. 39.
    Siddiqui RA et al (2005) Anticancer properties of propofol-docosahexaenoate and propofol-eicosapentaenoate on breast cancer cells. Breast Cancer Res 7(5):R645–R654PubMedCentralPubMedCrossRefGoogle Scholar
  40. 40.
    Sanlioglu AD et al (2005) Surface TRAIL decoy receptor-4 expression is correlated with trail resistance in MCF7 breast cancer cells. BMC Cancer 5:54PubMedCentralPubMedCrossRefGoogle Scholar
  41. 41.
    Dontu G et al (2003) In vitro propagation and transcriptional profiling of human mammary stem/progenitor cells. Genes Dev 17(10):1253–1270PubMedCentralPubMedCrossRefGoogle Scholar
  42. 42.
    Ginestier C et al (2007) ALDH1 is a marker of normal and malignant human mammary stem cells and a predictor of poor clinical outcome. Cell Stem Cell 1(5):555–567PubMedCentralPubMedCrossRefGoogle Scholar
  43. 43.
    Zhuang Y, Miskimins WK (2008) Cell cycle arrest in metformin treated breast cancer cells involves activation of AMPK, downregulation of cyclin D1, and requires p27Kip1 or p21Cip1. J Mol Signal 3:18PubMedCentralPubMedCrossRefGoogle Scholar
  44. 44.
    Cochrane DR et al (2010) MicroRNAs link estrogen receptor alpha status and dicer levels in breast cancer. Horm Cancer 1(6):306–319PubMedCrossRefGoogle Scholar
  45. 45.
    Pandey PR et al (2011) Resveratrol suppresses growth of cancer stem-like cells by inhibiting fatty acid synthase. Breast Cancer Res Treat 130(2):387–398PubMedCentralPubMedCrossRefGoogle Scholar
  46. 46.
    Funabashi H et al (1989) Binding site of cerulenin in fatty acid synthetase. J Biochem 105(5):751–755PubMedGoogle Scholar
  47. 47.
    Pizer ES et al (1996) Inhibition of fatty acid synthesis induces programmed cell death in human breast cancer cells. Cancer Res 56(12):2745–2747PubMedGoogle Scholar
  48. 48.
    Chlebowski RT (2012) Obesity and breast cancer outcome: adding to the evidence. J Clin Oncol 30(2):126–128PubMedCrossRefGoogle Scholar
  49. 49.
    Chlebowski RT et al (2012) Diabetes, metformin, and breast cancer in postmenopausal women. J Clin Oncol 30(23):2844–2852PubMedCentralPubMedCrossRefGoogle Scholar
  50. 50.
    Knowler WC et al (2002) Reduction in the incidence of type 2 diabetes with lifestyle intervention or metformin. N Engl J Med 346(6):393–403PubMedCrossRefGoogle Scholar
  51. 51.
    Jiralerspong S et al (2009) Metformin and pathologic complete responses to neoadjuvant chemotherapy in diabetic patients with breast cancer. J Clin Oncol 27(20):3297–3302PubMedCentralPubMedCrossRefGoogle Scholar
  52. 52.
    Knobloch M et al (2013) Metabolic control of adult neural stem cell activity by Fasn-dependent lipogenesis. Nature 493(7431):226–230PubMedCentralPubMedCrossRefGoogle Scholar
  53. 53.
    Wang X et al (2013) PPARgamma maintains ERBB2-positive breast cancer stem cells. Oncogene 32(49):5512–5521PubMedCentralPubMedCrossRefGoogle Scholar
  54. 54.
    Milgraum LZ et al (1997) Enzymes of the fatty acid synthesis pathway are highly expressed in situ breast carcinoma. Clin Cancer Res 3(11):2115–2120PubMedGoogle Scholar
  55. 55.
    Esslimani Sahla M et al (2007) Increased expression of fatty acid synthase and progesterone receptor in early steps of human mammary carcinogenesis. Int J Cancer 120(2):224–229PubMedCrossRefGoogle Scholar
  56. 56.
    Chalbos D et al (1992) Progestin‐induced fatty acid synthetase in human mammary tumors: from molecular to clinical studies. J Steroid Biochem Mol Biol 43(1‐3):223–228PubMedCrossRefGoogle Scholar
  57. 57.
    Joyeux C, Chalbos D, Rochefort H (1990) Effects of progestins and menstrual cycle on fatty acid synthetase and progesterone receptor in human mammary glands. J Clin Endocrinol Metab 70(5):1438–1444PubMedCrossRefGoogle Scholar
  58. 58.
    Chalbos D et al (1990) Expression of the progestin‐induced fatty acid synthetase in benign mastopathies and breast cancer as measured by RNA in situ hybridization. J Natl Cancer Inst 82(7):602–606PubMedCrossRefGoogle Scholar
  59. 59.
    Chalbos D et al (1990) Regulation of fatty acid synthetase by progesterone in normal and tumoral human mammary glands. Rev Esp Fisiol 46(1):43–46PubMedGoogle Scholar
  60. 60.
    Chambon M et al (1989) Progestins and androgens stimulate lipid accumulation in T47D breast cancer cells via their own receptors. J Steroid Biochem 33(5):915–922PubMedCrossRefGoogle Scholar
  61. 61.
    Joyeux C, Rochefort H, Chalbos D (1989) Progestin increases gene transcription and messenger ribonucleic acid stability of fatty acid synthetase in breast cancer cells. Mol Endocrinol 3(4):681–686PubMedCrossRefGoogle Scholar
  62. 62.
    Chalbos D et al (1987) Fatty acid synthetase and its mRNA are induced by progestins in breast cancer cells. J Biol Chem 262(21):9923–9926PubMedGoogle Scholar
  63. 63.
    Martel PM et al (2006) S14 protein in breast cancer cells: direct evidence of regulation by SREBP‐1c, superinduction with progestin, and effects on cell growth. Exp Cell Res 312(3):278–288PubMedGoogle Scholar
  64. 64.
    Menendez JA, Lupu R, Colomer R (2005) Obesity, fatty acid synthase, and cancer: serendipity or forgotten causal linkage? Mol Genet Metab 84(3):293–295PubMedCrossRefGoogle Scholar
  65. 65.
    Menendez JA et al (2004) Novel signaling molecules implicated in tumor‐associated fatty acid synthase‐dependent breast cancer cell proliferation and survival: role of exogenous dietary fatty acids, p53‐p21WAF1/CIP1, ERK1/2 MAPK, p27KIP1, BRCA1, and NF‐kappaB. Int J Oncol 24(3):591–608PubMedGoogle Scholar
  66. 66.
    Seguin F et al (2012) The fatty acid synthase inhibitor orlistat reduces experimental metastases and angiogenesis in B16‐F10 melanomas. Br J Cancer 107(6):977–987PubMedCentralPubMedCrossRefGoogle Scholar
  67. 67.
    Zaytseva YY et al (2012) Inhibition of fatty acid synthase attenuates CD44‐associated signaling and reduces metastasis in colorectal cancer. Cancer Res 72(6):1504–1517PubMedCentralPubMedCrossRefGoogle Scholar
  68. 68.
    Fryer LG, Parbu‐Patel A, Carling D (2002) The Anti‐diabetic drugs rosiglitazone and metformin stimulate AMP‐activated protein kinase through distinct signaling pathways. J Biol Chem 277(28):25226–25232PubMedCrossRefGoogle Scholar
  69. 69.
    Musi N et al (2002) Metformin increases AMP‐activated protein kinase activity in skeletal muscle of subjects with type 2 diabetes. Diabetes 51(7):2074–2081PubMedCrossRefGoogle Scholar
  70. 70.
    Zhou G et al (2001) Role of AMP‐activated protein kinase in mechanism of metformin action. J Clin Invest 108(8):1167–1174PubMedCentralPubMedCrossRefGoogle Scholar
  71. 71.
    Dowling RJ et al (2007) Metformin inhibits mammalian target of rapamycin‐dependent translation initiation in breast cancer cells. Cancer Res 67(22):10804–10812PubMedCrossRefGoogle Scholar
  72. 72.
    Guo D et al (2009) The AMPK agonist AICAR inhibits the growth of EGFRvIII‐expressing glioblastomas by inhibiting lipogenesis. Proc Natl Acad Sci U S A 106(31):12932–12937PubMedCentralPubMedCrossRefGoogle Scholar
  73. 73.
    Zhao X et al (2012) Regulation of lipogenesis by cyclin‐dependent kinase 8‐mediated control of SREBP‐1. J Clin Invest 122(7):2417–2427PubMedCentralPubMedCrossRefGoogle Scholar
  74. 74.
    Lee KH et al (2011) Targeting energy metabolic and oncogenic signaling pathways in triple‐negative breast cancer by a novel adenosine monophosphate‐activated protein kinase (AMPK) activator. J Biol Chem 286(45):39247–39258PubMedCentralPubMedCrossRefGoogle Scholar
  75. 75.
    Bao B et al (2012) Metformin inhibits cell proliferation, migration and invasion by attenuating CSC function mediated by deregulating miRNAs in pancreatic cancer cells. Cancer Prev Res (Phila) 5(3):355–364CrossRefGoogle Scholar
  76. 76.
    Blandino G et al (2012) Metformin elicits anticancer effects through the sequential modulation of DICER and c‐MYC. Nat Commun 3:865PubMedCrossRefGoogle Scholar
  77. 77.
    Oliveras‐Ferraros C et al (2011) Micro(mi)RNA expression profile of breast cancer epithelial cells treated with the anti‐diabetic drug metformin: induction of the tumor suppressor miRNA let‐7a and suppression of the TGFbeta‐induced oncomiR miRNA‐181a. Cell Cycle 10(7):1144–1151PubMedCrossRefGoogle Scholar
  78. 78.
    Loftus TM et al (2000) Reduced food intake and body weight in mice treated with fatty acid synthase inhibitors. Science 288(5475):2379–2381PubMedCrossRefGoogle Scholar
  79. 79.
    Mao JH et al (2012) microRNA‐195 suppresses osteosarcoma cell invasion and migration in vitro by targeting FASN. Oncol Lett 4(5):1125–1129PubMedCentralPubMedGoogle Scholar
  80. 80.
    Park JH et al (2011) Murine hepatic miRNAs expression and regulation of gene expression in diet‐induced obese mice. Mol Cells 31(1):33–38PubMedCentralPubMedCrossRefGoogle Scholar
  81. 81.
    Shirasaki T et al (2013) MicroRNA‐27a regulates lipid metabolism and inhibits hepatitis C virus replication in human hepatoma cells. J Virol 87(9):5270–5286PubMedCentralPubMedCrossRefGoogle Scholar
  82. 82.
    Zhong D et al (2013) MicroRNA‐613 represses lipogenesis in HepG2 cells by downregulating LXRalpha. Lipids Health Dis 12:32PubMedCentralPubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  • Reema S. Wahdan-Alaswad
    • 1
  • Dawn R. Cochrane
    • 1
  • Nicole S. Spoelstra
    • 1
  • Erin N. Howe
    • 1
  • Susan M. Edgerton
    • 1
  • Steven M. Anderson
    • 1
  • Ann D. Thor
    • 1
  • Jennifer K. Richer
    • 1
    Email author
  1. 1.Department of PathologyUniversity of Colorado Anschutz Medical CampusAuroraUSA

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