Downregulation of both EGFR and ErbB3 improves the cellular response to pemetrexed in an established pemetrexed‑resistant lung adenocarcinoma A549 cell line

Yu,Zhuang; Li,Xiu-Mei; Liu,Shi-Hai; Liu,Bing; Gao,Cai-Hong; Hou,Xin

doi:10.3892/or.2014.3027

Downregulation of both EGFR and ErbB3 improves the cellular response to pemetrexed in an established pemetrexed‑resistant lung adenocarcinoma A549 cell line

Authors:
- Zhuang Yu
- Xiu-Mei Li
- Shi-Hai Liu
- Bing Liu
- Cai-Hong Gao
- Xin Hou
View Affiliations

Affiliations: Department of Oncology, The Affiliated Hospital of Qingdao University, Qingdao 266003, P.R. China, Central Laboratory, The Affiliated Hospital of Qingdao University, Qingdao 266003, P.R. China, Department of Medicine, The Commercial Workers' Hospital of Qingdao, Qingdao 266011, P.R. China
Published online on: February 18, 2014 https://doi.org/10.3892/or.2014.3027
Pages: 1818-1824

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

Epidermal growth factor receptor (EGFR) and ErbB3 (HER3) play important roles in the regulation of cell proliferation, differentiation, anti-apoptosis and chemoresistance; however, their dysregulation in pemetrexed (PEM) resistance remains unclear. The aim of the present study was to clarify the relationship between PEM resistance and gene expression of EGFR and ErbB3, by establishing the PEM-resistant lung adenocarcinoma A549 cell line, A549/PEM. Compared with A549 cells, the A549/PEM cells were significantly more resistant to PEM (P=0.0024). The downregulation of S phase and arrest at G1 stage were detected in the A549/PEM cell line when compared to the A549 cells (P<0.05). The apoptosis rate of A549/PEM cells was much lower than that of the A549 cells after a 24 h continuous exposure to PEM (P<0.001). Real-time PCR and western blotting demonstrated the overexpression of EGFR and ErbB3 in A549/PEM cells. However, downregulation of EGFR or ErbB3 by lentiviral delivered shRNAs in A549/PEM cells showed no significant correlation with PEM sensitivity while silencing both EGFR and ErbB3 increased the cellular response to PEM in the A549/PEM cells and significantly decreased phosphorylation of STAT3, AKT and ERK. Together, these data suggest that either high expression of EGFR or ErbB3 plays a critical role in the cellular response to PEM in human lung adenocarcinoma cells though EGFR/ErbB3-dependent pathways.

View Figures

View References

1	Jemal A, Bray F, Center MM, Ferlay J, Ward E and Forman D: Global cancer statistics. CA Cancer J Clin. 61:69–90. 2011. View Article : Google Scholar
2	Herbst RS, Heymach JV and Lippman SM: Lung cancer. N Engl J Med. 359:1367–1380. 2008. View Article : Google Scholar : PubMed/NCBI
3	Schiller JH, Harrington D, Belani CP, et al: Comparison of four chemotherapy regimens for advanced non-small-cell lung cancer. N Engl J Med. 346:92–98. 2002. View Article : Google Scholar
4	Assaraf YG: Molecular basis of antifolate resistance. Cancer Metastasis Rev. 26:153–181. 2007. View Article : Google Scholar
5	Furrukh M, Al-Moundhri M, Zahid KF, Kumar S and Burney I: Customised, individualised treatment of metastatic non-small-cell lung carcinoma (NSCLC). Sultan Qaboos Univ Med J. 13:202–217. 2013. View Article : Google Scholar : PubMed/NCBI
6	Longo-Sorbello GS, Chen B, Budak-Alpdogan T and Bertino JR: Role of pemetrexed in non-small cell lung cancer. Cancer Invest. 25:59–66. 2007. View Article : Google Scholar
7	Scagliotti GV, Parikh P, von Pawel J, et al: Phase III study comparing cisplatin plus gemcitabine with cisplatin plus pemetrexed in chemotherapy-naive patients with advanced-stage non-small-cell lung cancer. J Clin Oncol. 26:3543–3551. 2008. View Article : Google Scholar : PubMed/NCBI
8	Esteban E, Casillas M and Cassinello A: Pemetrexed in first-line treatment of non-small cell lung cancer. Cancer Treat Rev. 35:364–373. 2009. View Article : Google Scholar : PubMed/NCBI
9	Ceppi P, Volante M, Saviozzi S, et al: Squamous cell carcinoma of the lung compared with other histotypes shows higher messenger RNA and protein levels for thymidylate synthase. Cancer. 107:1589–1596. 2006. View Article : Google Scholar : PubMed/NCBI
10	Zhang D, Ochi N, Takigawa N, et al: Establishment of pemetrexed-resistant non-small cell lung cancer cell lines. Cancer Lett. 309:228–235. 2011. View Article : Google Scholar : PubMed/NCBI
11	Wu SG, Yang CH, Yu CJ, et al: Good response to pemetrexed in patients of lung adenocarcinoma with epidermal growth factor receptor (EGFR) mutations. Lung Cancer. 72:333–339. 2011. View Article : Google Scholar : PubMed/NCBI
12	Benavente S, Huang S, Armstrong EA, et al: Establishment and characterization of a model of acquired resistance to epidermal growth factor receptor targeting agents in human cancer cells. Clin Cancer Res. 15:1585–1592. 2009. View Article : Google Scholar : PubMed/NCBI
13	Giltnane JM, Moeder CB, Camp RL and Rimm DL: Quantitative multiplexed analysis of ErbB family coexpression for primary breast cancer prognosis in a large retrospective cohort. Cancer. 115:2400–2409. 2009. View Article : Google Scholar : PubMed/NCBI
14	Sharma SV and Settleman J: ErbBs in lung cancer. Exp Cell Res. 315:557–571. 2009. View Article : Google Scholar : PubMed/NCBI
15	Hayashi M, Inokuchi M, Takagi Y, et al: High expression of HER3 is associated with a decreased survival in gastric cancer. Clin Cancer Res. 14:7843–7849. 2008. View Article : Google Scholar : PubMed/NCBI
16	Mimeault M, Hauke R and Batra SK: Recent advances on the molecular mechanisms involved in the drug resistance of cancer cells and novel targeting therapies. Clin Pharmacol Ther. 83:673–691. 2008. View Article : Google Scholar : PubMed/NCBI
17	Schmidt M and Lichtner RB: EGF receptor targeting in therapy-resistant human tumors. Drug Resist Updat. 5:11–18. 2002. View Article : Google Scholar : PubMed/NCBI
18	Desbois-Mouthon C: The HER3/ErbB3 receptor: a promising target in cancer drug therapy. Gastroenterol Clin Biol. 34:255–259. 2010. View Article : Google Scholar : PubMed/NCBI
19	Blivet-Van Eggelpoël MJ, Chettouh H, Fartoux L, et al: Epidermal growth factor receptor and HER-3 restrict cell response to sorafenib in hepatocellular carcinoma cells. J Hepatol. 57:108–115. 2012.PubMed/NCBI
20	Lu S, Zheng W, Ji L, et al: Synthesis, characterization, screening and docking analysis of 4-anilinoquinazoline derivatives as tyrosine kinase inhibitors. Eur J Med Chem. 61:84–94. 2013. View Article : Google Scholar : PubMed/NCBI
21	Zhou Y, Ling XL, Li SW, Li XQ and Yan B: Establishment of a human hepatoma multidrug resistant cell line in vitro. World J Gastroenterol. 16:2291–2297. 2010. View Article : Google Scholar : PubMed/NCBI
22	Ikari A, Sato T, Watanabe R, Yamazaki Y and Sugatani J: Increase in claudin-2 expression by an EGFR/MEK/ERK/c-Fos pathway in lung adenocarcinoma A549 cells. Biochim Biophys Acta. 1823:1110–1118. 2012. View Article : Google Scholar : PubMed/NCBI
23	Kaszubiak A, Holm PS and Lage H: Overcoming the classical multidrug resistance phenotype by adenoviral delivery of anti-MDR1 short hairpin RNAs and ribozymes. Int J Oncol. 31:419–430. 2007.PubMed/NCBI
24	Amati P and Lago C: Sensitivity to amphotericin B of in vitro established cell lines. Nature. 247:466–469. 1974. View Article : Google Scholar : PubMed/NCBI
25	Snow K and Judd W: Characterisation of adriamycin- and amsacrine-resistant human leukaemic T cell lines. Br J Cancer. 63:17–28. 1991. View Article : Google Scholar : PubMed/NCBI
26	Lu X, Errington J, Curtin NJ, Lunec J and Newell DR: The impact of p53 status on cellular sensitivity to antifolate drugs. Clin Cancer Res. 7:2114–2123. 2001.
27	Hsu HF, Huang KH, Lu KJ, et al: Typhonium blumei extract inhibits proliferation of human lung adenocarcinoma A549 cells via induction of cell cycle arrest and apoptosis. J Ethnopharmacol. 135:492–500. 2011. View Article : Google Scholar
28	Heinemann L, Simpson GR, Annels NE, et al: The effect of cell cycle synchronization on tumor sensitivity to reovirus oncolysis. Mol Ther. 18:2085–2093. 2010. View Article : Google Scholar : PubMed/NCBI
29	Hanauske AR, Eismann U, Oberschmidt O, et al: In vitro chemosensitivity of freshly explanted tumor cells to pemetrexed is correlated with target gene expression. Invest New Drugs. 25:417–423. 2007. View Article : Google Scholar : PubMed/NCBI
30	Ozasa H, Oguri T, Uemura T, et al: Significance of thymidylate synthase for resistance to pemetrexed in lung cancer. Cancer Sci. 101:161–166. 2010. View Article : Google Scholar : PubMed/NCBI
31	Wu MF, Hsiao YM, Huang CF, et al: Genetic determinants of pemetrexed responsiveness and nonresponsiveness in non-small cell lung cancer cells. J Thorac Oncol. 5:1143–1151. 2010. View Article : Google Scholar : PubMed/NCBI
32	Sebastian S, Settleman J, Reshkin SJ, Azzariti A, Bellizzi A and Paradiso A: The complexity of targeting EGFR signalling in cancer: from expression to turnover. Biochim Biophys Acta. 1766:120–139. 2006.PubMed/NCBI
33	Durmuş Tekir S, Yalçin Arga K and Ulgen KO: Drug targets for tumorigenesis: insights from structural analysis of EGFR signaling network. J Biomed Inform. 42:228–236. 2009.PubMed/NCBI
34	Fresno Vara JA, Casado E, de Castro J, Cejas P, Belda-Iniesta C and González-Barón M: PI3K/Akt signalling pathway and cancer. Cancer Treat Rev. 30:193–204. 2004.PubMed/NCBI
35	Kosaka T, Yatabe Y, Endoh H, et al: Analysis of epidermal growth factor receptor gene mutation in patients with non-small cell lung cancer and acquired resistance to gefitinib. Clin Cancer Res. 12:5764–5769. 2006. View Article : Google Scholar : PubMed/NCBI
36	Yamamoto H, Toyooka S and Mitsudomi T: Impact of EGFR mutation analysis in non-small cell lung cancer. Lung Cancer. 63:315–321. 2009.
37	Cappuzzo F, Hirsch FR, Rossi E, et al: Epidermal growth factor receptor gene and protein and gefitinib sensitivity in non-small-cell lung cancer. J Natl Cancer Inst. 97:643–655. 2005. View Article : Google Scholar : PubMed/NCBI
38	Grøvdal LM, Kim J, Holst MR, Knudsen SL, Grandal MV and van Deurs B: EGF receptor inhibitors increase ErbB3 mRNA and protein levels in breast cancer cells. Cell Signal. 24:296–301. 2012.PubMed/NCBI
39	Kawano O, Sasaki H, Endo K, et al: ErbB3 mRNA expression correlated with specific clinicopathologic features of Japanese lung cancers. J Surg Res. 146:43–48. 2008. View Article : Google Scholar : PubMed/NCBI