title: Small Molecule T315 Promotes Casitas B-Lineage Lymphoma-Dependent Degradation of Epidermal Growth Factor Receptor via Y1045 Autophosphorylation.
abstract: BACKGROUND
  Despite the fact that tyrosine kinase inhibitors (TKIs) have been found effective in treating patients harboring activating mutations of epidermal growth factor receptor (EGFR), an acquired secondary mutation, T790M, which lowers the affinity to TKIs, can lead to EGFR TKI resistance after this standard treatment.
  
  
  OBJECTIVE
  To evaluate the effect of small molecule T315 on EGFR degradation and its therapeutic efficacy in vitro and in vivo.
  
  
  METHODS
  Lung adenocarcinoma cells were treated with T315, and cell proliferation and apoptotic proportion were determined by the CellTiter 96 AQueous MTS (3-[4,5-dimethylthiazol-2-yl]-5-[3-carboxymethoxyphenyl]-2-[4-sulfophenyl]-2H-tetrazolium, inner salt) assay and flow cytometry. The effects of T315 on EGFR mRNA and protein levels, autophosphorylation, ubiquitination, and degradation were evaluated by real-time polymerase chain reaction and Western blot, respectively. Direct targeting of T315 to EGFR was confirmed by the in vitro kinase assay and mass spectrometry. Finally, the preclinical effect of T315 was validated in the murine xenograft model in combination with a second-generation TKI, afatinib.
  
  
  RESULTS
  We identified T315 as a novel, potent small molecule for suppressing cancer cell proliferation in vitro and in vivo. The therapeutic effect was verified after T315 was combined with a second-generation TKI, afatinib, compared with a single drug administration. We found a new mechanism of action, in that T315 appears to directly bind EGFR and triggers EGFR-Y1045 autophosphorylation, whereby its degradation is triggered through the ubiquitin-proteasome pathway.
  
  
  CONCLUSIONS
  Our evidence suggests that T315 is a novel class of anticancer drug that is able to inhibit the growth of EGFR-TKI-resistant lung adenocarcinoma cells by inducing the degradation of EGFR.
authors.affiliation: 1 Graduate Institute of Life Sciences, National Defense Medical Center, Taipei, Taiwan.
authors.forename: Kuo-Yen
authors.initials: KY
authors.lastname: Huang
authors.affiliation: 2 Research Center for Tumor Medical Science, China Medical University, Taichung, Taiwan.
authors.forename: Shih-Han
authors.initials: SH
authors.lastname: Kao
authors.affiliation: 3 Institute of Biomedical Sciences.
authors.forename: Wen-Lung
authors.initials: WL
authors.lastname: Wang
authors.affiliation: 4 Research Center for Industry of Human Ecology.|5 Graduate Institute of Health Industry Technology, Chang Gung University of Science and Technology, Kwei-San, Tao-Yuan, Taiwan.
authors.forename: Chi-Yuan
authors.initials: CY
authors.lastname: Chen
authors.affiliation: 6 Department of Medical Research, Taichung Veterans General Hospital, Taichung, Taiwan.
authors.forename: Tzu-Hung
authors.initials: TH
authors.lastname: Hsiao
authors.affiliation: 7 Division of Medicinal Chemistry and Pharmacognosy, College of Pharmacy, and.
authors.forename: Santosh B
authors.initials: SB
authors.lastname: Salunke
authors.affiliation: 8 Institute of Biomedical Sciences, National Chung Hsing University, Taichung, Taiwan.
authors.forename: Jeremy J W
authors.initials: JJ
authors.lastname: Chen
authors.affiliation: 9 Department of Clinical and Laboratory Sciences and Medical Biotechnology, National Taiwan University College of Medicine, Taipei, Taiwan.|10 NTU Center of Genomic Medicine, College of Medicine, National Taiwan University, Taipei, Taiwan.
authors.forename: Kang-Yi
authors.initials: KY
authors.lastname: Su
authors.affiliation: 3 Institute of Biomedical Sciences.
authors.forename: Shuenn-Chen
authors.initials: SC
authors.lastname: Yang
authors.affiliation: 11 Institute of Clinical Medicine, National Cheng Kung University College of Medicine, Tainan, Taiwan; and.
authors.forename: Tse-Ming
authors.initials: TM
authors.lastname: Hong
authors.affiliation: 12 Institute of Biological Chemistry, Academia Sinica, Taipei, Taiwan.|13 Division of Medicinal Chemistry, College of Pharmacy and Comprehensive Cancer Center, The Ohio State University, Columbus, Ohio.
authors.forename: Ching-Shih
authors.initials: CS
authors.lastname: Chen
authors.affiliation: 1 Graduate Institute of Life Sciences, National Defense Medical Center, Taipei, Taiwan.|3 Institute of Biomedical Sciences.|10 NTU Center of Genomic Medicine, College of Medicine, National Taiwan University, Taipei, Taiwan.|14 Department of Internal Medicine, National Taiwan University Hospital, Taipei, Taiwan.
authors.forename: Pan-Chyr
authors.initials: PC
authors.lastname: Yang
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000187:drug effects / Q000235:genetics; D000230:Adenocarcinoma / Q000188:drug therapy* / Q000235:genetics; D000077192:Adenocarcinoma of Lung; D000077716:Afatinib; D000818:Animals; D000970:Antineoplastic Agents / Q000009:adverse effects / Q000627:therapeutic use; D015153:Blotting, Western; D049109:Cell Proliferation / Q000187:drug effects; D004338:Drug Combinations; D019008:Drug Resistance, Neoplasm / Q000187:drug effects*; D057075:Enzyme Assays; D066246:ErbB Receptors / Q000037:antagonists & inhibitors* / Q000187:drug effects / Q000235:genetics; D006801:Humans; D008175:Lung Neoplasms / Q000188:drug therapy* / Q000235:genetics; D013058:Mass Spectrometry; D051379:Mice; D009154:Mutation / Q000235:genetics; D047428:Protein Kinase Inhibitors / Q000009:adverse effects / Q000627:therapeutic use*; D050721:Proto-Oncogene Proteins c-cbl / Q000187:drug effects / Q000235:genetics; D011799:Quinazolines / Q000009:adverse effects / Q000627:therapeutic use*; D060888:Real-Time Polymerase Chain Reaction; D014407:Tumor Cells, Cultured; D023041:Xenograft Model Antitumor Assays
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D000970:Antineoplastic Agents; C118387:Cblb protein, mouse; D004338:Drug Combinations; D047428:Protein Kinase Inhibitors; D011799:Quinazolines; D000077716:Afatinib; D050721:Proto-Oncogene Proteins c-cbl; C512478:EGFR protein, human; D066246:ErbB Receptors
keywords: T315; casitas B-lineage lymphoma; epidermal growth factor receptor degradation; lung adenocarcinoma