Cancer Therapy: Preclinical ImportanceofEGFR/ERCC1 InteractionFollowingRadiation- Induced DNA Damage

Purpose: The epidermal growth factor receptor (EGFR) plays an important role in cellular response to chemotherapy and radiotherapy through modulation of DNA repair. EGFR activates DNA-dependent protein kinase (DNA-PK) stimulating repair ofDNA strand breaks (SB) and interstrand crosslinks (ICL).We investigated the role of EGFR in repair of ionizing radiation (IR)-induced SB independently of DNA-PK. ExperimentalDesign: TheEGFR interactomewas investigated viamass spectrometry. IR-induced EGFR– ERCC1bindingwas validated biochemically and via proximity ligation assay in different cell lines including theM059KandM059J glioma cell lines, proficient anddeficient for the expressionofDNAPKcs, respectively. EGFR–ERCC1 functional significance following IR-induced SBwas investigated in knockdown experiments with the Comet and gH2AX foci assays. The effect of this interaction was tested with EGFR–ERCC1 knockdown in combination with gefitinib and NU7026 using the MTT and apoptosis assays. Results: This study demonstrates that EGFR inhibition further impairs IR-induced DNA repair in cells lacking expression of DNAPKcs or in combination with the DNAPK inhibitor NU7026. Our data suggest a role for EGFR in DNA repair independent of DNAPKcs but dependent on ERCC1. Alkaline comet and gH2AX foci assays in cells depleted of EGFR, ERCC1, or EGFR–ERCC1 expression demonstrated involvement of this interaction in DNA repair. Cellular survival and apoptosis data correlate with levels of residual DNA damage underlying the importance of this complex following SB. Conclusion: These data emphasize the importance of understanding the various mechanisms by which EGFRmodulates DNA repair to optimize targeted therapy for patients with cancer. Clin Cancer Res; 20(13); 3496–506. 2014 AACR.