BUB1 regulates non-homologous end joining pathway to mediate radioresistance in triple-negative breast cancer
Recommended Citation
Sriramulu S, Thoidingjam S, Chen WM, Hassan O, Siddiqui F, Brown SL, Movsas B, Green MD, Davis AJ, Speers C, Walker E, and Nyati S. BUB1 regulates non-homologous end joining pathway to mediate radioresistance in triple-negative breast cancer. J Exp Clin Cancer Res 2024; 43(1):163.
Document Type
Article
Publication Date
6-11-2024
Publication Title
Journal of experimental & clinical cancer research
Abstract
BACKGROUND: Triple-negative breast cancer (TNBC) is a highly aggressive form of breast cancer subtype often treated with radiotherapy (RT). Due to its intrinsic heterogeneity and lack of effective targets, it is crucial to identify novel molecular targets that would increase RT efficacy. Here we demonstrate the role of BUB1 (cell cycle Ser/Thr kinase) in TNBC radioresistance and offer a novel strategy to improve TNBC treatment.
METHODS: Gene expression analysis was performed to look at genes upregulated in TNBC patient samples compared to other subtypes. Cell proliferation and clonogenic survivals assays determined the IC(50) of BUB1 inhibitor (BAY1816032) and radiation enhancement ratio (rER) with pharmacologic and genomic BUB1 inhibition. Mammary fat pad xenografts experiments were performed in CB17/SCID. The mechanism through which BUB1 inhibitor sensitizes TNBC cells to radiotherapy was delineated by γ-H2AX foci assays, BLRR, Immunoblotting, qPCR, CHX chase, and cell fractionation assays.
RESULTS: BUB1 is overexpressed in BC and its expression is considerably elevated in TNBC with poor survival outcomes. Pharmacological or genomic ablation of BUB1 sensitized multiple TNBC cell lines to cell killing by radiation, although breast epithelial cells showed no radiosensitization with BUB1 inhibition. Kinase function of BUB1 is mainly accountable for this radiosensitization phenotype. BUB1 ablation also led to radiosensitization in TNBC tumor xenografts with significantly increased tumor growth delay and overall survival. Mechanistically, BUB1 ablation inhibited the repair of radiation-induced DNA double strand breaks (DSBs). BUB1 ablation stabilized phospho-DNAPKcs (S2056) following RT such that half-lives could not be estimated. In contrast, RT alone caused BUB1 stabilization, but pre-treatment with BUB1 inhibitor prevented stabilization (t(1/2), ~8 h). Nuclear and chromatin-enriched fractionations illustrated an increase in recruitment of phospho- and total-DNAPK, and KAP1 to chromatin indicating that BUB1 is indispensable in the activation and recruitment of non-homologous end joining (NHEJ) proteins to DSBs. Additionally, BUB1 staining of TNBC tissue microarrays demonstrated significant correlation of BUB1 protein expression with tumor grade.
CONCLUSIONS: BUB1 ablation sensitizes TNBC cell lines and xenografts to RT and BUB1 mediated radiosensitization may occur through NHEJ. Together, these results highlight BUB1 as a novel molecular target for radiosensitization in women with TNBC.
Medical Subject Headings
Humans; Triple Negative Breast Neoplasms; Animals; Female; Mice; Radiation Tolerance; Cell Line, Tumor; DNA End-Joining Repair; Protein Serine-Threonine Kinases; Xenograft Model Antitumor Assays; Cell Proliferation; Gene Expression Regulation, Neoplastic; Mice, SCID
PubMed ID
38863037
Volume
43
Issue
1
First Page
163
Last Page
163
