title: Resistance to tyrosine kinase inhibition by mutant epidermal growth factor receptor variant III contributes to the neoplastic phenotype of glioblastoma multiforme.
abstract: OBJECTIVE
  We have reported previously that tumors expressing wild-type epidermal growth factor receptor (EGFR) in a murine model are sensitive to the EGFR tyrosine kinase inhibitor gefitinib, whereas tumors expressing mutant EGFR variant III (EGFRvIII) are resistant. Determination of how this differential inhibition occurs may be important to patient selection and treatment criteria, as well as the design of future therapeutics for glioblastoma multiforme.
  
  
  METHODS
  We have determined and quantified how treatment with gefitinib at commonly used, noncytotoxic doses affects neoplastic functions ascribed to EGFRvIII, including downstream signaling by Akt, DNA synthesis, and cellular invasion. In doing so, we have tested and compared a series of wild-type and mutant EGFRvIII-expressing fibroblast and glioblastoma cell lines in vitro after treatment with gefitinib.
  
  
  RESULTS
  The results of these experiments demonstrate that short-term treatment with gefitinib (approximately 24 h) does not reduce phosphorylation of EGFRvIII, whereas EGFR phosphorylation is inhibited in a dose-dependent manner. However, after daily treatment with gefitinib, phosphorylation declines for EGFRvIII by day 3 and later. Nevertheless, after 7 days of daily treatment, cells that express and are dependent on EGFRvIII for tumorigenic growth are not effectively growth inhibited. This may be due in part to phosphorylation of Akt, which is inhibited in EGFR-expressing cells after treatment with gefitinib, but is unaffected in cells expressing EGFRvIII. Cell cycle analysis shows that nascent DNA synthesis in EGFR-expressing cells is inhibited in a dose-dependent manner by gefitinib, yet is unaffected in EGFRvIII-expressing cells with increasing dosage. Furthermore, cells expressing EGFRvIII demonstrate greater invasive capability with increasing gefitinib concentration when compared with cells expressing EGFR after treatment.
  
  
  CONCLUSIONS
  We conclude that the neoplastic phenotype of EGFRvIII is relatively resistant to gefitinib and requires higher doses, repeated dosing, and longer exposure to decrease receptor phosphorylation. However, this decrease does not effectively inhibit the biologically relevant processes of DNA synthesis, cellular growth, and invasion in cells expressing EGFRvIII.
authors.affiliation: Department of Surgery, Division of Neurosurgery, Duke University Medical Center, Durham, North Carolina 27710, USA.
authors.forename: Chris A
authors.initials: CA
authors.lastname: Learn
authors.forename: Tristan L
authors.initials: TL
authors.lastname: Hartzell
authors.forename: Carol J
authors.initials: CJ
authors.lastname: Wikstrand
authors.forename: Gary E
authors.initials: GE
authors.lastname: Archer
authors.forename: Jeremy N
authors.initials: JN
authors.lastname: Rich
authors.forename: Allan H
authors.initials: AH
authors.lastname: Friedman
authors.forename: Henry S
authors.initials: HS
authors.lastname: Friedman
authors.forename: Darell D
authors.initials: DD
authors.lastname: Bigner
authors.forename: John H
authors.initials: JH
authors.lastname: Sampson
mesh_terms: D000818:Animals; D002455:Cell Division / Q000187:drug effects / Q000235:genetics; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000187:drug effects / Q000235:genetics; D004305:Dose-Response Relationship, Drug; D019008:Drug Resistance, Neoplasm / Q000235:genetics; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D066246:ErbB Receptors / Q000235:genetics* / Q000378:metabolism; D000077156:Gefitinib; D005909:Glioblastoma / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D006801:Humans; D051379:Mice; D009154:Mutation; D010766:Phosphorylation / Q000187:drug effects; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt; D011799:Quinazolines / Q000494:pharmacology*; D041701:Swiss 3T3 Cells; D013997:Time Factors
chemical_list: D004791:Enzyme Inhibitors; D011518:Proto-Oncogene Proteins; D011799:Quinazolines; D066246:ErbB Receptors; D011505:Protein-Tyrosine Kinases; C494918:AKT1 protein, human; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D000077156:Gefitinib