Loss of EGFR confers acquired resistance to AZD9291 in an EGFR-mutant non-small cell lung cancer cell line with an epithelial–mesenchymal transition phenotype
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AZD9291 is an irreversible, small-molecule inhibitor which has potency against mutant EGFR- and T790M-resistant mutation. Despite the encouraging efficacy in clinical, the acquired resistance will finally occur. Further study will need to be done to identify the acquired resistance mechanisms and determine the next treatment.
We established an AZD9291-resistant cell line (HCC827/AZDR) from parental HCC827 cell line through stepwise pulsed selection of AZD9291. The expression of EGFR and its downstream pathways were determined by western blot analysis or immunofluorescence assay. The sensitivity to indicated agents were evaluated by MTS.
Compared with parental HCC827 cells, the HCC827/AZDR cells showed high resistance to AZD9291 and other EGFR-TKIs, and exhibited a mesenchymal-like phenotype. Almost complete loss of EGFR expression was observed in HCC827/AZDR cells. But the activation of downstream pathway, MAPK signaling, was found in HCC827/AZDR cells even in the presence of AZD9291. Inhibition of MAPK signaling had no effect on cell viability of HCC827/AZDR and could not reverse AZD9291 resistance because of the subsequent activation of AKT signaling. When treated with the combination of AKT and MAPK inhibitor, HCC827/AZDR showed remarkable growth inhibition.
Loss of EGFR could be proposed as a potential acquired resistance mechanism of AZD9291 in EGFR-mutant NSCLC cells with an EMT phenotype. Despite the loss of EGFR, the activation of MAPK pathway which had crosstalk with AKT pathway could maintain the proliferation and survival of resistant cells. Blocking MAPK and AKT signaling may be a potential therapeutic strategy following AZD9291 resistance.
KeywordsAZD9291 Resistance EGFR loss NSCLC Potential therapy
Bovine serum albumin
Breakthrough therapy designation
Epidermal growth factor receptor
Fetal bovine serum
Mitogen-activated protein kinases
Non-small cell lung cancer
Overall response rate
Protein Kinase B
Quality of life
Tyrosine kinase inhibitors
We would like to thank Beta Pharma (Zhejiang, China) and Shihe Gene Bio-Technology (Nanjing, China) for some technical assistance.
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflict of interest. The authors are responsible for the content and writing of the paper.
Research involving human participants and/or animals
This article does not contain studies of human participants or animals performed by any of the authors.
- Byers LA, Diao L, Wang J, Saintigny P, Girard L et al (2013) An epithelial-mesenchymal transition gene signature predicts resistance to EGFR and PI3K inhibitors and identifies Axl as a therapeutic target for overcoming EGFR inhibitor resistance. Clin Cancer Res 19:279–290. https://doi.org/10.1158/1078-0432.CCR-12-1558 CrossRefPubMedGoogle Scholar
- Del RM, Tiseo M, Bordi P, D’Incecco A, Camerini A et al (2016) Contribution of KRAS mutations and c.2369C> T (p.T790M) EGFR to acquired resistance to EGFR-TKIs in EGFR mutant NSCLC: a study on circulating tumor. DNA Oncotarget 29:266–276. https://doi.org/10.18632/oncotarget.6957 CrossRefGoogle Scholar
- Della Corte CM, Malapelle U, Vigliar E, Pepe F, Troncone G et al (2017) Efficacy of continuous EGFR-inhibition and role of Hedgehog in EGFR acquired resistance in human lung cancer cells with activating mutation of. EGFR Oncotarget 8:23020–23032. https://doi.org/10.18632/oncotarget.15479 PubMedCrossRefGoogle Scholar
- Eberlein CA, Stetson D, Markovets AA, Al-Kadhimi KJ, Lai Z et al (2015) Acquired resistance to the mutant-selective EGFR inhibitor AZD9291 is associated with increased dependence on RAS signaling in preclinical. Models Cancer Res 75:2489–2500. https://doi.org/10.1158/0008-5472.CAN-14-3167 CrossRefPubMedGoogle Scholar
- Fukuoka M, Wu YL, Thongprasert S, Sunpaweravong P, Leong SS et al (2011) Biomarker analyses and final overall survival results from a phase III, randomized, open-label, first-line study of gefitinib versus carboplatin/paclitaxel in clinically selected patients with advanced non-small-cell lung cancer in Asia (IPASS). J Clin Oncol 29:2866–2874. https://doi.org/10.1200/jco.2010.33.4235 CrossRefPubMedGoogle Scholar
- Fustaino V, Presutti D, Colombo T, Cardinali B, Papoff G et al (2017) Characterization of epithelial-mesenchymal transition intermediate/hybrid phenotypes associated to resistance to EGFR inhibitors in non-small cell lung cancer cell lines. Oncotarget 8:103340–103363. https://doi.org/10.18632/oncotarget.21132 CrossRefPubMedPubMedCentralGoogle Scholar
- Goss G, Tsai C-M, Shepherd FA, Bazhenova L, Lee JS et al (2016) Osimertinib for pretreated EGFR Thr790Met-positive advanced non-small-cell lung cancer (AURA2): a multicentre, open-label, single-arm, phase 2 study. Lancet Oncol 17:1643–1652. https://doi.org/10.1016/s1470-2045(16)30508-3 CrossRefPubMedGoogle Scholar
- Hirai H, Sootome H, Nakatsuru Y, Miyama K, Taguchi S et al (2010) MK-2206, an allosteric Akt inhibitor, enhances antitumor efficacy by standard chemotherapeutic agents or molecular targeted drugs in vitro and in vivo. Mol Cancer Ther 9:1956–1967. https://doi.org/10.1158/1535-7163.mct-09-1012 CrossRefPubMedGoogle Scholar
- Jin Y, Shao Y, Shi X, Lou G, Zhang Y et al (2016) Mutational profiling of non-small-cell lung cancer patients resistant to first-generation EGFR tyrosine kinase inhibitors using next generation sequencing. Oncotarget 7:61755–61763. https://doi.org/10.18632/oncotarget.11237 PubMedPubMedCentralCrossRefGoogle Scholar
- Ma C, Wei S, Song Y (2011) T790M and acquired resistance of EGFR TKI: a literature review of clinical reports. J Thorac Dis 3:10–18. https://doi.org/10.3978/j.issn.2072-1439.2010.12.02 PubMedPubMedCentralCrossRefGoogle Scholar
- Mirzoeva OK, Das D, Heiser LM, Bhattacharya S, Siwak D et al (2009) Basal subtype and MAPK/ERK kinase (MEK)-phosphoinositide 3-kinase feedback signaling determine susceptibility of breast cancer cells to MEK inhibition. Cancer Res 69:565–572. https://doi.org/10.1158/0008-5472.CAN-08-3389 CrossRefPubMedPubMedCentralGoogle Scholar
- Niederst MJ, Hu H, Mulvey HE, Lockerman EL, Garcia AR et al (2015) The allelic context of the C797S mutation acquired upon treatment with third-generation EGFR inhibitors impacts sensitivity to subsequent treatment strategies. Clin Cancer Res 21:3924–3933. https://doi.org/10.1158/1078-0432.ccr-15-0560 CrossRefPubMedPubMedCentralGoogle Scholar
- Ramalingam S, Reungwetwattana T, Chewaskulyong B (2017a) Osimertinib vs standard of care (SoC) EGFR-TKI as first-line therapy in patients (pts) with EGFRm advanced NSCLC: FLAURA. Paper presented at the EUROPEAN SOCIETY FOR MEDICAL ONCOLOGY 2017 (ESMO 2017)Google Scholar
- Remon J, Moran T, Majem M, Reguart N, Dalmau E, Marquez-Medina D, Lianes P (2014) Acquired resistance to epidermal growth factor receptor tyrosine kinase inhibitors in EGFR-mutant non-small cell lung cancer: a new era begins. Cancer Treat Rev 40:93–101. https://doi.org/10.1016/j.ctrv.2013.06.002 CrossRefPubMedGoogle Scholar
- Wu YL, Zhou C, Hu CP, Feng J, Lu S et al (2014) Afatinib versus cisplatin plus gemcitabine for first-line treatment of Asian patients with advanced non-small-cell lung cancer harbouring EGFR mutations (LUX-Lung 6): an open-label, randomised phase 3 trial. Lancet Oncol 15:213–222. https://doi.org/10.1016/s1470-2045(13)70604-1 CrossRefPubMedGoogle Scholar
- Yang J, Ramalingam SS, Jänne PA, Cantarini M, Mitsudomi T (2016) LBA2_PR: Osimertinib (AZD9291) in pre-treated pts with T790M-positive advanced NSCLC: updated Phase 1 (P1) and pooled Phase 2 (P2) results. J Thorac Oncol 11:S152-S153. https://doi.org/10.1016/s1556-0864(16)30325-2 CrossRefGoogle Scholar
- Zhou C, Wu YL, Chen G, Feng J, Liu XQ et al (2011) Erlotinib versus chemotherapy as first-line treatment for patients with advanced EGFR mutation-positive non-small-cell lung cancer (OPTIMAL, CTONG-0802): a multicentre, open-label, randomised, phase 3 study. Lancet Oncol 12:735–742. https://doi.org/10.1016/s1470-2045(11)70184-x CrossRefPubMedGoogle Scholar