Silibinin downregulates MMP2 expression via Jak2/STAT3 pathway and inhibits the migration and invasive potential in MDA-MB-231 cells

Byun,Hyo  Joo; Darvin,Pramod; Kang,Dong  Young; Sp,Nipin; Joung,Youn  Hee; Park,Jong  Hwan; Kim,Sun  Jin; Yang,Young  Mok

doi:10.3892/or.2017.5588

Silibinin downregulates MMP2 expression via Jak2/STAT3 pathway and inhibits the migration and invasive potential in MDA-MB-231 cells

Authors:
- Hyo Joo Byun
- Pramod Darvin
- Dong Young Kang
- Nipin Sp
- Youn Hee Joung
- Jong Hwan Park
- Sun Jin Kim
- Young Mok Yang
View Affiliations

Affiliations: Department of Pathology, School of Medicine, Institute of Biomedical Science and Technology, Konkuk University, Chungju 27478, Republic of Korea, Trinity Court, Cardiff CF24 0AA, Wales, UK
Published online on: April 19, 2017 https://doi.org/10.3892/or.2017.5588
Pages: 3270-3278

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )

Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )

Cited By (CrossRef): 0 citations Loading Articles...

Abstract

Worldwide, breast cancer (BCa) is the most common cancer in women. Among its subtypes, triple-negative breast cancer (TNBC) is an aggressive form associated with diminished survival. TNBCs are characterized by their absence, or minimal expression, of the estrogen and progesterone receptors, as well as the human epidermal growth factor receptor 2 (i.e. ER-/-, PR-/-, Her2-/Low). Consequently, treatment for this subtype of BCa remains problematic. Silibinin, a derivative of the flavonoid silymarin, is reported to have anticancer activities against hepatic and non-small cell lung cancers. We hypothesized that silibinin might inhibit cell-extracellular matrix interactions via the regulation, expression, and activation of STAT3 in TNBCs, which could directly inhibit metastasis in silibinin-treated BCa cells. Using proliferation assays, we found that exposure to silibinin at a concentration of 200 µM inhibited the proliferation of breast cancer (BCa) cells; this concentration also inhibited phosphorylation of STAT3 and its principal upstream kinase, Jak2. Furthermore, we found that silibinin inhibited the nuclear translocation of STAT3, as well as its binding to the MMP2 gene promoter. The ability of silibinin to inhibit metastasis was further studied using an in vitro invasion assay. The results confirm the role of STAT3 as a critical mediator in the invasive potential of BCa cells, and STAT3 knock-down resulted in inhibition of invasion. The invasion ability of silibinin-treated BCa cells was studied in detail with the expression of MMP2. Prevention of STAT3 activation also resulted in the inhibition of MMP2 expression. Use of a small interfering RNA to knock down STAT3 (siSTAT3) allowed us to confirm the role of STAT3 in regulating MMP2 expression, as well as the mechanism of action of silibinin in inhibiting MMP2. Taken together, we found that silibinin inhibits the Jak2/STAT3/MMP2 signaling pathway, and inhibits the proliferation, migration, and invasion of triple-negative BCa cells.

View Figures

View References

1	Cleator S, Heller W and Coombes RC: Triple-negative breast cancer: Therapeutic options. Lancet Oncol. 8:235–244. 2007. View Article : Google Scholar : PubMed/NCBI
2	Kuo WH, Chang YY, Lai LC, Tsai MH, Hsiao CK, Chang KJ and Chuang EY: Molecular characteristics and metastasis predictor genes of triple-negative breast cancer: A clinical study of triple-negative breast carcinomas. PLoS One. 7:e458312012. View Article : Google Scholar : PubMed/NCBI
3	Chen JQ and Russo J: ERalpha-negative and triple negative breast cancer: Molecular features and potential therapeutic approaches. Biochim Biophys Acta. 1796:162–175. 2009.PubMed/NCBI
4	Garcia R, Bowman TL, Niu G, Yu H, Minton S, Muro-Cacho CA, Cox CE, Falcone R, Fairclough R, Parsons S, et al: Constitutive activation of Stat3 by the Src and JAK tyrosine kinases participates in growth regulation of human breast carcinoma cells. Oncogene. 20:2499–2513. 2001. View Article : Google Scholar : PubMed/NCBI
5	Garcia R, Yu CL, Hudnall A, Catlett R, Nelson KL, Smithgall T, Fujita DJ, Ethier SP and Jove R: Constitutive activation of Stat3 in fibroblasts transformed by diverse oncoproteins and in breast carcinoma cells. Cell Growth Differ. 8:1267–1276. 1997.PubMed/NCBI
6	Siveen KS, Sikka S, Surana R, Dai X, Zhang J, Kumar AP, Tan BK, Sethi G and Bishayee A: Targeting the STAT3 signaling pathway in cancer: Role of synthetic and natural inhibitors. Biochim Biophys Acta. 1845:136–154. 2014.PubMed/NCBI
7	Imada K and Leonard WJ: The Jak-STAT pathway. Mol Immunol. 37:1–11. 2000. View Article : Google Scholar : PubMed/NCBI
8	Darnell JE Jr: STATs and gene regulation. Science. 277:1630–1635. 1997. View Article : Google Scholar : PubMed/NCBI
9	Thomas SJ, Snowden JA, Zeidler MP and Danson SJ: The role of JAK/STAT signalling in the pathogenesis, prognosis and treatment of solid tumours. Br J Cancer. 113:365–371. 2015. View Article : Google Scholar : PubMed/NCBI
10	Bowman T, Garcia R, Turkson J and Jove R: STATs in oncogenesis. Oncogene. 19:2474–2488. 2000. View Article : Google Scholar : PubMed/NCBI
11	Rundhaug JE: Matrix metalloproteinases, angiogenesis, and cancer: commentary re: A. C. Lockhart et al., Reduction of wound angiogenesis in patients treated with BMS-275291, a broad spectrum matrix metalloproteinase inhibitor. Clin. Cancer Res. 9:00-00, 2003. Clin Cancer Res 9. 551–554. 2003.
12	Verma RP and Hansch C: Matrix metalloproteinases (MMPs): Chemical-biological functions and (Q)SARs. Bioorg Med Chem. 15:2223–2268. 2007. View Article : Google Scholar : PubMed/NCBI
13	Visse R and Nagase H: Matrix metalloproteinases and tissue inhibitors of metalloproteinases: Structure, function, and biochemistry. Circ Res. 92:827–839. 2003. View Article : Google Scholar : PubMed/NCBI
14	Brooks PC, Strömblad S, Sanders LC, von Schalscha TL, Aimes RT, Stetler-Stevenson WG, Quigley JP and Cheresh DA: Localization of matrix metalloproteinase MMP-2 to the surface of invasive cells by interaction with integrin αvβ3. Cell. 85:683–693. 1996. View Article : Google Scholar : PubMed/NCBI
15	Rojiani MV, Alidina J, Esposito N and Rojiani AM: Expression of MMP-2 correlates with increased angiogenesis in CNS metastasis of lung carcinoma. Int J Clin Exp Pathol. 3:775–781. 2010.PubMed/NCBI
16	Zheng H, Takahashi H, Murai Y, Cui Z, Nomoto K, Niwa H, Tsuneyama K and Takano Y: Expressions of MMP-2, MMP-9 and VEGF are closely linked to growth, invasion, metastasis and angiogenesis of gastric carcinoma. Anticancer Res. 26A:3579–3583. 2006.
17	Xie TX, Huang FJ, Aldape KD, Kang SH, Liu M, Gershenwald JE, Xie K, Sawaya R and Huang S: Activation of stat3 in human melanoma promotes brain metastasis. Cancer Res. 66:3188–3196. 2006. View Article : Google Scholar : PubMed/NCBI
18	Xie TX, Wei D, Liu M, Gao AC, Ali-Osman F, Sawaya R and Huang S: Stat3 activation regulates the expression of matrix metalloproteinase-2 and tumor invasion and metastasis. Oncogene. 23:3550–3560. 2004. View Article : Google Scholar : PubMed/NCBI
19	Lee JW, Kwak HJ, Lee JJ, Kim YN, Lee JW, Park MJ, Jung SE, Hong SI, Lee JH and Lee JS: HSP27 regulates cell adhesion and invasion via modulation of focal adhesion kinase and MMP-2 expression. Eur J Cell Biol. 87:377–387. 2008. View Article : Google Scholar : PubMed/NCBI
20	Valenzuela A and Garrido A: Biochemical bases of the pharmacological action of the flavonoid silymarin and of its structural isomer silibinin. Biol Res. 27:105–112. 1994.PubMed/NCBI
21	Singh RP and Agarwal R: Cosmeceuticals and silibinin. Clin Dermatol. 27:479–484. 2009. View Article : Google Scholar : PubMed/NCBI
22	Loguercio C and Festi D: Silybin and the liver: From basic research to clinical practice. World J Gastroenterol. 17:2288–2301. 2011. View Article : Google Scholar : PubMed/NCBI
23	Kosina P, Kren V, Gebhardt R, Grambal F, Ulrichová J and Walterová D: Antioxidant properties of silybin glycosides. Phytother Res. 16 Suppl 1:S33–S39. 2002. View Article : Google Scholar : PubMed/NCBI
24	Dehmlow C, Murawski N and de Groot H: Scavenging of reactive oxygen species and inhibition of arachidonic acid metabolism by silibinin in human cells. Life Sci. 58:1591–1600. 1996. View Article : Google Scholar : PubMed/NCBI
25	Tyagi AK, Singh RP, Agarwal C, Chan DC and Agarwal R: Silibinin strongly synergizes human prostate carcinoma DU145 cells to doxorubicin-induced growth Inhibition, G₂-M arrest, and apoptosis. Clin Cancer Res. 8:3512–3519. 2002.PubMed/NCBI
26	Agarwal C, Singh RP, Dhanalakshmi S, Tyagi AK, Tecklenburg M, Sclafani RA and Agarwal R: Silibinin upregulates the expression of cyclin-dependent kinase inhibitors and causes cell cycle arrest and apoptosis in human colon carcinoma HT-29 cells. Oncogene. 22:8271–8282. 2003. View Article : Google Scholar : PubMed/NCBI
27	Bosch-Barrera J and Menendez JA: Silibinin and STAT3: A natural way of targeting transcription factors for cancer therapy. Cancer Treat Rev. 41:540–546. 2015. View Article : Google Scholar : PubMed/NCBI
28	Tyagi A, Agarwal C, Dwyer-Nield LD, Singh RP, Malkinson AM and Agarwal R: Silibinin modulates TNF-α and IFN-γ mediated signaling to regulate COX2 and iNOS expression in tumorigenic mouse lung epithelial LM2 cells. Mol Carcinog. 51:832–842. 2012. View Article : Google Scholar : PubMed/NCBI
29	Lim EJ, Hong DY, Park JH, Joung YH, Darvin P, Kim SY, Na YM, Hwang TS, Ye SK, Moon ES, et al: Methylsulfonylmethane suppresses breast cancer growth by down-regulating STAT3 and STAT5b pathways. PLoS One. 7:e333612012. View Article : Google Scholar : PubMed/NCBI
30	Björklund M and Koivunen E: Gelatinase-mediated migration and invasion of cancer cells. Biochim Biophys Acta. 1755:37–69. 2005.PubMed/NCBI
31	Bauer KR, Brown M, Cress RD, Parise CA and Caggiano V: Descriptive analysis of estrogen receptor (ER)-negative, progesterone receptor (PR)-negative, and HER2-negative invasive breast cancer, the so-called triple-negative phenotype: A population-based study from the California cancer Registry. Cancer. 109:1721–1728. 2007. View Article : Google Scholar : PubMed/NCBI
32	Dent R, Hanna WM, Trudeau M, Rawlinson E, Sun P and Narod SA: Pattern of metastatic spread in triple-negative breast cancer. Breast Cancer Res Treat. 115:423–428. 2009. View Article : Google Scholar : PubMed/NCBI
33	Gluz O, Liedtke C, Gottschalk N, Pusztai L, Nitz U and Harbeck N: Triple-negative breast cancer - current status and future directions. Ann Oncol. 20:1913–1927. 2009. View Article : Google Scholar : PubMed/NCBI
34	Chu SC, Chiou HL, Chen PN, Yang SF and Hsieh YS: Silibinin inhibits the invasion of human lung cancer cells via decreased productions of urokinase-plasminogen activator and matrix metalloproteinase-2. Mol Carcinog. 40:143–149. 2004. View Article : Google Scholar : PubMed/NCBI
35	Nambiar D, Prajapati V, Agarwal R and Singh RP: In vitro and in vivo anticancer efficacy of silibinin against human pancreatic cancer BxPC-3 and PANC-1 cells. Cancer Lett. 334:109–117. 2013. View Article : Google Scholar : PubMed/NCBI
36	Singh RP, Deep G, Blouin MJ, Pollak MN and Agarwal R: Silibinin suppresses in vivo growth of human prostate carcinoma PC-3 tumor xenograft. Carcinogenesis. 28:2567–2574. 2007. View Article : Google Scholar : PubMed/NCBI
37	Yu H, Lee H, Herrmann A, Buettner R and Jove R: Revisiting STAT3 signalling in cancer: New and unexpected biological functions. Nat Rev Cancer. 14:736–746. 2014. View Article : Google Scholar : PubMed/NCBI
38	Darvin P, Joung YH, Sp N, Kang DY, Byun HJ, Hwang DY, Cho KH, Park KD, Lee HK and Yang YM: Sorghum polyphenol suppresses the growth as well as metastasis of colon cancer xenografts through co-targeting jak2/STAT3 and PI3K/Akt/mTOR pathways. J Funct Foods. 15:193–206. 2015. View Article : Google Scholar
39	Darvin P, Baeg SJ, Joung YH, Sp N, Kang DY, Byun HJ, Park JU and Yang YM: Tannic acid inhibits the Jak2/STAT3 pathway and induces G1/S arrest and mitochondrial apoptosis in YD-38 gingival cancer cells. Int J Oncol. 47:1111–1120. 2015.PubMed/NCBI
40	Egeblad M and Werb Z: New functions for the matrix metalloproteinases in cancer progression. Nat Rev Cancer. 2:161–174. 2002. View Article : Google Scholar : PubMed/NCBI
41	Bauvois B: New facets of matrix metalloproteinases MMP-2 and MMP-9 as cell surface transducers: Outside-in signaling and relationship to tumor progression. Biochim Biophys Acta. 1825:29–36. 2012.PubMed/NCBI
42	Yilmaz M and Christofori G: EMT, the cytoskeleton, and cancer cell invasion. Cancer Metastasis Rev. 28:15–33. 2009. View Article : Google Scholar : PubMed/NCBI