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The reverse effect of X-ray irradiation on acquired gefitinib resistance in non-small cell lung cancer cell line NCI-H1975 in vitro

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Abstract

The clinical efficacy of gefitinib in the treatment of non-small cell lung cancer (NSCLC) with mutations in exon 18, 19 or 21 of epidermal growth factor receptor (EGFR) is limited by the acquired resistance to the drug. To explore whether X-ray irradiation could reverse the acquired gefitinib resistance in NSCLC cell in vitro. We chose a human NSCLC cell line NCI-H1975 to establish acquired gefitinib-resistant cell line named as NCI-H1975/GR. NCI-H1975/GR was irradiated with X-ray and then named as NCI-H1975/GR/XR. In the three cell lines, subsequently cell growth curves and cell population doubling time were calculated by cell proliferation assay, the changes of cell viability were evaluated by trypan blue dye exclusion method and MTT assay, the cell cycle distribution and apoptosis were investigated by flow cytometry, the expressions of E-cadherin and vimentin used to indicate epithelial-mesenchymal transition (EMT) were determined by western blot analysis, the protein expressions in EGFR/KRAS/BRAF transduction pathway were detected by immunocytochemistry, and the mutations of EGFR, KRAS and BRAF were detected by high resolution melting analysis and direct sequencing. We found that the X-ray irradiation enhanced the growth inhibitory effects of gefitinib on the acquired gefitinib-resistant cell line. Of NCI-H1975/GR/XR following gefitinib treatment, the IC50 decreased significantly, the cell proportion of phase G0/G1 was slightly higher, and the apoptosis cell proportion was significantly higher than those of NCI-H1975/GR. In addition, the reversal of EMT being present in NCI-H1975/GR cells was likely appearing in NCI-H1975/GR/XR cells. These results indicated that the acquired gefitinib resistance could be reversed by X-ray irradiation in NSCLC cell line NCI-H1975 harboring both the L858R and T790M mutation in vitro.

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References

  • Ando M, Okamoto I, Yamamoto N, Takeda K, Tamura K, Seto T, Ariyoshi Y, Fukuoka M (2006) Predictive factors for interstitial lung disease, antitumor response, and survival in non-small-cell lung cancer patients treated with gefitinib. J Clin Oncol 24(16):2549–2556

    Article  CAS  PubMed  Google Scholar 

  • Balak MN, Gong Y, Riely GJ, Somwar R, Li AR, Zakowski MF, Chiang A, Yang G, Ouerfelli O, Kris MG, Ladanyi M, Miller VA, Pao W (2006) Novel D761Y and common secondary T790 M mutations in epidermal growth factor receptor-mutant lung adenocarcinomas with acquired resistance to kinase inhibitors. Clin Cancer Res 12(21):6494–6501

    Article  CAS  PubMed  Google Scholar 

  • Bean J, Brennan C, Shih JY, Riely G, Viale A, Wang L, Chitale D, Motoi N, Szoke J, Broderick S, Balak M, Chang WC, Yu CJ, Gazdar A, Pass H, Rusch V, Gerald W, Huang SF, Yang PC, Miller V, Ladanyi M, Yang CH, Pao W (2007) MET amplification occurs with or without T790 M mutations in EGFR mutant lung tumors with acquired resistance to gefitinib or erlotinib. Proc Natl Acad Sci USA 104(52):20932–20937

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Chang CC, Chi KH, Kao SJ, Hsc PS, Tsang YW, Chang HJ, Yeh YW, Hsieh YS, Jiang JS (2011) Upfront gefitinib/erlotinib treatment followed by concomitant radiotherapy for advanced lung cancer: a monoinstitutional experience. Lung Cancer 73(2):189–194

    Article  PubMed  Google Scholar 

  • Colquhoun AJ, Mchugh LA, Tulchinsky E, Kriajevska M, Mellon JK (2007) Combination treatment with ionising radiation and gefitinib (‘Iressa’, ZD1839), an epidermal growth factor receptor (EGFR) inhibitor, significantly inhibits bladder cancer cell growth in vitro and in vivo. J Radiat Res 48(5):351–360

    Article  CAS  PubMed  Google Scholar 

  • Costa DB, Kobayashi S, Tenen DG, Huberman MS (2007) Pooled analysis of the prospective trials of gefitinib monotherapy for EGFR-mutant non-small cell lung cancers. Lung Cancer 58(1):95–103

    Article  PubMed Central  PubMed  Google Scholar 

  • Costa DB, Schumer ST, Tenen DG, Kobayashi S (2008) Differential responses to erlotinib in epidermal growth factor receptor (EGFR)-mutated lung cancers with acquired resistance to gefitinib carrying the L747S or T790 M secondary mutations. J Clin Oncol 26(7):1182–1184

    Article  PubMed  Google Scholar 

  • Dong S, Zhang XC, Cheng H, Zhu JQ, Chen ZH, Zhang YF, Xie Z, Wu LY (2012) Everolimus synergizes with gefitinib in non-small-cell lung cancer cell lines resistant to epidermal growth factor receptor tyrosine kinase inhibitors. Cancer Chemother Pharmacol 70(5):707–716

    Article  CAS  PubMed  Google Scholar 

  • Engelman JA, Zejnullahu K, Mitsudomi T, Song Y, Hyland C, Park JO, Lindeman N, Gale CM, Zhao X, Christensen J, Kosaka T, Holmes AJ, Rogers AM, Cappuzzo F, Mok T, Lee C, Johnson BE, Cantley LC, Jänne PA (2007) MET amplification leads to gefitinib resistance in lung cancer by activating ERBB3 signaling. Science 316(5827):1039–1043

    Article  CAS  PubMed  Google Scholar 

  • Ercan D, Xu C, Yanagita M, Monast CS, Pratilas CA, Montero J, Butaney M, Shimamura T, Sholl L, Ivanova EV, Tadi M, Rogers A, Repellin C, Capelletti M, Maertens O, Goetz EM, Letai A, Garraway LA, Lazzara MJ, Rosen N, Gray NS, Wong KK, Jänne PA (2012) Reactivation of ERK signaling causes resistance to EGFR kinase inhibitors. Cancer Discov 2(10):934–947

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Faber AC, Corcoran RB, Ebi H, Sequist LV, Waltman BA, Chung E, Incio J, Digumarthy SR, Pollack SF, Song Y, Muzikansky A, Lifshits E, Roberge S, Coffman EJ, Benes CH, Gómez HL, Baselga J, Arteaga CL, Rivera MN, Dias-Santagata D, Jain RK, Engelman JA (2011) BIM expression in treatment-naïve cancers predicts responsiveness to kinase inhibitors. Cancer Discov 1(4):352–365

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Fukuoka M, Yano S, Giaccone G, Tamura T, Nakagawa K, Douillard JY, Nishiwaki Y, Vansteenkiste J, Kudoh S, Rischin D, Eek R, Horai T, Noda K, Takata I, Smit E, Averbuch S, Macleod A, Feyereislova A, Dong RP, Baselga J (2003) Multi-institutional randomized phase II trial of gefitinib for previously treated patients with advanced non-small-cell lung cancer. J Clin Oncol 21(12):2237–2246

    Article  CAS  PubMed  Google Scholar 

  • Gazdar AF, Shigematsu H, Herz J, Minna JD (2004) Mutations and addiction to EGFR: the Achilles ‘heal’ of lung cancers? Trends Mol Med 10(10):481–486

    Article  CAS  PubMed  Google Scholar 

  • Hayes DN, Monti S, Parmigiani G, Gilks CB, Naoki K, Bhattacharjee A, Socinski MA, Perou C, Meyerson M (2006) Gene expression profiling reveals re -producible human lung adenocarcinoma subtypes in multiple in dependent patient cohorts. J Clin Oncol 24(31):5079–5090

    Article  CAS  PubMed  Google Scholar 

  • Hu B, Mu HP, Zhang YQ, Su CY, Song JT, Meng C, Liu DX (2014) Prognostic significance of TBX2 expression in non-small cell lung cancer. J Mol Histol. [Epub ahead of print]

  • Huang MH, Lee JH, Chang YJ, Tsai HH, Lin YL, Lin AM, Yang JC (2013) MEK inhibitors reverse resistance in epidermal growth factor receptor mutation lung cancer cells with acquired resistance to gefitinib. Mol Oncol 7(1):112–120

    Article  CAS  PubMed  Google Scholar 

  • Kaneda H, Tamura K, Kurata T, Uejima H, Nakagawa K, Fukuoka M (2004) Retrospective analysis of the predictive factors associated with the response and survival benefit of gefitinib in patients with advanced non-small-cell lung cancer. Lung Cancer 46(2):247–254

    Article  PubMed  Google Scholar 

  • Lynch TJ, Bell DW, Sordella R, Gurubhagavatula S, Okimoto RA, Brannigan BW, Harris PL, Haserlat SM, Supko JG, Haluska FG, Louis DN, Christiani DC, Settleman J, Haber DA (2004) Activating mutations in the epidermal growth factor receptor underlying responsiveness of non-small-cell lung cancer to gefitinib. N Engl J Med 350(21):2129–2139

    Article  CAS  PubMed  Google Scholar 

  • Morabito A, Costanzo R, Rachiglio AM, Pasquale R, Sandomenico C, Franco R, Montanino A, De Lutio E, Rocco G, Normanno N (2013) Activity of gefitinib in a non-small-cell lung cancer patient with both activating and resistance EGFR mutations. J Thorac Oncol 8(7):e59–e60

    Article  CAS  PubMed  Google Scholar 

  • Nguyen KSH, Neal JW (2012) First line treatment of EGFR-mutant non-small-cell lung cancer: the role of erlotinib and other tyrosine kinase inhibitors. Biologics 6:337–344

    CAS  PubMed Central  PubMed  Google Scholar 

  • Ohashi K, Sequist LV, Arcila ME, Moran T, Chmielecki J, Lin YL, Pan Y, Wang L, de Stanchina E, Shien K, Aoe K, Toyooka S, Kiura K, Fernandez-Cuesta L, Fidias P, Yang JC, Miller VA, Riely GJ, Kris MG, Engelman JA, Vnencak-Jones CL, Dias-Santagata D, Ladanyi M, Pao W (2012) Lung cancers with acquired resistance to EGFR inhibitors occasionally harbor BRAF gene mutations but lack mutations in KRAS, NRAS, or MEK1. Proc Natl Acad Sci USA 109(31):E2127–E2133

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Paez JG, Janne PA, Lee JC, Tracy S, Greulich H, Gabriel S, Herman P, Kaye FJ, Lindeman N, Boggon TJ, Naoki K, Sasaki H, Fujii Y, Eck MJ, Sellers WR, Johnson BE, Meyerson M (2004) EGFR mutations in lung cancer: correlation with clinical response to gefitinib therapy. Science 304(5676):1497–1500

    Article  CAS  PubMed  Google Scholar 

  • Pao W, Miller VA (2005) Epidermal growth factor receptor mutations, small-molecule kinase inhibitors, and non-small cell lung cancer: current knowledge and future directions. J Clin Oncol 23(11):2556–2568

    Article  CAS  PubMed  Google Scholar 

  • Pao W, Miller V, Zakowski M, Doherty J, Politi K, Sarkaria I, Singh B, Heelan R, Rusch V, Fulton L, Mardis E, Kupfer D, Wilson R, Kris M, Varmus H (2004) EGF receptor gene mutations are common in lung cancers from ‘‘neversmokers’’ and are associated with sensitivity of tumors to gefitinib and erlotinib. Proc Natl Acad Sci USA 101(36):13306–13311

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Pao W, Miller VA, Politi KA, Riely GJ, Somwar R, Zakowski MF, Kris MG, Varmus H (2005) Acquired resistance of lung adenocarcinomas to gefitinib or erlotinib is associated with a second mutation in the EGFR kinase domain. PLoS Med 2(3):e73

    Article  PubMed Central  PubMed  Google Scholar 

  • Rho JK, Choi YJ, Lee JK, Ryoo BY, Na II, Yang SH, Kim CH, Lee JC (2009) Epithelial to mesenchymal transition derived from repeated exposure to gefitnib determined the sensitivity to EGFR inhibitors in A549, a non-small cell lung cancer cell line. Lung Cancer 63(2):219–226

    Article  PubMed  Google Scholar 

  • Riely GJ, Politi KA, Miller VA, Pao W (2006) Update on epidermal growth factor receptor mutations in non-small cell lung cancer. Clin Cancer Res 12(24):7232–7241

    Article  CAS  PubMed  Google Scholar 

  • Sequist LV, Waltman BA, Dias-Santagata D, Digumarthy S, Turke AB, Fidias P, Bergethon K, Shaw AT, Gettinger S, Cosper AK, Akhavanfard S, Heist RS, Temel J, Christensen JG, Wain JC, Lynch TJ, Vernovsky K, Mark EJ, Lanuti M, Iafrate AJ, Mino-Kenudson M, Engelman JA (2011) Genotypic and histological evolution of lung cancers acquiring resistance to EGFR inhibitors. Sci Trans Med 3(75):75ra26

    Article  Google Scholar 

  • Shigematsu H, Gazdar AF (2006) Somatic mutations of epidermal growth factor receptor signaling pathway in lung cancers. Int J Cancer 118(2):257–262

    Article  CAS  PubMed  Google Scholar 

  • Soria JC, Mok TS, Cappuzzo F, Janne PA (2012) EGFR-mutated oncogene-addicted non-small cell lung cancer: current trends and future prospects. Cancer Treat Rev 38(5):416–430

    Article  CAS  PubMed  Google Scholar 

  • Suda K, Tomizawa K, Fujii M, Murakami H, Osada H, Maehara Y, Yatabe Y, Sekido Y, Mitsudomi T (2011) Epithelial to mesenchymal transition in an epidermal growth factor receptor-mutant lung cancer cell line with acquired resistance to erlotinib. J Thoracic Oncol 6(7):1152–1161

    Article  Google Scholar 

  • Takano T, Ohe Y, Kusumoto M, Tateishi U, Yamamoto S, Nokihara H, Yamamoto N, Sekine I, Kunitoh H, Tamura T, Kodama T, Saijo N (2004) Risk factors for interstitial lung disease and predictive factors for tumor response in patients with advanced non-small cell lung cancer treated with gefitinib. Lung Cancer 45(1):93–104

    Article  PubMed  Google Scholar 

  • Tamura K, Fukuoka M (2005) Gefitinib in non-small cell lung cancer. Expert Opin Pharmacother 6(6):985–993

    Article  CAS  PubMed  Google Scholar 

  • Thomson S, Petti F, Sujka-Kwok I, Mercado P, Bean J, Monaghan M, Seymour SL, Argast GM, Epstein DM, Haley JD (2011) A systems view of epithelial-mesenchymal transition signaling states. Clin Exp Metastasis 28(2):137–155

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Xu Y, Liu H, Chen J, Zhou Q (2010) Acquired resistance of lung adenocarcinoma to EGFR-tyrosine kinase inhibitors gefitinib and erlotinib. Cancer Biol Ther 9(8):572–582

    Article  CAS  PubMed  Google Scholar 

  • Xu RT, Shen H, Guo RH, Sun J, Gao W, Shu YQ (2012) Combine therapy of gefitinib and fulvestrant enhances antitumor effects on NSCLC cell lines with acquired resistance to gefitinib. Biomed Pharmacother 66:384–389

    Article  CAS  PubMed  Google Scholar 

  • Yauch RL, Januario T, Eberhard DA, Cavet G, Zhu W, Fu L, Pham TQ, Soriano R, Stinson J, Seshagiri S, Modrusan Z, Lin CY, O’Neill V, Amler LC (2005) Epithelial versus mesenchymal phenotype determines in vitro sensitivity and predicts clinical activity of erlotinib in lung cancer patients. Clin Cancer Res 11(24 pt 1):8686–8698

    Article  CAS  PubMed  Google Scholar 

  • Zhang T, Zhang DM, Zhao D, Hou XM, Liu XJ, Ling XL, Ma SC (2014) The prognostic value of osteopontin expression in non-small cell lung cancer: a meta-analysis. J Mol Histol. [Epub ahead of print]

  • Zhuang HQ, Wang J, Yuan ZY, Zhao LJ, Wang P, Wang CL (2009) The drug-resistance to gefitinib in PTEN low expression cancer cells is reversed by irradiation in vitro. J Exp Clin Cancer Res 28:123

    Article  PubMed Central  PubMed  Google Scholar 

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Acknowledgments

We thank Dr Yinghai Chen for providing technical assistance. This work was supported by the National Natural Science Foundation of China (81071805). This study was supported by DaLian Merricom gene diagnosis technology Co., Ltd.

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Authors and Affiliations

  1. Dalian Medical University, Dalian, 116044, People’s Republic of China

    Jing Wang, Hong Wei, Weipeng Lv, Ling Lv & Shen Lv

  2. Laboratory Center, Second Affiliated Hospital of Dalian Medical University, 467#, Zhongshan Road, Dalian, 116027, Liaoning Province, People’s Republic of China

    Jing Wang, Baoxia Zhao, Mei Li & Shen Lv

  3. College of Medical Laboratory, Dalian Medical University, Dalian, 116044, People’s Republic of China

    Baoxia Zhao

  4. Department of Pathology, College of Basic Medical Sciences, Dalian Medical University, Dalian, 116044, Liaoning Province, People’s Republic of China

    Bo Song

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  1. Jing Wang
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Correspondence to Shen Lv.

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Wang, J., Wei, H., Zhao, B. et al. The reverse effect of X-ray irradiation on acquired gefitinib resistance in non-small cell lung cancer cell line NCI-H1975 in vitro. J Mol Hist 45, 641–652 (2014). https://doi.org/10.1007/s10735-014-9583-2

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