inhibiting BRAF Oncogene-Mediated Radioresistance Effectively Radiosensitizes BRAF~(V6OOE)-Mutant Thyroid Cancer Cells by Constraining DNA Double-Strand Break Repair

摘要

Purpose: Activating BRAF mutations, most commonly BRAF~(V600E), are a major oncogenic driver of many cancers. We explored whether BRAF~(V600E) promotes radiation resistance and whether selectively targeting BRAF~(V600E) with a BRAF inhibitor (vemurafenib, BRAFi) sensitizes BRAF~(V600E) thyroid cancer cells to radiotherapy. Experimental Design: Immunoblotting, neutral comet, immunocytochemistry, functional reporter, and dono-genic assays were used to analyze the outcome and molecular characteristics following radiotherapy with or without BRAF~(V600E) or vemurafenib in thyroid cancer cells. Results: BRAF~(V600E) thyroid cancer cell lines were associated with resistance to ionizing radiation (IR), and expression of braf~(v600e) into Wild-type BRAF thyroid cancer cells led to IR resistance. BRAFi inhibited ERK signaling in BRAF~(V600E) mutants, but not BRAF wild-type thyroid cancer cell lines. BRAFi selectively radiosensitized and delayed resolution of IR-induced yH2AX nuclear foci in BRAF~(V600E) cells. Moreover, BRAFi impaired global DNA repair and altered the resolution of 53BP1 and RAD51 nuclear foci in BRAF~(V600E) cells following IR. BRAF~(V600E) mutants displayed enhanced nonhomologous end-joining (NHEJ) repair activity, which was abolished by BRAFi. Intriguingly, BRAF~(V600E) mutation led to upregulation of XLF, a component of NHEJ, which was prevented by BRAFi. Importantly, BRAFi in combination with radiotherapy resulted in marked and sustained tumor regression of BRAF~(V600E) thyroid tumor xenografts. Conclusions: BRAF~(V600E) mutation promotes NHEJ activity leading to radioresistance and BRAFi selectively radiosensitizes BRAF~(V600E) thyroid cancer cells through inhibiting NHEJ. Our findings suggest that combining BRAFi and radiation may improve the therapeutic outcome of patients with BRAF~(V600E)-mutant thyroid cancer.