Importance of EGFR/ERCC1 interaction following radiation-induced DNA damage.

Abstract

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 of DNA 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. The EGFR interactome was investigated via mass spectrometry. IR-induced EGFR-ERCC1 binding was validated biochemically and via proximity ligation assay in different cell lines including the M059K and M059J glioma cell lines, proficient and deficient for the expression of DNAPKcs, respectively. EGFR-ERCC1 functional significance following IR-induced SB was investigated in knockdown experiments with the Comet and γH2AX 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. 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 γH2AX 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. These data emphasize the importance of understanding the various mechanisms by which EGFR modulates DNA repair to optimize targeted therapy for patients with cancer.