title: Attenuation of the DNA damage response by transforming growth factor-beta inhibitors enhances radiation sensitivity of non-small-cell lung cancer cells in vitro and in vivo.
abstract: OBJECTIVE
  To determine whether transforming growth factor (TGF)-β inhibition increases the response to radiation therapy in human and mouse non-small-cell lung carcinoma (NSCLC) cells in vitro and in vivo.
  
  
  METHODS
  TGF-β-mediated growth response and pathway activation were examined in human NSCLC NCI-H1299, NCI-H292, and A549 cell lines and murine Lewis lung cancer (LLC) cells. Cells were treated in vitro with LY364947, a small-molecule inhibitor of the TGF-β type 1 receptor kinase, or with the pan-isoform TGF-β neutralizing monoclonal antibody 1D11 before radiation exposure. The DNA damage response was assessed by ataxia telangiectasia mutated (ATM) or Trp53 protein phosphorylation, γH2AX foci formation, or comet assay in irradiated cells. Radiation sensitivity was determined by clonogenic assay. Mice bearing syngeneic subcutaneous LLC tumors were treated with 5 fractions of 6 Gy and/or neutralizing or control antibody.
  
  
  RESULTS
  The NCI-H1299, A549, and LLC NSCLC cell lines pretreated with LY364947 before radiation exposure exhibited compromised DNA damage response, indicated by decreased ATM and p53 phosphorylation, reduced γH2AX foci, and increased radiosensitivity. The NCI-H292 cells were unresponsive. Transforming growth factor-β signaling inhibition in irradiated LLC cells resulted in unresolved DNA damage. Subcutaneous LLC tumors in mice treated with TGF-β neutralizing antibody exhibited fewer γH2AX foci after irradiation and significantly greater tumor growth delay in combination with fractionated radiation.
  
  
  CONCLUSIONS
  Inhibition of TGF-β before radiation attenuated DNA damage recognition and increased radiosensitivity in most NSCLC cells in vitro and promoted radiation-induced tumor control in vivo. These data support the rationale for concurrent TGF-β inhibition and RT to provide therapeutic benefit in NSCLC.
authors.affiliation: Department of Radiation Oncology, New York University School of Medicine, New York, New York.
authors.forename: Shisuo
authors.initials: S
authors.lastname: Du
authors.affiliation: Department of Radiation Oncology, New York University School of Medicine, New York, New York.
authors.forename: Sophie
authors.initials: S
authors.lastname: Bouquet
authors.affiliation: Department of Radiation Oncology, New York University School of Medicine, New York, New York.
authors.forename: Chen-Hao
authors.initials: CH
authors.lastname: Lo
authors.affiliation: Department of Radiation Oncology, New York University School of Medicine, New York, New York.
authors.forename: Ilenia
authors.initials: I
authors.lastname: Pellicciotta
authors.affiliation: Department of Radiation Oncology, New York University School of Medicine, New York, New York.
authors.forename: Shiva
authors.initials: S
authors.lastname: Bolourchi
authors.affiliation: Varian Medical Systems, Palo Alto, California.
authors.forename: Renate
authors.initials: R
authors.lastname: Parry
authors.affiliation: Department of Radiation Oncology, New York University School of Medicine, New York, New York. Electronic address: mhbarcellos-hoff@nyumc.org.
authors.forename: Mary Helen
authors.initials: MH
authors.lastname: Barcellos-Hoff
mesh_terms: D000596:Amino Acids / Q000494:pharmacology*; D000818:Animals; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D057134:Antibodies, Neutralizing / Q000494:pharmacology; D064007:Ataxia Telangiectasia Mutated Proteins / Q000378:metabolism; D015153:Blotting, Western / Q000379:methods; D002289:Carcinoma, Non-Small-Cell Lung / Q000473:pathology / Q000532:radiotherapy*; D045744:Cell Line, Tumor; D017024:Chemotherapy, Adjuvant; D020552:Comet Assay; D004249:DNA Damage / Q000187:drug effects*; D006657:Histones / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D008175:Lung Neoplasms / Q000473:pathology / Q000532:radiotherapy*; D051379:Mice; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D011836:Radiation Tolerance / Q000187:drug effects*; D000077293:Receptor, Transforming Growth Factor-beta Type I; D018125:Receptors, Transforming Growth Factor beta / Q000037:antagonists & inhibitors / Q000378:metabolism; D016212:Transforming Growth Factor beta / Q000037:antagonists & inhibitors* / Q000378:metabolism; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D014966:Xanthenes / Q000494:pharmacology*
chemical_list: D000596:Amino Acids; D000911:Antibodies, Monoclonal; D057134:Antibodies, Neutralizing; C468783:H2AX protein, human; D006657:Histones; C479758:LY366457; D018125:Receptors, Transforming Growth Factor beta; D016212:Transforming Growth Factor beta; D016159:Tumor Suppressor Protein p53; D014966:Xanthenes; D064007:Ataxia Telangiectasia Mutated Proteins; D017346:Protein Serine-Threonine Kinases; D000077293:Receptor, Transforming Growth Factor-beta Type I