title: K-RAS(V12) induces autocrine production of EGFR ligands and mediates radioresistance through EGFR-dependent Akt signaling and activation of DNA-PKcs.
abstract: OBJECTIVE
  It is known that postirradiation survival of tumor cells presenting mutated K-RAS is mediated through autocrine activation of epidermal growth factor receptor (EGFR). In this study the molecular mechanism of radioresistance of cells overexpressing mutated K-RAS(V12) was investigated.
  
  
  METHODS
  Head-and-neck cancer cells (FaDu) presenting wild-type K-RAS were transfected with empty vector or vector expressing mutated K-RAS(V12). The effect of K-RAS(V12) on autocrine production of EGFR ligands, activation of EGFR downstream pathways, DNA damage repair, and postirradiation survival was analyzed.
  
  
  RESULTS
  Conditioned medium collected from K-RAS(V12)-transfected cells enhanced activation of the phosphatidylinositol-3-kinase-Akt pathway and increased postirradiation survival of wild-type K-RAS parental cells when compared with controls. These effects were reversed by amphiregulin (AREG)-neutralizing antibody. In addition, secretion of the EGFR ligands AREG and transforming growth factor α was significantly increased upon overexpression of K-RAS(V12). Expression of mutated K-RAS(V12) resulted in an increase in radiation-induced DNA-dependent protein kinase catalytic subunit (DNA-PKcs) phosphorylation at S2056. This increase was accompanied by increased repair of DNA double-strand breaks. Abrogation of DNA-PKcs phosphorylation by serum depletion or AREG-neutralizing antibody underscored the role of autocrine production of EGFR ligands, namely, AREG, in regulating DNA-PKcs activation in K-RAS mutated cells.
  
  
  CONCLUSIONS
  These data indicate that radioresistance of K-RAS mutated tumor cells is at least in part due to constitutive production of EGFR ligands, which mediate enhanced repair of DNA double-strand breaks through the EGFR-phosphatidylinositol-3-kinase-Akt cascade.
authors: Division of Radiobiology and Molecular Environmental Research, Department of Radiation Oncology, Eberhard Karls University, Tübingen, Germany.
authors: Minjmaa
authors: M
authors: Minjgee
authors: Mahmoud
authors: M
authors: Toulany
authors: Rainer
authors: R
authors: Kehlbach
authors: Klaudia
authors: K
authors: Giehl
authors: H Peter
authors: HP
authors: Rodemann
mesh_terms: D066258:Amphiregulin; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000502:physiology; D017077:Culture Media, Conditioned; D053903:DNA Breaks, Double-Stranded; D004260:DNA Repair / Q000502:physiology*; D051747:DNA-Activated Protein Kinase / Q000378:metabolism*; D066255:EGF Family of Proteins; D066246:ErbB Receptors / Q000378:metabolism*; D011905:Genes, ras; D006023:Glycoproteins / Q000276:immunology / Q000378:metabolism*; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000276:immunology / Q000378:metabolism*; D008024:Ligands; D009154:Mutation; D058539:Phosphatidylinositol 3-Kinase / Q000378:metabolism; D010766:Phosphorylation; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D016283:Proto-Oncogene Proteins p21(ras) / Q000235:genetics / Q000502:physiology*; D011836:Radiation Tolerance / Q000502:physiology*; D015398:Signal Transduction / Q000502:physiology; D014162:Transfection; D016211:Transforming Growth Factor alpha / Q000378:metabolism*
chemical_list: C587848:AREG protein, human; D066258:Amphiregulin; D000911:Antibodies, Monoclonal; D017077:Culture Media, Conditioned; D066255:EGF Family of Proteins; D006023:Glycoproteins; D036341:Intercellular Signaling Peptides and Proteins; D008024:Ligands; D016211:Transforming Growth Factor alpha; D058539:Phosphatidylinositol 3-Kinase; D066246:ErbB Receptors; D051747:DNA-Activated Protein Kinase; D051057:Proto-Oncogene Proteins c-akt; D016283:Proto-Oncogene Proteins p21(ras)