K-RAS(V12) induces autocrine production of EGFR ligands and mediates radioresistance through EGFR-dependent Akt signaling and activation of DNA-PKcs

Int J Radiat Oncol Biol Phys. 2011 Dec 1;81(5):1506-14. doi: 10.1016/j.ijrobp.2011.05.057. Epub 2011 Oct 8.

Abstract

Purpose: 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 and materials: 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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amphiregulin
  • Antibodies, Monoclonal / pharmacology
  • Cell Line, Tumor
  • Cell Survival / physiology
  • Culture Media, Conditioned
  • DNA Breaks, Double-Stranded
  • DNA Repair / physiology*
  • DNA-Activated Protein Kinase / metabolism*
  • EGF Family of Proteins
  • ErbB Receptors / metabolism*
  • Genes, ras
  • Glycoproteins / immunology
  • Glycoproteins / metabolism*
  • Humans
  • Intercellular Signaling Peptides and Proteins / immunology
  • Intercellular Signaling Peptides and Proteins / metabolism*
  • Ligands
  • Mutation
  • Phosphatidylinositol 3-Kinase / metabolism
  • Phosphorylation
  • Proto-Oncogene Proteins c-akt / metabolism
  • Proto-Oncogene Proteins p21(ras) / genetics
  • Proto-Oncogene Proteins p21(ras) / physiology*
  • Radiation Tolerance / physiology*
  • Signal Transduction / physiology
  • Transfection
  • Transforming Growth Factor alpha / metabolism*

Substances

  • AREG protein, human
  • Amphiregulin
  • Antibodies, Monoclonal
  • Culture Media, Conditioned
  • EGF Family of Proteins
  • Glycoproteins
  • Intercellular Signaling Peptides and Proteins
  • Ligands
  • Transforming Growth Factor alpha
  • Phosphatidylinositol 3-Kinase
  • ErbB Receptors
  • DNA-Activated Protein Kinase
  • Proto-Oncogene Proteins c-akt
  • Proto-Oncogene Proteins p21(ras)