Gefitinib Radiosensitizes Non–Small Cell Lung Cancer Cells by Suppressing Cellular DNA Repair Capacity

Abstract

Purpose: Overexpression of the epidermal growth factor receptor (EGFR) promotes unregulated growth, inhibits apoptosis, and likely contributes to clinical radiation resistance of non^small cell lung cancer (NSCLC). Molecular blockade of EGFR signaling is an attractive therapeutic strategy for enhancing the cytotoxic effects of radiotherapy that is currently under investigation in preclinical and clinical studies. In the present study, we have investigated the mechanism by which gefitinib, a selective EGFR tyrosine kinase inhibitor, restores the radiosensitivity of NSCLC cells. Experimental Design: Two NSCLC cell lines, A549 and H1299, were treated with 1 Amol/L gefitinib for 24 h before irradiation and then tested for clonogenic survival and capacity for repairing DNA double strand breaks (DSB). Four different repair assays were used: host cell reactivation, detection of g-H2AX and pNBS1 repair foci using immunofluorescence microscopy, the neutral comet assay, and pulsed-field gel electrophoresis. Results: In clonogenic survival experiments, gefitinib had significant radiosensitizing effects on both cell lines. Results from all four DNA damage repair analyses in cultured A549 and H1299 cells showed that gefitinib had a strong inhibitory effect on the repair of DSBs after ionizing radiation.The presence of DSBs was especially prolonged during the first 2 h of repair compared with controls. Immunoblot analysis of selected repair proteins indicated that pNBS1 activation was prolonged by gefitinib correlating with its effect on pNBS1-labeled repair foci. Conclusions: Overall, we conclude that gefitinib enhances the radioresponse of NSCLC cells by suppressing cellular DNA repair capacity, thereby prolonging the presence of radiation-induced DSBs.

Management of non-small cell lung cancer with EGFR mutation: the role of radiotherapy in the era of tyrosine kinase inhibitor therapy—opportunities and challenges

“…Preclinical studies showed that EGFR TKI, such as gefitinib and erlotinib, have radiosensitizing effects at multiple levels including cell cycle arrest, apoptosis, accelerated repopulation and DNA damage repair (50). However, in previous clinical studies, incorporation of TKI in the combined chemoradiotherapy for locally advanced NSCLC was not associated with improved treatment outcomes (51); possible explanations include the cell cycle specific antagonism when combined with concurrent chemotherapy and the unselected patients in these trials.…”

Downregulation of Annexin A1 is correlated with radioresistance in nasopharyngeal carcinoma

“…It is well known that radioresistance is associated with DNA damage repair, apoptosis and cell cycle checkpoints (31)(32)(33). IR is able to induce DNA damage and apoptosis via reactive oxygen species generated by radiolytic hydrolysis (34).…”

Role of epidermal growth factor receptor in DNA damage repair

“…There are different opinions about whether tyrosine kinase inhibitors that target nuclear kinase activity of EGFR have an effect on the redistribution of DNA-PK. Gefitinib (a tyrosine kinase inhibitor) was observed to suppress nuclear DNA-PK levels in head and neck squamous cell carcinoma cells [13], but not in nonsmall cell lung carcinoma (NSCLC) cell lines A549 [14]. In NSCLC cell lines, unlike wild-type (WT) EGFR, receptors with common oncogenic TKD mutations, L858R or ∆E746-E750, are more sensitive to ionizing radiation and are defective in radiation induced translocation to the nucleus; receptors with these mutations fail to bind the catalytic and regulatory subunits of the DNA-PK.…”

“…Because Nbs1 functions in a protein complex MRN, an abnormal abundance of Nbs1 may sequester other repair proteins away from the sites of DSBs. Nbs1 is a substrate for the ATM kinase; Nbs1 phosphorylation is a possible connection between EGFR>ERK>ATM>Nbs1, and should be examined in further studies [14].…”