title: Acquisition of cancer stem cell-like properties in non-small cell lung cancer with acquired resistance to afatinib.
abstract: Afatinib is an irreversible epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitor (TKI) that is known to be effective against the EGFR T790M variant, which accounts for half of the mechanisms of acquired resistance to reversible EGFR-TKIs. However, acquired resistance to afatinib was also observed in clinical use. Thus, elucidating and overcoming the mechanisms of resistance are important issues in the treatment of non-small cell lung cancer. In this study, we established various afatinib-resistant cell lines and investigated the resistance mechanisms. EGFR T790M mutations were not detected using direct sequencing in established resistant cells. Several afatinib-resistant cell lines displayed MET amplification, and these cells were sensitive to the combination of afatinib plus crizotinib. As a further investigation, a cell line that acquired resistance to afatinib plus crizotinib, HCC827-ACR, was established from one of the MET amplified-cell lines. Several afatinib-resistant cell lines including HCC827-ACR displayed epithelial-to-mesenchymal transition (EMT) features and epigenetic silencing of miR-200c, which is a suppresser of EMT. In addition, these cell lines also exhibited overexpression of ALDH1A1 and ABCB1, which are putative stem cell markers, and resistance to docetaxel. In conclusion, we established afatinib-resistant cells and found that MET amplification, EMT, and stem cell-like features are observed in cells with acquired resistance to EGFR-TKIs. This finding may provide clues to overcoming resistance to EGFR-TKIs.
authors.affiliation: Department of Thoracic, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan.|Department of Clinical Genomic Medicine, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan.
authors.forename: Shinsuke
authors.identifier: http://orcid.org/0000-0001-8643-5037
authors.initials: S
authors.lastname: Hashida
authors.affiliation: Department of Thoracic, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan.
authors.forename: Hiromasa
authors.initials: H
authors.lastname: Yamamoto
authors.affiliation: Department of Thoracic, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan.|Department of Clinical Genomic Medicine, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan.
authors.forename: Kazuhiko
authors.initials: K
authors.lastname: Shien
authors.affiliation: Department of Thoracic, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan.
authors.forename: Yuichiro
authors.initials: Y
authors.lastname: Miyoshi
authors.affiliation: Department of Thoracic, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan.
authors.forename: Tomoaki
authors.initials: T
authors.lastname: Ohtsuka
authors.affiliation: Department of Thoracic, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan.
authors.forename: Ken
authors.initials: K
authors.lastname: Suzawa
authors.affiliation: Department of Thoracic, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan.
authors.forename: Mototsugu
authors.initials: M
authors.lastname: Watanabe
authors.affiliation: Department of Thoracic, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan.
authors.forename: Yuho
authors.initials: Y
authors.lastname: Maki
authors.affiliation: Department of Thoracic, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan.
authors.forename: Junichi
authors.initials: J
authors.lastname: Soh
authors.affiliation: Department of Thoracic, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan.
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Asano
authors.affiliation: Department of Thoracic, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan.
authors.forename: Kazunori
authors.initials: K
authors.lastname: Tsukuda
authors.affiliation: Department of Thoracic, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan.
authors.forename: Shinichiro
authors.initials: S
authors.lastname: Miyoshi
authors.affiliation: Department of Thoracic, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan.|Department of Clinical Genomic Medicine, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan.
authors.forename: Shinichi
authors.initials: S
authors.lastname: Toyooka
mesh_terms: D018435:ATP Binding Cassette Transporter, Subfamily B / Q000096:biosynthesis; D000077716:Afatinib; D000444:Aldehyde Dehydrogenase / Q000096:biosynthesis; D000080924:Aldehyde Dehydrogenase 1 Family; D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology; D001483:Base Sequence; D002289:Carcinoma, Non-Small-Cell Lung / Q000235:genetics / Q000473:pathology*; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D000077547:Crizotinib; D019175:DNA Methylation / Q000235:genetics; D000077143:Docetaxel; D019008:Drug Resistance, Neoplasm; D058750:Epithelial-Mesenchymal Transition / Q000187:drug effects; D066246:ErbB Receptors / Q000037:antagonists & inhibitors; D005260:Female; D006801:Humans; D008175:Lung Neoplasms / Q000235:genetics / Q000473:pathology*; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D035683:MicroRNAs / Q000235:genetics; D009154:Mutation / Q000187:drug effects; D014411:Neoplastic Stem Cells / Q000187:drug effects* / Q000473:pathology; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors; D011720:Pyrazoles / Q000494:pharmacology; D011725:Pyridines / Q000494:pharmacology; D011799:Quinazolines / Q000494:pharmacology*; D050697:Retinal Dehydrogenase; D017422:Sequence Analysis, DNA; D043823:Taxoids / Q000494:pharmacology
chemical_list: C513055:ABCB1 protein, human; D018435:ATP Binding Cassette Transporter, Subfamily B; D000970:Antineoplastic Agents; C533896:MIRN200 microRNA, human; D035683:MicroRNAs; D047428:Protein Kinase Inhibitors; D011720:Pyrazoles; D011725:Pyridines; D011799:Quinazolines; D043823:Taxoids; D000077143:Docetaxel; D000077716:Afatinib; D000077547:Crizotinib; D000080924:Aldehyde Dehydrogenase 1 Family; D000444:Aldehyde Dehydrogenase; C510223:ALDH1A1 protein, human; D050697:Retinal Dehydrogenase; C512478:EGFR protein, human; D066246:ErbB Receptors; D011505:Protein-Tyrosine Kinases
keywords: Afatinib; EGFR-TKI; cancer stem cells; drug resistance; non-small cell lung cancer