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Tumor Biology

, Volume 37, Issue 4, pp 5337–5346 | Cite as

Mechanism of metformin action in MCF-7 and MDA-MB-231 human breast cancer cells involves oxidative stress generation, DNA damage, and transforming growth factor β1 induction

  • Poliana Camila Marinello
  • Thamara Nishida Xavier da Silva
  • Carolina Panis
  • Amanda Fouto Neves
  • Kaliana Larissa Machado
  • Fernando Henrique Borges
  • Flávia Alessandra Guarnier
  • Sara Santos Bernardes
  • Júlio Cesar Madureira de-Freitas-Junior
  • José Andrés Morgado-Díaz
  • Rodrigo Cabral Luiz
  • Rubens Cecchini
  • Alessandra Lourenço Cecchini
Original Article

Abstract

The participation of oxidative stress in the mechanism of metformin action in breast cancer remains unclear. We investigated the effects of clinical (6 and 30 μM) and experimental concentrations of metformin (1000 and 5000 μM) in MCF-7 and in MDA-MB-231 cells, verifying cytotoxicity, oxidative stress, DNA damage, and intracellular pathways related to cell growth and survival after 24 h of drug exposure. Clinical concentrations of metformin decreased metabolic activity of MCF-7 cells in the MTT assay, which showed increased oxidative stress and DNA damage, although cell death and impairment in the proliferative capacity were observed only at higher concentrations. The reduction in metabolic activity and proliferation in MDA-MB-231 cells was present only at experimental concentrations after 24 h of drug exposition. Oxidative stress and DNA damage were induced in this cell line at experimental concentrations. The drug decreased cytoplasmic extracellular signal-regulated kinases 1 and 2 (ERK1/2) and AKT and increased nuclear p53 and cytoplasmic transforming growth factor β1 (TGF-β1) in both cell lines. These findings suggest that metformin reduces cell survival by increasing reactive oxygen species, which induce DNA damage and apoptosis. A relationship between the increase in TGF-β1 and p53 levels and the decrease in ERK1/2 and AKT was also observed. These findings suggest the mechanism of action of metformin in both breast cancer cell lineages, whereas cell line specific undergoes redox changes in the cells in which proliferation and survival signaling are modified. Taken together, these results highlight the potential clinical utility of metformin as an adjuvant during the treatment of luminal and triple-negative breast cancer.

Keywords

Metformin MCF-7 MDA-MB-231 Oxidative stress Breast cancer 

Abbreviations

ERK1/2

Extracellular signal-regulated kinases 1 and 2

AKT

Protein kinase B

TGF-β1

Transforming growth factor β1

AMPK

Adenosine-5′-monophosphate-activated protein kinase

mTOR

Mammalian target of rapamycin

HER-2

Human epidermal growth factor receptor 2

TNBC

Triple-negative breast cancer

MTT

2-(3,5-Diphenyltetrazol-2-ium-2-yl)-4,5-dimethyl-1,3-thiazole bromide

PBS

Phosphate-buffered saline

EB

Ethidium bromide

AO

Acridine orange

MDA

Malondialdehyde

8-Oh-dG

8-Hydroxy-2-deoxyguanosine

ANOVA

Analysis of variance

SOD

Superoxide dismutase

OS

Oxidative stress

ROS

Reactive oxygen species

Notes

Acknowledgments

The authors are grateful to J.A. Vargas and P.S.R. Dionízio-Filho, from the Department of General Pathology of the State University of Londrina, for their excellent technical assistance.

Compliance with ethical standards

Conflicts of interest

None

Supplementary material

13277_2015_4395_Fig7_ESM.gif (341 kb)
Supplementary Figure 1

Metformin increases nuclear p53 levels and cytoplasmic TGF-β1 levels and reduces cytoplasmic ERK1/2 and AKT in human breast cancer cells. Immunocytochemistry analysis of MCF-7 cells (a) and MDA-MB-231 cells (b) exposed to different metformin concentrations (6, 30, 1000, and 5000 μM) for 24 h. Illustrative panel showing a picture selected for each experimental condition. For the metformin 5000 μM, where the effects were more pronounced, a picture focusing on a single cell was selected (GIF 340 kb).

13277_2015_4395_MOESM1_ESM.tif (3.9 mb)
High Resolution Image (TIF 3950 kb).

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Copyright information

© International Society of Oncology and BioMarkers (ISOBM) 2015

Authors and Affiliations

  • Poliana Camila Marinello
    • 1
  • Thamara Nishida Xavier da Silva
    • 2
  • Carolina Panis
    • 3
  • Amanda Fouto Neves
    • 1
  • Kaliana Larissa Machado
    • 1
  • Fernando Henrique Borges
    • 3
  • Flávia Alessandra Guarnier
    • 2
  • Sara Santos Bernardes
    • 1
  • Júlio Cesar Madureira de-Freitas-Junior
    • 4
  • José Andrés Morgado-Díaz
    • 4
  • Rodrigo Cabral Luiz
    • 1
  • Rubens Cecchini
    • 3
  • Alessandra Lourenço Cecchini
    • 1
  1. 1.Laboratory of Molecular PathologyState University of LondrinaLondrinaBrazil
  2. 2.Laboratory of Pathophysiology and Muscle AdaptationState University of LondrinaLondrinaBrazil
  3. 3.Laboratory of Pathophysiology and Free RadicalsState University of LondrinaLondrinaBrazil
  4. 4.Brazilian National Cancer InstituteINCARio de JaneiroBrazil

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