title: Compound Library Screening Identified Cardiac Glycoside Digitoxin as an Effective Growth Inhibitor of Gefitinib-Resistant Non-Small Cell Lung Cancer via Downregulation of α-Tubulin and Inhibition of Microtubule Formation.
abstract: Non-small-cell lung cancer (NSCLC) dominates over 85% of all lung cancer cases. Epidermal growth factor receptor (EGFR) activating mutation is a common situation in NSCLC. In the clinic, molecular-targeting with Gefitinib as a tyrosine kinase inhibitor (TKI) for EGFR downstream signaling is initially effective. However, drug resistance frequently happens due to additional mutation on EGFR, such as substitution from threonine to methionine at amino acid position 790 (T790M). In this study, we screened a traditional Chinese medicine (TCM) compound library consisting of 800 single compounds in TKI-resistance NSCLC H1975 cells, which contains substitutions from leucine to arginine at amino acid 858 (L858R) and T790M mutation on EGFR. Attractively, among these compounds there are 24 compounds CC50 of which was less than 2.5 μM were identified. We have further investigated the mechanism of the most effective one, Digitoxin. It showed a significantly cytotoxic effect in H1975 cells by causing G2 phase arrest, also remarkably activated 5' adenosine monophosphate-activated protein kinase (AMPK). Moreover, we first proved that Digitoxin suppressed microtubule formation through decreasing α-tubulin. Therefore, it confirmed that Digitoxin effectively depressed the growth of TKI-resistance NSCLC H1975 cells by inhibiting microtubule polymerization and inducing cell cycle arrest.
authors.affiliation: State Key Laboratory of Quality Research in Chinese Medicine/Macau Institute for Applied Research in Medicine and Health, Macau University of Science and Technology, Taipa, Macau SAR, China. mmmiazh@gmail.com.
authors.forename: Yi-Ze
authors.initials: YZ
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Quality Research in Chinese Medicine/Macau Institute for Applied Research in Medicine and Health, Macau University of Science and Technology, Taipa, Macau SAR, China. aurora329@163.com.
authors.forename: Xi
authors.initials: X
authors.lastname: Chen
authors.affiliation: State Key Laboratory of Quality Research in Chinese Medicine/Macau Institute for Applied Research in Medicine and Health, Macau University of Science and Technology, Taipa, Macau SAR, China. fxx022@163.com.
authors.forename: Xing-Xing
authors.initials: XX
authors.lastname: Fan
authors.affiliation: Department of Thoracic Surgery, The 1st Affiliated Hospital of Guangzhou Medical University, Guangzhou Institute of Respiratory Disease, State Key Laboratory of Respiratory Disease, Guangzhou 510120, China. hejx@vip.163.com.
authors.forename: Jian-Xing
authors.initials: JX
authors.lastname: He
authors.affiliation: Department of Thoracic Surgery, The 1st Affiliated Hospital of Guangzhou Medical University, Guangzhou Institute of Respiratory Disease, State Key Laboratory of Respiratory Disease, Guangzhou 510120, China. kingmal17@126.com.
authors.forename: Jun
authors.initials: J
authors.lastname: Huang
authors.affiliation: Department of Thoracic Surgery, The 1st Affiliated Hospital of Guangzhou Medical University, Guangzhou Institute of Respiratory Disease, State Key Laboratory of Respiratory Disease, Guangzhou 510120, China. oneplusten@126.com.
authors.forename: Da-Kai
authors.initials: DK
authors.lastname: Xiao
authors.affiliation: State Key Laboratory of Quality Research in Chinese Medicine/Macau Institute for Applied Research in Medicine and Health, Macau University of Science and Technology, Taipa, Macau SAR, China. ylzhou@must.edu.mo.
authors.forename: Yan-Ling
authors.initials: YL
authors.lastname: Zhou
authors.affiliation: State Key Laboratory of Quality Research in Chinese Medicine/Macau Institute for Applied Research in Medicine and Health, Macau University of Science and Technology, Taipa, Macau SAR, China. steven4238@hotmail.com.
authors.forename: Sen-You
authors.initials: SY
authors.lastname: Zheng
authors.affiliation: State Key Laboratory of Quality Research in Chinese Medicine/Macau Institute for Applied Research in Medicine and Health, Macau University of Science and Technology, Taipa, Macau SAR, China. ntxjhui@gmail.com.
authors.forename: Jia-Hui
authors.initials: JH
authors.lastname: Xu
authors.affiliation: State Key Laboratory of Quality Research in Chinese Medicine/Macau Institute for Applied Research in Medicine and Health, Macau University of Science and Technology, Taipa, Macau SAR, China. xjyao@must.edu.mo.
authors.forename: Xiao-Jun
authors.initials: XJ
authors.lastname: Yao
authors.affiliation: State Key Laboratory of Quality Research in Chinese Medicine/Macau Institute for Applied Research in Medicine and Health, Macau University of Science and Technology, Taipa, Macau SAR, China. lliu@must.edu.mo.
authors.forename: Liang
authors.initials: L
authors.lastname: Liu
authors.affiliation: State Key Laboratory of Quality Research in Chinese Medicine/Macau Institute for Applied Research in Medicine and Health, Macau University of Science and Technology, Taipa, Macau SAR, China. lhleung@must.edu.mo.
authors.forename: Elaine Lai-Han
authors.initials: EL
authors.lastname: Leung
mesh_terms: D055372:AMP-Activated Protein Kinases / Q000096:biosynthesis; D017209:Apoptosis / Q000187:drug effects; D002289:Carcinoma, Non-Small-Cell Lung / Q000188:drug therapy* / Q000235:genetics; D002301:Cardiac Glycosides / Q000008:administration & dosage* / Q000737:chemistry; D059447:Cell Cycle Checkpoints / Q000187:drug effects; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D004074:Digitoxin / Q000008:administration & dosage* / Q000737:chemistry; D019008:Drug Resistance, Neoplasm / Q000187:drug effects / Q000235:genetics*; D066246:ErbB Receptors / Q000096:biosynthesis / Q000235:genetics; D000077156:Gefitinib; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D008516:Medicine, Chinese Traditional; D008870:Microtubules / Q000187:drug effects* / Q000473:pathology; D009154:Mutation; D011799:Quinazolines / Q000627:therapeutic use; D014404:Tubulin / Q000096:biosynthesis / Q000235:genetics
chemical_list: D002301:Cardiac Glycosides; D011799:Quinazolines; D014404:Tubulin; D004074:Digitoxin; C512478:EGFR protein, human; D066246:ErbB Receptors; D055372:AMP-Activated Protein Kinases; D000077156:Gefitinib
keywords: Digitoxin; EGFR; NSCLC; TKI resistance; cell cycle; microtubule; natural product compound library; α-tubulin