Pro-survival AKT and ERK signaling from EGFR and mutant EGFRvIII enhances DNA double-strand break repair in human glioma cells

Abstract

The Epidermal growth factor receptor (EGFR) is frequently dysregulated in malignant glioma that leads to increased resistance to cancer therapy. Up-regulation of wild type or expression of mutants such as EGFR variant III (EGFRvIII), the most common EGFR mutation in malignant glioma, is associated with tumor radioresistance and poor clinical outcome. This radioresistance is thought to be the result of a strong cytoprotective response fueled by signaling via AKT and ERK that is heightened by radiation in the clinical dose range. Several groups including ours have shown that this response may modulate DNA repair. Herein, we show that expression of EGFRvIII promoted Pro-survival AKT and ERK signaling from EGFR and mutant EGFRvIII enhances DNA double-strand break repair in

human glioma cells γ-H2AX foci resolution, a surrogate for double-strand break (DSB) repair, and thus enhanced DNA repair. Conversely, small molecule inhibitors targeting EGFR, MEK, and the expression of dominant-negative EGFR (EGFR-CD533) significantly reduced the resolution of γ-H2AX foci. When homologous recombination repair (HRR) and non-homologous end joining (NHEJ) were specifically examined, we found that EGFRvIII stimulated and CD533 compromised HRR and NHEJ, respectively. Furthermore, NHEJ was significantly blocked by inhibitors of AKT and ERK signaling pathways. Moreover, expression of EGFRvIII and CD533 increased and reduced, respectively, the formation of phospho-DNA-PKcs and -ATM repair foci, and RAD51 expression levels, indicating that DSB repair is regulated at multiple levels. Altogether, signaling from EGFR and EGFRvIII promotes both HRR and NHEJ that is likely one contributing factor towards the radioresistance of malignant gliomas.