title: Targeting mTOR to overcome epidermal growth factor receptor tyrosine kinase inhibitor resistance in non-small cell lung cancer cells.
abstract: OBJECTIVE
  Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) have shown dramatic clinical benefits in advanced non-small cell lung cancer (NSCLC); however, resistance remains a serious problem in clinical practice. The present study analyzed mTOR-associated signaling-pathway differences between the EGFR TKI-sensitive and -resistant NSCLC cell lines and investigated the feasibility of targeting mTOR with specific mTOR inhibitor in EGFR TKI resistant NSCLC cells.
  
  
  METHODS
  We selected four different types of EGFR TKI-sensitive and -resistant NSCLC cells: PC9, PC9GR, H1650 and H1975 cells as models to detect mTOR-associated signaling-pathway differences by western blot and Immunoprecipitation and evaluated the antiproliferative effect and cell cycle arrest of ku-0063794 by MTT method and flow cytometry.
  
  
  RESULTS
  In the present study, we observed that mTORC2-associated Akt ser473-FOXO1 signaling pathway in a basal state was highly activated in resistant cells. In vitro mTORC1 and mTORC2 kinase activities assays showed that EGFR TKI-resistant NSCLC cell lines had higher mTORC2 kinase activity, whereas sensitive cells had higher mTORC1 kinase activity in the basal state. The ATP-competitive mTOR inhibitor ku-0063794 showed dramatic antiproliferative effects and G1-cell cycle arrest in both sensitive and resistant cells. Ku-0063794 at the IC50 concentration effectively inhibited both mTOR and p70S6K phosphorylation levels; the latter is an mTORC1 substrate and did not upregulate Akt ser473 phosphorylation which would be induced by rapamycin and resulted in partial inhibition of FOXO1 phosphorylation. We also observed that EGFR TKI-sensitive and -resistant clinical NSCLC tumor specimens had higher total and phosphorylated p70S6K expression levels.
  
  
  CONCLUSIONS
  Our results indicate mTORC2-associated signaling-pathway was hyperactivated in EGFR TKI-resistant cells and targeting mTOR with specific mTOR inhibitors is likely a good strategy for patients with EGFR mutant NSCLC who develop EGFR TKI resistance; the potential specific roles of mTORC2 in EGFR TKI-resistant NSCLC cells were still unknown and should be further investigated.
authors.affiliation: Guangdong Lung Cancer Institute, Medical Research Center of Guangdong General Hospital and Guangdong Academy of Medical Sciences, Guangzhou, China.
authors.forename: Shi-Jiang
authors.initials: SJ
authors.lastname: Fei
authors.forename: Xu-Chao
authors.initials: XC
authors.lastname: Zhang
authors.forename: Song
authors.initials: S
authors.lastname: Dong
authors.forename: Hua
authors.initials: H
authors.lastname: Cheng
authors.forename: Yi-Fang
authors.initials: YF
authors.lastname: Zhang
authors.forename: Ling
authors.initials: L
authors.lastname: Huang
authors.forename: Hai-Yu
authors.initials: HY
authors.lastname: Zhou
authors.forename: Zhi
authors.initials: Z
authors.lastname: Xie
authors.forename: Zhi-Hong
authors.initials: ZH
authors.lastname: Chen
authors.forename: Yi-Long
authors.initials: YL
authors.lastname: Wu
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology*; D002289:Carcinoma, Non-Small-Cell Lung / Q000188:drug therapy / Q000378:metabolism*; D045744:Cell Line, Tumor; D019008:Drug Resistance, Neoplasm / Q000187:drug effects; D066246:ErbB Receptors / Q000037:antagonists & inhibitors* / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D009025:Morpholines / Q000494:pharmacology*; D047428:Protein Kinase Inhibitors / Q000627:therapeutic use*; D011743:Pyrimidines / Q000494:pharmacology*; D015398:Signal Transduction / Q000187:drug effects; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism*
chemical_list: D000970:Antineoplastic Agents; D009025:Morpholines; D047428:Protein Kinase Inhibitors; D011743:Pyrimidines; C541932:Ku 0063794; C546842:MTOR protein, human; D066246:ErbB Receptors; D058570:TOR Serine-Threonine Kinases