Research ArticleExperimental Studies

Metformin Inhibits Tumor Cell Migration via Down-regulation of MMP9 in Tamoxifen-resistant Breast Cancer Cells

SOON YOUNG JANG, AEREE KIM, JIN KYOUNG KIM, CHUNGYEL KIM, YOUL-HEE CHO, JONG-HO KIM, CHUL HWAN KIM and JI-YUN LEE
Anticancer Research August 2014, 34 (8) 4127-4134;

Abstract

Metformin is the most widely used anti-diabetic drug in the world. Recent evidence indicates that metformin could potentially inhibit tumorigenesis. In the present study, we found that metformin inhibited cell migration and invasion of phorbol 12-myristate 13-acetate-induced MCF-7 and tamoxifen-resistant MCF-7 breast cancer cells. This inhibition was correlated with the modulation of matrix metalloproteinase-9 (MMP9) via the suppression of its expression and proteolytic activity. These results indicate that metformin leads to the suppression of migration and invasion through regulation of MMP9 and it may have potential as an anticancer drug for therapy in human breast cancer, especially of chemoresistant cancer cells.

Cancer cell invasion and metastasis represent a series of events in the tumor microenvironment. One such event is the proteolytic degradation of extracellular matrix components (1, 2). Certain proteases, such as serine proteases, cysteine proteases, and matrix metalloproteinases (MMPs), contribute to invasion and metastasis. MMPs can be divided into four sub-classes: collagenase, gelatinase, stromelysin, and membrane-associated MMPs (3). Recent studies have described MMPs as critical regulators of the tumor microenvironment, as well as being involved in tumor progression, metastasis, invasion, and inflammation (4). Human MMPs, such as gelatinase-A (MMP2) and gelatinase-B (MMP9), are key enzymes that are responsible for the degradation of type-IV collagen, which is a major component of the basement membrane. These two MMPs are known to be associated with tumor migration and invasion in several types of cancers (1). Regulation of MMP9 is important for several biological processes, including angiogenesis, inflammatory response, and wound healing (5). The elevated expression of MMP9 occurs in response to inflammatory and oncogenic signals and is involved in several pathological processes such as metastasis, tumor-induced angiogenesis, lupus, and asthma (5, 6). The expression of MMP9 can be induced at the transcriptional level in response to different agents, such as growth factors, interleukins, tumor necrosis factor (TNF)-α and phorbol 12-myristate 13-acetate (PMA). Modulation of the MMP9 gene is achieved via a 2.2-kb upstream regulatory sequence containing binding sites for activator protein 1 (AP1), nuclear factor kappa-light-chain-enhancer of activated B cells (NFκB), and specificity protein 1 (SP1) (7, 8). Two AP1 binding sites (proximal and distal) have been shown to contribute to the transcriptional induction in response to several stimuli (9). However, all DNA binding sites, namely AP1, NFκB, and SP1, are required for full activation of MMP9 (10).

The oral anti-diabetic drug metformin belongs to the family of bi-guanides and is the most widely used anti-hyperglycemic drug in the world. Metformin has been shown to suppress the energy-sensitive AMP-activated protein kinase/mammalian target of rapamycin signaling pathway, which leads to reduced protein synthesis and cell proliferation. Recent retrospective epidemiological studies have shown that metformin treatment is associated with reduced risk of various types of cancer, such as prostate, breast, lung, and pancreatic cancer (11-13). In addition, several groups have reported the inhibitory effect of metformin on melanoma cell proliferation through the induction of autophagic cell death (14, 15). These results suggest that metformin might potentially be used as an anticancer drug for different types of cancer.

Tamoxifen resistance presents a serious challenge to the treatment of patients with breast cancer. In spite of an initial response to such therapies, many patients will eventually show tumor relapse and disease progression (16, 17). Previous studies have shown that acquisition of tamoxifen resistance in breast cancer cells is indicative of an increased metastatic ability, including invasion and migration.

In the present study, we investigated whether metformin exerts its effects through inhibition of MMP9 in MCF-7 cells as well as tamoxifen-resistant TamR-MCF-7 breast cancer cells.

Materials and Methods

Materials and reagents. Metformin was purchased from Sigma (St. Louis, MO, USA), and antibodies against MMP2 and MMP9 were purchased from Cell Signaling (Boston, MA, USA). Antibodies against proliferating cell nuclear antigen, p65, c-JUN, and c-FOS were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). PMA was purchased from Calbiochem (San Diego, CA, USA). MMP9 ELISA kits were purchased from R&D Systems (Abingdon, UK).

Cell lines and culture. The human breast cancer cell lines MCF-7 were obtained from America Type Culture Collection (Mannassa, VA, USA) and Tamoxifen-resistant MCF-7 were gifts from Dr. A. Kim, Korea University Guro Hospital. MCF-7 cells were grown in RPMI 1640 medium supplemented with 10% fetal bovine serum. Stable TamR-MCF-7 cells were maintained in phenol red-free RPMI medium supplemented with 10% charcoal-stripped and steroid-depleted fetal bovine serum (FBS).

Transwell invasion assays. The invasiveness of tumor cells was assessed by an invasion assay in Transwell chambers consisting of a Transwell membrane (8 μm pore size, 6.5 mm diameter) (Corning Life Science, Corning, NY, USA) coated with Matrigel (100 μg/ml, 100 μl/well). Cells were seeded onto the upper wells in the presence of different concentrations (5 mM to 100 mM) of metformin. The bottom chambers of the Transwell were filled with cell growth medium. Cells were fixed, stained, and counted under a light microscope after 24 h incubation.

In-gel zymography. MMP activities were assayed as described elsewhere (18). Briefly, 5×105 cells in a six-well plate were cultured in serum-free medium for 16 to 24 h, and the conditioned medium thus produced was separated on a sodium dodecyl sulfate –polyacrylamide electrophoresis gel (by SDS–PAGE) containing 1 mg/ml gelatin. The gel was washed with buffer I (Tris–HCl; pH 7.5) and 2.5% Triton X-100), incubated overnight in buffer II (150 mM NaCl, 5 mM CaCl2, and 50 mM Tris–HCl; pH 7.6) at 37°C, and stained with Coomassie blue. Clear bands indicated where MMPs had degraded the gelatin.

Cell migration assay. Cell migration was assessed using a wound-healing assay. Cells were seeded at 2×104 cells per 6-well plate. After scraping the cell monolayer with a sterile micropipette tip, the wells were washed with serum-free medium and incubated several times with different concentrations (5 mM to 100 mM) of metformin. The first image of each scratch was acquired at time zero. Each scratch was examined and captured at the same location, and the healed area was measured after 24 h.

Western blotting. Cell extracts were prepared using RIPA buffer (1× phosphate buffered saline (PBS), 1% NP40, 0.5% sodium deoxycholate, 0.1% SDS containing 100 μl of 10 mg/ml phenylmethanesulfonylfluoride or phenylmethylsulfonyl fluoride (PMSF), and one tablet of the complete mini protease inhibitors) after washing cells twice with 1×PBS. The protein lysates were resolved by SDS–PAGE and transferred using nitrocellulose membranes (Whatman PROTEAN® BA83, 0.2 μm). The membranes were incubated with buffer containing 0.1% Tween 20 and 5% skim milk and were then exposed to the desired primary antibody. After treatment with a suitable secondary antibody, the immunoreactive bands were visualized using the standard enhanced chemiluminescence (Pierce, Rockford, IL, USA) method.

Reverse transcription-PCR (RT-PCR). Total RNA was extracted from the treated cells, and cDNA was synthesized from 1 μg of total RNA using a High Capacity cDNA Synthesis Kit (Applied Biosystems, Foster city, CA, USA) for the RT-PCR analysis. The following PCR primers were used: MMP9 sense: 5’- TTTGACAGCGACAAGAAGTGG-3’, antisense: 5’- TCCCATC CTTGAACAAATACA-3’; MMP2 sense: 5’- CATTCCGCTTCCA GGGCACAT-3’, antisense: 5’- GCTCCTGAATGCCCTTGAT GTCA-3’; Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) sense: 5’-GCCATCGTCACCAACTGGGAC-3’, antisense: 5’-CGATTTCCCGCTCGGCCGTGG-3’. The PCR products were analyzed using agarose gel electrophoresis and visualized after treatment with ethidium bromide.

Luciferase reporter assay. MCF-7 and TamR-MCF-7 cells were transiently co-transfected with 0.5 μg MMP9-, NFκB-, and AP1-luciferase plasmids and 0.5 μg pSV-β-galactosidase reporter vector using Lipofectamine 2000 (Invitrogen, Carlsbad, CA, USA) transfection reagent. The MMP9- and NF-κB-luciferase plasmids were kindly provided by Dr. I. Shin (Hanyang University, South Korea) and the AP1-luciferase plasmid was kindly provided by Dr. S. Jang (University of Ulsan College of Medicine, South Korea). After transfection for 24 h, the cells were treated with metformin for the described period, 24 h. Cell extracts were prepared for the luciferase assays. The luciferase activity was normalized by β-galactosidase activity.

Electrophoretic mobility shift assay (EMSA). The oligonucleotides were labeled with (γ-32P) ATP and incubated with nuclear extracts for 30 min using gel shift assay kit (Promega, Madison, WI, USA). The DNA–protein complexes were resolved on non-denaturing and non-reducing 6% acrylamide gels. The probes used for EMSA were as follows: for AP1, 5’-CGC TTG ATG ACT CAG CCG GAA-3’; and for NFκB, 5’-AGT TGA GGG GAC TTT CCC GAA C-3’.

ELISA assay. Cells were cultured at 5×105 cells per well in 6-well plates. At approximately 80% confluence, the cells were stimulated by metformin (5 mM to 10 mM) in the absence and presence of 100 nM PMA. The supernatants were collected after 24 h, and the level of MMP9 in the cultured media were determined using an ELISA kit (R&D Systems, Abingdon, UK).

Statistical analysis. The results are represented as the mean±standard error (SE), and the statistical comparisons between groups were performed using one-way ANOVA followed by Student's t-test. A value of p≤0.05 was considered statistically significant.

Figure 1.
Figure 1.

Suppression of migration and invasion by metformin in MCF-7 and tamoxifen-resistant TamR-MCF7 human breast cancer cells. A: Cultures of MCF-7 (upper panel) and TamR-MCF-7 (lower panel) cells were ‘wounded’ by scratching with a pipette tip and incubated with or without 100 nM PMA and the indicated concentrations of metformin. Representative images of wound healing were taken at the time of the scratch and 24 h later. B: Invasion was determined using a Transwell assay after treatment of MCF-7 (upper panel) and TamR-MCF-7 (lower panel) cells with the indicated concentrations of metformin, with or without 100 nM PMA for 24 h. Representative photomicrographs show membrane-associated cells were stained with eosin. The data represent the average of three independent experiments±SD. Statistically significant difference (p≤0.05) from *PMA treatment only, #from the control group.

Results

Metformin suppresses migration and invasion in both MCF-7 and TamR-MCF-7 cells. Since TamR-MCF-7 cells display enhanced invasive capacity (19), we first examined the inhibitory effects of metformin on the invasive potency of MCF-7 cells and TamR-MCF-7 cells using in vitro migration and invasion assays. MCF-7 cells treated with PMA, of which the effects on cancer cell migration have been previously described (20), presented a 2- to 3-fold increase in migration and invasion. However, treatment with metformin inhibited PMA-induced migration and invasion in a somewhat dose-dependent manner (Figure 1A and B, upper panel). In addition, we confirmed the anti-invasive effects of metformin on TamR-MCF-7 cells. Metformin inhibited the migration and invasion of TamR-MCF-7 cells in a manner similar to its effects on MCF-7 cells (Figure 1A and B, lower panel). These results indicate that metformin effectively inhibits migration and invasion of MCF-7 and TamR-MCF-7 cells.

Metformin reduces expression and secretion of MMP9. It is known that MMP9 and MMP2 are critical for invasion and migration in several cancer cell lines (21). Therefore, we examined the effect of metformin on the levels of both MMP2 and MMP9 in MCF-7 and TamR-MCF-7 cells. As shown in Figure 2A, metformin reduced MMP9 expression in both PMA-treated MCF-7 cells and TamR-MCF-7 cells in a dose-dependent manner. However, the expression of MMP2 was not significantly affected by PMA nor by metformin treatment in either cell line (Figure 2A). To further test the effect of metformin on MMP activity, which is related to the invasion and metastasis of human cancer, gelatin zymography was performed in PMA-treated MCF-7 cells and TamR-MCF-7 cells. The results showed that metformin reduced MMP9 activity in both cell lines (Figure 2B). To confirm the inhibitory effects of metformin on MMP9, an ELISA assay was performed to estimate MMP9 secretion. It was shown that a 24-h treatment of metformin significantly inhibited the secretion of MMP9 from PMA-treated MCF-7 cells (Figure 2C). These results indicate that metformin inhibits cell migration and invasion through down-regulation of MMP9 in both MCF-7 and TamR-MCF7 cells.

Figure 2.
Figure 2.

The reduction of MMP9 expression and secretion by metformin in MCF-7 and tamoxifen-resistant TamR-MCF7 human breast cancer cells. A: MMP2 and MMP9 protein and mRNA levels in MCF-7 and TamR-MCF-7 cells were evaluated using western blot analysis and semi-quantitative RT-PCR. Expression levels of β-actin and GAPDH were included as internal control. The asterisk represents an unspecified band. B: MCF-7 (upper panel) and TamR-MCF-7 (lower panel) cells in serum-free medium were treated with the indicated concentrations of metformin with or without 100 nM PMA for 24 h. The cell-conditioned medium was collected after treatment and assayed using gelatin zymography to analyze the activity of MMP9. Assays were performed several times. C: MMP9 secreted by MCF-7 cells in the conditioned media was quantified using ELISA. Data are the means±SD of triplicate experiments. **Statistically significant difference from PMA treatment only (p≤0.001).

Figure 3.
Figure 3.

The inhibition of MMP9 promoter activity by metformin in MCF-7 and tamoxifen-resistant TamR-MCF-7 human breast cancer cells. The MCF-7 and Tam-MCF-7 cells were transfected with MMP9-Luc (0.5 μg) (A), NFκB-Luc (0.5 μg) (B), and AP1-Luc (0.5 μg) (C) reporter plasmids. After 24 h, cells were treated with the indicated concentrations of metformin for 24 h (for MCF-7 cells in the presence of 100 nM PMA) and harvested for luciferase assays. The luciferase activity was normalized to that of β-galactosidase. Data are the means±SD of triplicate experiments and denote a statistically significant difference at *p≤0.05 and **p≤0.001 from PMA treatment only, and at #p≤0.05 and ##p≤0.001 from the control.

Metformin inhibits transcription of MMP9. To investigate the molecular mechanisms underlying the inhibitory effects of metformin on MMP9 expression, we performed a promoter assay in MCF-7 and TamR-MCF-7 cells. We used a luciferase reporter plasmid assay containing the minimal response elements, NFκB and AP1, located in the -710 bp region upstream of the transcription start site of the human MMP9 gene. After MCF-7 cells were transfected with MMP9-Luc, the promoter activity in response to PMA, as well as to PMA with metformin, were examined. MMP9-Luc activity showed an 8-fold increase in response to PMA and 75% repression of this response on treatment with both PMA and 10 mM metformin (Figure 3A, upper panel). Additionally, metformin treatment of TamR-MCF7 cells inhibited MMP9-Luc activity in a dose-dependent manner (Figure 3A, lower panel). This suggests that metformin can inhibit MMP9 expression at the transcriptional level. To confirm this observation, luciferase reporter plasmids containing tandem repeats of the AP1- or NFκB-binding sites were used to study the effects of metformin on the promoter activity. Treatment with metformin had no effect on NFκB-Luc activity in MCF-7 cells (Figure 3B, upper panel) but significantly reduced NFκB-Luc activity in TamR-MCF-7 cells at concentrations ranging from 25 to 75 mM (Figure 3B, lower panel). However, AP1-Luc activity was reduced by metformin in a dose-dependent manner in both MCF-7 and TamR-MCF-7 cells (Figure 3C). These results indicate that the AP1 transcription factor contributed to the inhibition of MMP9 by metformin in MCF-7 and TamR-MCF7 cells. In addition, TamR-MCF-7 cells may employ another signaling pathway for the regulation of MMP9.

Metformin inhibits specific transcription factor binding to the MMP9 promoter region. To determine whether metformin leads to the repression of MMP9 transcription by inhibiting the binding of transcription factors such as AP1 or NFκB to the proximal promoter region, we performed EMSA. Nuclear extracts from MCF-7 cells were prepared and used to detect AP1 binding to the proximal AP1 sequence within the MMP9 promoter. As shown in Figure 4A, metformin significantly reduced PMA-induced AP1 DNA-binding activity in MCF-7 cells. Nuclear extracts from TamR-MCF-7 cells treated with different concentrations of metformin for 24 h were isolated and analyzed for AP1 and NFκB DNA-binding activities. Metformin reduced the NFκB as well as AP1 DNA-binding activities in a dose-dependent manner (Figure 4A). These data were consistent with the promoter analysis (Figure 3), suggesting that metformin inhibits MMP9 expression through DNA binding activity of AP1 and/or NFκB transcription factors. To identify the subunit of the AP1 transcription factor that is regulated by metformin, we examined the expression of c-FOS and c-JUN through nuclear fractionation. Our data show that metformin reduced PMA-induced c-JUN expression but had little effect on the expression of c-FOS or p65 in MCF-7 cells (Figure 4B). Furthermore, metformin markedly inhibited p65 and c-FOS expression in TamR-MCF-7 cells but had no effect on the expression of c-JUN (Figure 4B). These results suggest that the mechanism of metformin inhibition of MMP9 is based on the DNA binding activity of different transcription factors in MCF-7 (AP1 only) and TamR-MCF-7 cells (AP1 and NFκB).

Figure 4.
Figure 4.

The effects of metformin on the DNA-binding activities and expression in MCF-7 and tamoxifen-resistant TamR-MCF-7 human breast cancer cells. A: MCF-7 cells were treated with metformin for 24 h in the presence of 100 nM PMA, and nuclear extracts (5 μg) were prepared. The nuclear extracts were incubated with radiolabeled oligonucleotides containing the AP1 motif in the MMP9 promoter. Nuclear extracts (5 μg) from TamR-MCF-7 cells treated with metformin for 24 h were mixed with radioactive oligonucleotides containing the AP1 and NFκB motifs in the MMP9 promoter. Bound complexes were analyzed by EMSA. B: The nuclear fraction was extracted from MCF-7 cells treated with metformin and 100 nM PMA for 24 h, and from TamR-MCF-7 cells treated with metformin for 24 h. The nuclear levels of AP1 (c-FOS and c-JUN) and NFκB (p65) subunits were determined by western blot analysis using subunit-specific antibodies. PCNA expression was included as an internal control.

Discussion

TamR-MCF-7 cells express increased levels of epidermal growth factor receptor (EGFR), have enhanced motility, and a fibroblast-like phenotype (19, 22). The progression of tamoxifen-resistant tumors is one of the challenges encountered in endocrine therapy of breast cancer. Several studies have supported the hypothesis that tamoxifen resistance occurs via augmentation of altered signaling pathways. Specifically, EGFR, EGFR2/HER2, and insulin-like growth factor-1 receptor signaling pathways are elevated, and the activity of kinases, such as extracellular regulated kinase 1/2, p38, and protein kinase B, is also increased (23).

MMP9 is a 92-kDa type-IV collagenase that belongs to the gelatinase group of human MMPs. The expression of MMP9 is regulated by diverse growth factors, cytokines, and xenobiotics, such as PMA (24), and is related to the invasion, metastasis, and angiogenesis of various types of cancer cell lines. Therefore, developing drugs that inhibit MMP9 could be useful in human cancer therapy.

Metformin, which reduces blood glucose levels and enhances insulin sensitivity, is the most commonly prescribed oral drug in patients with type II diabetes. Metformin treatment has growth-inhibitory effects involving the activation of AMP-activated kinase and serine/threonine protein kinases, which serve as energy sensors in all eukaryotic cell types (25, 26). Recently, several studies have shown that metformin can inhibit cell proliferation in several cancer cell lines including MCF-7 cells (25).

In the present study, we observed that metformin was effective in inhibiting in vitro invasion through down-regulation of MMP9 in both MCF-7 and TamR-MCF-7 cells. Metformin suppressed the PMA-induced activity of MMP9 in MCF-7 cells, whereas the activity of MMP2 was not significantly affected. Similar data were also found for TamR-MCF7 cells. To determine the underlying mechanism, we performed several experiments including gelatin zymography, real time (RT)-PCR, western blot analysis, and luciferase reporter gene assay. Based on the above data, we found that the inhibitory effect of metformin on MMP9 activity was due to its effects on transcriptional regulation. In addition, we observed that the major target of metformin in the regulation of MMP9 was AP-1 transcription factor, whereas NFκB had little effect in MCF-7 cells. However, metformin modulated both AP1 and NFκB, key transcription factors related in the regulation of MMP9 gene expression in TamR-MCF-7 cells. Finally, we evaluated the suppression of DNA-binding activity by metformin from the MMP9 promoter in both MCF-7 and TamR-MCF-7 cells. Importantly, we found that treatment with metformin dramatically reduces DNA-binding activity of the transcription factors AP1/NFκB in both cell lines. Our results demonstrate that metformin suppression of AP1 or NFκB is an effective strategy for blocking MMP9 induction. Thus, the regulation of AP1 and NFκB, which is downstream of several signaling pathways such as Focal Adhesion Kinase, phosphoinositide 3-kinase/protein kinase B, and Mitogen-activated protein kinases, is essentially involved in the cascade of tumor invasion and metastasis (27).

In conclusion, we demonstrate, as far as we are aware for the first time, that metformin inhibits MCF-7 and TamR-MCF-7 breast cancer cell migration and invasion through MMP9 regulation. This finding brings new clues to the understanding of the differences inaction of metformin between MCF-7 and TamR-MCF-7 cells. Metformin treatment might be considered in preventing the invasion and metastasis of human malignant tumors, such as drug-resistant breast cancer.

Acknowledgements

This work was supported by grant K1133691 from Korea University, and by the Basic Science Research Program (2014R1A2A2A01003566) from the National Research Foundation of Korea (NRF) grant, funded by the Ministry of Education, Science and Technology (MEST), Republic of Korea.

Footnotes

  • Declaration of Interest

    None.

  • Received April 7, 2014.
  • Revision received June 3, 2014.
  • Accepted June 4, 2014.

References

  1. ↵
    1. Chambers AF,
    2. Matrisian LM
    : Changing views of the role of matrix metalloproteinases in metastasis. J Natl Cancer Inst 89(17): 1260-1270, 1997.
    OpenUrlAbstract/FREE Full Text
  2. ↵
    1. Woodhouse EC,
    2. Chuaqui RF,
    3. Liotta LA
    : General mechanisms of metastasis. Cancer 80(8 Suppl): 1529-1537, 1997.
    OpenUrlCrossRefPubMed
  3. ↵
    1. Yan C,
    2. Boyd DD
    : Regulation of matrix metalloproteinase gene expression. J Cell Physiol 211(1): 19-26, 2007.
    OpenUrlCrossRefPubMed
  4. ↵
    1. Kessenbrock K,
    2. Plaks V,
    3. Werb Z
    : Matrix metalloproteinases: regulators of the tumor microenvironment. Cell 141(1): 52-67, 2010.
    OpenUrlCrossRefPubMed
  5. ↵
    1. Galewska Z,
    2. et al
    : Gelatinase matrix metalloproteinase (MMP)-2 and MMP-9 of the umbilical cord blood in preeclampsia. Clin Chem Lab Med 46(4): 517-22, 2008.
    OpenUrlPubMed
  6. ↵
    1. Ram M,
    2. Sherer Y,
    3. Shoenfeld Y
    : Matrix metalloproteinase-9 and autoimmune diseases. J Clin Immunol 26(4): 299-307, 2006.
    OpenUrlCrossRefPubMed
  7. ↵
    1. Himelstein BP,
    2. et al
    : Transcriptional activation of the matrix metalloproteinase-9 gene in an H-ras and v-myc transformed rat embryo cell line. Oncogene 14(16): 1995-1998, 1997.
    OpenUrlCrossRefPubMed
  8. ↵
    1. Mohan R,
    2. et al
    : Curcuminoids inhibit the angiogenic response stimulated by fibroblast growth factor-2, including expression of matrix metalloproteinase gelatinase B J Biol Chem 275(14): 10405-10412, 2000.
    OpenUrlAbstract/FREE Full Text
  9. ↵
    1. Yan C,
    2. et al
    : Repression of 92-kDa type IV collagenase expression by MTA1 is mediated through direct interactions with the promoter via a mechanism, which is both dependent on and independent of histone deacetylation. J Biol Chem 278(4): 2309-2316, 2003.
    OpenUrlAbstract/FREE Full Text
  10. ↵
    1. Van den Steen PE,
    2. et al
    : Biochemistry and molecular biology of gelatinase B or matrix metalloproteinase-9 (MMP-9). Crit. Rev. Biochem. Mol Biol 37(6): 375-536, 2002.
    OpenUrl
  11. ↵
    1. Ben Sahra I,
    2. et al
    : Metformin in cancer therapy: a new perspective for an old antidiabetic drug? Mol Cancer Ther 9(5): 1092-1099, 2010.
    OpenUrlAbstract/FREE Full Text
    1. Pollak M
    : Metformin and other biguanides in oncology: advancing the research agenda. Cancer Prev Res (Phila) 3(9): 1060-1065, 2010.
    OpenUrlAbstract/FREE Full Text
  13. ↵
    1. Decensi A,
    2. et al
    : Metformin and cancer risk in diabetic patients: a systematic review and meta-analysis. Cancer Prev Res (Phila) 3(11): 1451-1461, 2010.
    OpenUrlAbstract/FREE Full Text
  14. ↵
    1. Tomic T,
    2. et al
    : Metformin inhibits melanoma development through autophagy and apoptosis mechanisms. Cell Death Dis 2: e199, 2011.
    OpenUrlCrossRefPubMed
  15. ↵
    1. Woodard J,
    2. Platanias LC
    : AMP-activated kinase (AMPK)-generated signals in malignant melanoma cell growth and survival. Biochem Biophys Res Commun 398(1): 135-139, 2010.
    OpenUrlCrossRefPubMed
  16. ↵
    1. Ali S,
    2. Coombes RC
    : Endocrine-responsive breast cancer and strategies for combating resistance. Nat Rev Cancer 2(2): 101-112, 2002.
    OpenUrlCrossRefPubMed
  17. ↵
    1. Osborne CK,
    2. Fuqua SA
    : Mechanisms of tamoxifen resistance. Breast Cancer Res Treat 32(1): 49-55, 1994.
    OpenUrlCrossRefPubMed
  18. ↵
    1. Han H,
    2. et al
    : CADPE inhibits PMA-stimulated gastric carcinoma cell invasion and matrix metalloproteinase-9 expression by FAK/MEK/ERK-mediated AP-1 activation. Mol Cancer Res 8(11): 1477-1488, 2010.
    OpenUrlAbstract/FREE Full Text
  19. ↵
    1. Hiscox S,
    2. et al
    : Tamoxifen resistance in MCF7 cells promotes EMT-like behaviour and involves modulation of beta-catenin phosphorylation. Int. J. Cancer 118(2): 290-301, 2006.
    OpenUrlCrossRefPubMed
  20. ↵
    1. Nomura N,
    2. et al
    : Phorbol 12-myristate 13-acetate (PMA)-induced migration of glioblastoma cells is mediated via p38MAPK/Hsp27 pathway. Biochem Pharmacol 74(5): 690-701, 2007.
    OpenUrlCrossRefPubMed
  21. ↵
    1. Hu B,
    2. et al
    : Fibulin-3 is uniquely upregulated in malignant gliomas and promotes tumor cell motility and invasion. Mol. Cancer Res 7(11): 1756-1770, 2009.
    OpenUrlAbstract/FREE Full Text
  22. ↵
    1. Hiscox S,
    2. et al
    : Elevated Src activity promotes cellular invasion and motility in tamoxifen resistant breast cancer cells. Breast Cancer Res Treat 97(3): 263-274, 2006.
    OpenUrlCrossRefPubMed
  23. ↵
    1. Farabegoli F,
    2. Papi A,
    3. Orlandi M
    : (−)-Epigallocatechin-3-gallate down-regulates EGFR, MMP-2, MMP-9 and EMMPRIN and inhibits the invasion of MCF-7 tamoxifen-resistant cells. Biosci Rep 31(2): 99-108, 2011.
    OpenUrlPubMed
  24. ↵
    1. Lee PP,
    2. et al
    : Functional significance of MMP-9 in tumor necrosis factor-induced proliferation and branching morphogenesis of mammary epithelial cells. Endocrinology 141(10): 3764-3773, 2000.
    OpenUrlCrossRefPubMed
  25. ↵
    1. Alimova IN,
    2. et al
    : Metformin inhibits breast cancer cell growth, colony formation and induces cell cycle arrest in vitro. Cell Cycle 8(6): 909-915, 2009.
    OpenUrlCrossRefPubMed
  26. ↵
    1. Cerezo M,
    2. et al
    : Metformin blocks melanoma invasion and metastasis development in AMPK/p53-dependent manner. Mol Cancer Ther 12(8): 1605-1615, 2013.
    OpenUrlAbstract/FREE Full Text
  27. ↵
    1. Sato T,
    2. et al
    : Inhibition of activator protein-1 binding activity and phosphatidylinositol 3-kinase pathway by nobiletin, a polymethoxy flavonoid, results in augmentation of tissue inhibitor of metalloproteinases-1 production and suppression of production of matrix metalloproteinases-1 and -9 in human fibrosarcoma HT-1080 cells. Cancer Res 62(4): 1025-1029, 2002.
    OpenUrlAbstract/FREE Full Text
Back to top

In this issue

Print
Download PDF
Article Alerts
Email Article
Citation Tools
Reprints and Permissions
Metformin Inhibits Tumor Cell Migration via Down-regulation of MMP9 in Tamoxifen-resistant Breast Cancer Cells
SOON YOUNG JANG, AEREE KIM, JIN KYOUNG KIM, CHUNGYEL KIM, YOUL-HEE CHO, JONG-HO KIM, CHUL HWAN KIM, JI-YUN LEE
Anticancer Research Aug 2014, 34 (8) 4127-4134;
Twitter logo Facebook logo Mendeley logo

Jump to section

Keywords