Open Access

Afatinib, an EGFR inhibitor, decreases EMT and tumorigenesis of Huh‑7 cells by regulating the ERK‑VEGF/MMP9 signaling pathway

  • Authors:
    • Yafei Chen
    • Xin Chen
    • Xiaojun Ding
    • Yingwei Wang
  • View Affiliations

  • Published online on: August 6, 2019     https://doi.org/10.3892/mmr.2019.10562
  • Pages: 3317-3325
  • Copyright: © Chen et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

Transcatheter arterial embolization (TAE) therapy has been used in the treatment of inoperable hepatocellular carcinoma (HCC). However, tumor recurrence and metastasis are common in patients after TAE, and these processes may be caused by circulating tumor cells (CTCs). Epithelial‑mesenchymal transition (EMT) serves important roles in CTCs, and abnormal expression and activation of epidermal growth factor receptor (EGFR) is common in cancer cells. Afatinib is an EGFR‑tyrosine kinase inhibitor (TKI). The present study aimed to investigate the effects of afatinib on EMT and tumorigenesis in HCC cells. Western blot analysis suggested that afatinib was able to effectively suppress overactivation of EGFR. Moreover, the expression levels of EMT‑ and metastasis‑associated genes were found to be modulated by afatinib through EGFR inhibition. In addition, Cell Counting Kit‑8 and Transwell assays suggested that the viability, migration and invasion of HCC cells were inhibited by afatinib through EGFR inhibition. Furthermore, the activity of the ERK signaling pathway and the expression levels of vascular endothelial growth factor (VEGF) and matrix metalloproteinase 9 (MMP9) were decreased following treatment with afatinib in vitro. Collectively, the present results suggested that the inhibitory effects of afatinib on EMT and tumorigenesis may be associated with the ERK‑VEGF/MMP9 signaling pathway. The present study provides new insights into understanding the mechanism underlying HCC and may facilitate the development of novel therapeutic strategies to treat HCC recurrence.

References

1 

Burkhart RA, Ronnekleiv-Kelly SM and Pawlik TM: Personalized therapy in hepatocellular carcinoma: Molecular markers of prognosis and therapeutic response. Surg Oncol. 26:138–145. 2017. View Article : Google Scholar : PubMed/NCBI

2 

Torre LA, Bray F, Siegel RL, Ferlay J, Lortet-Tieulent J and Jemal A: Global cancer statistics, 2012. CA Cancer J Clin. 65:87–108. 2015. View Article : Google Scholar : PubMed/NCBI

3 

Nagao E, Hirakawa M, Soeda H, Tsuruta S, Sakai H and Honda H: Transcatheter arterial embolization for chest wall metastasis of hepatocellular carcinoma. World J Radiol. 5:45–48. 2013. View Article : Google Scholar : PubMed/NCBI

4 

Wang R, Zhao N, Li S, Fang JH, Chen MX, Yang J, Jia WH, Yuan Y and Zhuang SM: MicroRNA-195 suppresses angiogenesis and metastasis of hepatocellular carcinoma by inhibiting the expression of VEGF, VAV2, and CDC42. Hepatology. 58:642–653. 2013. View Article : Google Scholar : PubMed/NCBI

5 

Germano D and Daniele B: TIE2-expressing monocytes as a diagnostic marker for hepatocellular carcinoma correlates with angiogenesis. Hepatobiliary Surg Nutr. 3:166–167. 2014.PubMed/NCBI

6 

Serrano MJ, Alvarez-Cubero MJ, De Miguel Pérez D, Rodríguez-Martínez A, Gonzalez-Herrera L, Robles-Fernandez I, Hernandez JE, Puche JLG and Lorente JA: Significance of EGFR expression in circulating tumor cells. Adv Exp Med Biol. 994:285–296. 2017. View Article : Google Scholar : PubMed/NCBI

7 

Schlessinger J: Ligand-induced, receptor-mediated dimerization and activation of EGF receptor. Cell. 110:669–672. 2002. View Article : Google Scholar : PubMed/NCBI

8 

Komposch K and Sibilia M: EGFR signaling in liver diseases. Int J Mol Sci. 17:E302015. View Article : Google Scholar : PubMed/NCBI

9 

Zhong L, Liao D, Zhang M, Zeng C, Li X, Zhang R, Ma H and Kang T: YTHDF2 suppresses cell proliferation and growth via destabilizing the EGFR mRNA in hepatocellular carcinoma. Cancer Lett. 442:252–261. 2019. View Article : Google Scholar : PubMed/NCBI

10 

Zhu L, Liu R, Zhang W, Qian S and Wang J: Application of EGFR inhibitor reduces circulating tumor cells during transcatheter arterial embolization. Clin Transl Oncol. 20:639–646. 2018. View Article : Google Scholar : PubMed/NCBI

11 

Keating GM: Afatinib: A review in advanced non-small cell lung cancer. Target Oncol. 11:825–835. 2016. View Article : Google Scholar : PubMed/NCBI

12 

Rhim AD, Mirek ET, Aiello NM, Maitra A, Bailey JM, McAllister F, Reichert M, Beatty GL, Rustgi AK, Vonderheide RH, et al: EMT and dissemination precede pancreatic tumor formation. Cell. 148:349–361. 2012. View Article : Google Scholar : PubMed/NCBI

13 

Li JY, Huang WX, Zhou X, Chen J and Li Z: Numb inhibits epithelial-mesenchymal transition via RBP-Jκ-dependent Notch1/PTEN/FAK signaling pathway in tongue cancer. BMC Cancer. 19:3912019. View Article : Google Scholar : PubMed/NCBI

14 

Peng H and Li H: The encouraging role of long noncoding RNA small nuclear RNA host gene 16 in epithelial-mesenchymal transition of bladder cancer via directly acting on miR-17-5p/metalloproteinases 3 axis. Mol Carcinog. 2019.(Epub ahead of print). View Article : Google Scholar

15 

Felipe Lima J, Nofech-Mozes S, Bayani J and Bartlett JM: EMT in breast carcinoma-a review. J Clin Med. 5:E652016. View Article : Google Scholar : PubMed/NCBI

16 

Toh Y, Pencil SD and Nicolson GL: A novel candidate metastasis-associated gene, mta1, differentially expressed in highly metastatic mammary adenocarcinoma cell lines. cDNA cloning, expression, and protein analyses. J Biol Chem. 269:22958–22963. 1994.PubMed/NCBI

17 

Toh Y, Pencil SD and Nicolson GL: Analysis of the complete sequence of the novel metastasis-associated candidate gene, mta1, differentially expressed in mammary adenocarcinoma and breast cancer cell lines. Gene. 159:97–104. 1995. View Article : Google Scholar : PubMed/NCBI

18 

Minard ME, Kim LS, Price JE and Gallick GE: The role of the guanine nucleotide exchange factor Tiam1 in cellular migration, invasion, adhesion and tumor progression. Breast Cancer Res Treat. 84:21–32. 2004. View Article : Google Scholar : PubMed/NCBI

19 

Chen B, Ding Y, Liu F, Ruan J, Guan J, Huang J, Ye X, Wang S, Zhang G, Zhang X, et al: Tiam1, overexpressed in most malignancies, is a novel tumor biomarker. Mol Med Rep. 5:48–53. 2012.PubMed/NCBI

20 

Yu LN, Zhang QL, Li X, Hua X, Cui YM, Zhang NJ, Liao WT and Ding YQ: Tiam1 transgenic mice display increased tumor invasive and metastatic potential of colorectal cancer after 1,2-dimethylhydrazine treatment. PLoS One. 8:e730772013. View Article : Google Scholar : PubMed/NCBI

21 

Ferrara N, Gerber HP and LeCouter J: The biology of VEGF and its receptors. Nat Med. 9:669–676. 2003. View Article : Google Scholar : PubMed/NCBI

22 

Kessenbrock K, Plaks V and Werb Z: Matrix metalloproteinases: Regulators of the tumor microenvironment. Cell. 141:52–67. 2010. View Article : Google Scholar : PubMed/NCBI

23 

Stetler-Stevenson WG: Matrix metalloproteinases in angiogenesis: A moving target for therapeutic intervention. J Clin Invest. 103:1237–1241. 1999. View Article : Google Scholar : PubMed/NCBI

24 

Bourboulia D and Stetler-Stevenson WG: Matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs): Positive and negative regulators in tumor cell adhesion. Semin Cancer Biol. 20:161–168. 2010. View Article : Google Scholar : PubMed/NCBI

25 

Gomes E and Rockwell P: p38 MAPK as a negative regulator of VEGF/VEGFR2 signaling pathway in serum deprived human SK-N-SH neuroblastoma cells. Neurosci Lett. 431:95–100. 2008. View Article : Google Scholar : PubMed/NCBI

26 

Wang L, Liu T, Nishioka M, Aguirre RL, Win SS and Okada N: Activation of ERK1/2 and cyclin D1 expression in oral tongue squamous cell carcinomas: Relationship between clinicopathological appearances and cell proliferation. Oral Oncol. 42:625–631. 2006. View Article : Google Scholar : PubMed/NCBI

27 

Li Q and Yang Z: Expression of phospho-ERK1/2 and PI3-K in benign and malignant gallbladder lesions and its clinical and pathological correlations. J Exp Clin Cancer Res. 28:652009. View Article : Google Scholar : PubMed/NCBI

28 

Yang Y, Park H, Yang Y, Kim TS, Bang SI and Cho D: Enhancement of cell migration by corticotropin-releasing hormone through ERK1/2 pathway in murine melanoma cell line, B16F10. Exp Dermatol. 16:22–27. 2007. View Article : Google Scholar : PubMed/NCBI

29 

Henry C, Llamosas E, Knipprath-Meszaros A, Schoetzau A, Obermann E, Fuenfschilling M, Caduff R, Fink D, Hacker N, Ward R, et al: Targeting the ROR1 and ROR2 receptors in epithelial ovarian cancer inhibits cell migration and invasion. Oncotarget. 6:40310–40326. 2015. View Article : Google Scholar : PubMed/NCBI

30 

Arocho A, Chen B, Ladanyi M and Pan Q: Validation of the 2-DeltaDeltaCt calculation as an alternate method of data analysis for quantitative PCR of BCR-ABL P210 transcripts. Diagn Mol Pathol. 15:56–61. 2006. View Article : Google Scholar : PubMed/NCBI

31 

Jayachandran A, Dhungel B and Steel JC: Epithelial-to- mesenchymal plasticity of cancer stem cells: Therapeutic targets in hepatocellular carcinoma. J Hematol Oncol. 9:742016. View Article : Google Scholar : PubMed/NCBI

32 

Wang H, Wu Q, Zhang Y, Zhang HN, Wang YB and Wang W: TGF-β1-induced epithelial-mesenchymal transition in lung cancer cells involves upregulation of miR-9 and downregulation of its target, E-cadherin. Cell Mol Biol Lett. 22:222017. View Article : Google Scholar : PubMed/NCBI

33 

Zhu G, Zhang Y, Wang Q, Che S, Yang Y, Chen L and Lin Z: The prognostic value of Tiam1 correlates with its roles in epithelial-mesenchymal transition progression and angiogenesis in lung adenocarcinoma. Cancer Manag Res. 11:1741–1752. 2019. View Article : Google Scholar : PubMed/NCBI

34 

Liu CY, Lin HH, Tang MJ and Wang YK: Vimentin contributes to epithelial-mesenchymal transition cancer cell mechanics by mediating cytoskeletal organization and focal adhesion maturation. Oncotarget. 6:15966–15983. 2015.PubMed/NCBI

35 

Lin X, Zheng L, Song H, Xiao J, Pan B, Chen H, Jin X and Yu H: Effects of microRNA-183 on epithelial-mesenchymal transition, proliferation, migration, invasion and apoptosis in human pancreatic cancer SW1900 cells by targeting MTA1. Exp Mol Pathol. 102:522–532. 2017. View Article : Google Scholar : PubMed/NCBI

36 

Roskoski R Jr: Small molecule inhibitors targeting the EGFR/ErbB family of protein-tyrosine kinases in human cancers. Pharmacol Res. 139:395–411. 2019. View Article : Google Scholar : PubMed/NCBI

37 

Ding D, Huang H, Jiang W, Yu W, Zhu H, Liu J, Saiyin H, Wu J, Huang H, Jiang S and Yu L: Reticulocalbin-2 enhances hepatocellular carcinoma proliferation via modulating the EGFR-ERK pathway. Oncogene. 36:6691–6700. 2017. View Article : Google Scholar : PubMed/NCBI

38 

Ye QH, Zhu WW, Zhang JB, Qin Y, Lu M, Lin GL, Guo L, Zhang B, Lin ZH, Roessler S, et al: GOLM1 modulates EGFR/RTK cell-surface recycling to drive hepatocellular carcinoma metastasis. Cancer Cell. 30:444–458. 2016. View Article : Google Scholar : PubMed/NCBI

39 

Scaltriti M and Baselga J: The epidermal growth factor receptor pathway: A model for targeted therapy. Clin Cancer Res. 12:5268–5272. 2006. View Article : Google Scholar : PubMed/NCBI

40 

Ikeda S, Tsigelny IF, Skjevik ÅA, Kono Y, Mendler M, Kuo A, Sicklick JK, Heestand G, Banks KC, Talasaz A, et al: Next-generation sequencing of circulating tumor DNA reveals frequent alterations in advanced hepatocellular carcinoma. Oncologist. 23:586–593. 2018. View Article : Google Scholar : PubMed/NCBI

41 

Li R, Yanjiao G, Wubin H, Yue W, Jianhua H, Huachuan Z, Rongjian S and Zhidong L: Secreted GRP78 activates EGFR-SRC-STAT3 signaling and confers the resistance to sorafeinib in HCC cells. Oncotargt. 8:19354–19364. 2017.

42 

Panvichian R, Tantiwetrueangdet A, Sornmayura P and Leelaudomlipi S: Missense mutations in exons 18–24 of EGFR in hepatocellular carcinoma tissues. Biomed Res Int. 2015:1718452015. View Article : Google Scholar : PubMed/NCBI

43 

Chen BH and Giudice LC: Dysfunctional uterine bleeding. West J Med. 169:280–284. 1998.PubMed/NCBI

44 

Solca F, Dahl G, Zoephel A, Bader G, Sanderson M, Klein C, Kraemer O, Himmelsbach F, Haaksma E and Adolf GR: Target binding properties and cellular activity of afatinib (BIBW 2992), an irreversible ErbB family blocker. J Pharmacol Exp Ther. 343:342–350. 2012. View Article : Google Scholar : PubMed/NCBI

45 

Tu Y, Wang C, Yang Z, Zhao B, Lai L, Yang Q, Zheng P and Zhu W: Discovery of novel quinazoline derivatives bearing semicarbazone moiety as potent EGFR kinase inhibitors. Comput Struct Biotechnol J. 16:462–478. 2018. View Article : Google Scholar : PubMed/NCBI

46 

Roof AK and Gutierrez-Hartmann A: Consider the context: Ras/ERK and PI3K/AKT/mTOR signaling outcomes are pituitary cell type-specific. Mol Cell Endocrinol. 463:87–96. 2018. View Article : Google Scholar : PubMed/NCBI

47 

Fang ZT, Wang GZ, Zhang W, Qu XD, Liu R, Qian S, Zhu L, Zhou B and Wang JH: Transcatheter arterial embolization promotes liver tumor metastasis by increasing the population of circulating tumor cells. OncoTargets Ther. 6:1563–1572. 2013.

48 

Ishiyama N, Lee SH, Liu S, Li GY, Smith MJ, Reichardt LF and Ikura M: Dynamic and static interactions between p120 catenin and E-cadherin regulate the stability of cell-cell adhesion. Cell. 141:117–128. 2010. View Article : Google Scholar : PubMed/NCBI

49 

Jiang WG, Grimshaw D, Martin TA, Davies G, Parr C, Watkins G, Lane J, Abounader R, Laterra J and Mansel RE: Reduction of stromal fibroblast-induced mammary tumor growth, by retroviral ribozyme transgenes to hepatocyte growth factor/scatter factor and its receptor, c-MET. Clin Cancer Res. 9:4274–4281. 2003.PubMed/NCBI

50 

Kalluri R and Weinberg RA: The basics of epithelial-mesenchymal transition. J Clin Invest. 119:1420–1428. 2009. View Article : Google Scholar : PubMed/NCBI

51 

Puisieux A: Role of epithelial-mesenchymal transition in tumor progression. Bull Acad Natl Med. 193:2017–2032; discussion 2032–2034. 2009.(In French). PubMed/NCBI

52 

Wang ZC, Gao Q, Shi JY, Guo WJ, Yang LX, Liu XY, Liu LZ, Ma LJ, Duan M, Zhao YJ, et al: Protein tyrosine phosphatase receptor S acts as a metastatic suppressor in hepatocellular carcinoma by control of epithermal growth factor receptor-induced epithelial-mesenchymal transition. Hepatology. 62:1201–1214. 2015. View Article : Google Scholar : PubMed/NCBI

53 

Deng L, Tang J, Yang H, Cheng C, Lu S, Jiang R and Sun B: MTA1 modulated by miR-30e contributes to epithelial-to-mesenchymal transition in hepatocellular carcinoma through an ErbB2-dependent pathway. Oncogene. 36:3976–3985. 2017. View Article : Google Scholar : PubMed/NCBI

54 

Hashida S, Yamamoto H, Shien K, Miyoshi Y, Ohtsuka T, Suzawa K, Watanabe M, Maki Y, Soh J, Asano H, et al: Acquisition of cancer stem cell-like properties in non-small cell lung cancer with acquired resistance to afatinib. Cancer Sci. 106:1377–1384. 2015. View Article : Google Scholar : PubMed/NCBI

55 

Kang X, Lu P, Cui Y, Wang Y, Zhang Q, Gong Y and Xu Z: Bufalin reverses hepatocyte growth factor-induced resistance to afatinib in H1975 lung cancer cells. Zhonghua Zhong Liu Za Zhi. 37:490–496. 2015.(In Chinese). PubMed/NCBI

56 

Song L, Li W, Zhang H, Liao W, Dai T, Yu C, Ding X, Zhang L and Li J: Over-expression of AEG-1 significantly associates with tumour aggressiveness and poor prognosis in human non-small cell lung cancer. J Pathol. 219:317–326. 2009. View Article : Google Scholar : PubMed/NCBI

57 

Jain RK, Duda DG, Clark JW and Loeffler JS: Lessons from phase III clinical trials on anti-VEGF therapy for cancer. Nat Clin Pract Oncol. 3:24–40. 2006. View Article : Google Scholar : PubMed/NCBI

58 

Ng EW, Shima DT, Calias P, Cunningham ET Jr, Guyer DR and Adamis AP: Pegaptanib, a targeted anti-VEGF aptamer for ocular vascular disease. Nat Rev Drug Discov. 5:123–132. 2006. View Article : Google Scholar : PubMed/NCBI

59 

Mukhopadhyay D, Nagy JA, Manseau EJ and Dvorak HF: Vascular permeability factor/vascular endothelial growth factor-mediated signaling in mouse mesentery vascular endothelium. Cancer Res. 58:1278–1284. 1998.PubMed/NCBI

60 

Zhang HH, Zhang Y, Cheng YN, Gong FL, Cao ZQ, Yu LG and Guo XL: Metformin incombination with curcumin inhibits the growth, metastasis, and angiogenesis of hepatocellular carcinoma in vitro and in vivo. Mol Carcinog. 57:44–56. 2018. View Article : Google Scholar : PubMed/NCBI

61 

Brown DM and Regillo CD: Anti-VEGF agents in the treatment of neovascular age-related macular degeneration: Applying clinical trial results to the treatment of everyday patients. Am J Ophthalmol. 144:627–637. 2007. View Article : Google Scholar : PubMed/NCBI

62 

Huang M, Huang B, Li G and Zeng S: Apatinib affect VEGF-mediated cell proliferation, migration, invasion via blocking VEGFR2/RAF/MEK/ERK and PI3K/AKT pathways in cholangiocarcinoma cell. BMC Gastroenterol. 18:1692018. View Article : Google Scholar : PubMed/NCBI

63 

Fearnley GW, Smith GA, Abdul-Zani I, Yuldasheva N, Mughal NA, Homer-Vanniasinkam S, Kearney MT, Zachary IC, Tomlinson DC, Harrison MA, et al: VEGF-A isoforms program differential VEGFR2 signal transduction, trafficking and proteolysis. Biol Open. 5:571–583. 2016. View Article : Google Scholar : PubMed/NCBI

64 

Almalki SG and Agrawal DK: ERK signaling is required for VEGF-A/VEGFR2-induced differentiation of porcine adipose-derived mesenchymal stem cells into endothelial cells. Stem Cell Res Ther. 8:1132017. View Article : Google Scholar : PubMed/NCBI

65 

Mu X, Zhao T, Xu C, Shi W, Geng B, Shen J, Zhang C, Pan J, Yang J, Hu S, et al: Oncometabolite succinate promotes angiogenesis by upregulating VEGF expression through GPR91-mediated STAT3 and ERK activation. Oncotarget. 8:13174–13185. 2017. View Article : Google Scholar : PubMed/NCBI

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October 2019
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APA
Chen, Y., Chen, X., Ding, X., & Wang, Y. (2019). Afatinib, an EGFR inhibitor, decreases EMT and tumorigenesis of Huh‑7 cells by regulating the ERK‑VEGF/MMP9 signaling pathway. Molecular Medicine Reports, 20, 3317-3325. https://doi.org/10.3892/mmr.2019.10562
MLA
Chen, Y., Chen, X., Ding, X., Wang, Y."Afatinib, an EGFR inhibitor, decreases EMT and tumorigenesis of Huh‑7 cells by regulating the ERK‑VEGF/MMP9 signaling pathway". Molecular Medicine Reports 20.4 (2019): 3317-3325.
Chicago
Chen, Y., Chen, X., Ding, X., Wang, Y."Afatinib, an EGFR inhibitor, decreases EMT and tumorigenesis of Huh‑7 cells by regulating the ERK‑VEGF/MMP9 signaling pathway". Molecular Medicine Reports 20, no. 4 (2019): 3317-3325. https://doi.org/10.3892/mmr.2019.10562