Time- and dose-dependent regulation of circular RNAs in the response of triple-negative breast cancer cells to ionizing radiation

BACKGROUND: Triple-negative breast cancer (TNBC) is the most aggressive breast cancer subtype, with few treatment options and an increased risk of radioresistance development. Circular RNAs (circRNAs) are non-coding molecules that have emerged as major regulators of cancer progression and therapy efficacy. In this study, we aimed to elucidate their role in radiation response mechanisms of TNBC cell lines. METHODS: BT-20, MDA-MB-231, and MDA-MB-468 cells were exposed to two different doses of ionizing radiation and analyzed at multiple time points to elucidate the expression profiles of ten circRNAs with regulatory potential, namely circTP53, circFOXO3, circCCNB1, circABCC1, circFBXW7, circHIPK3, circNCOR1, circADAM9, circUBAP2, and circATAD2. These circRNAs were pre-amplified and quantified using quantitative real-time PCR with divergent primers, while p53 phosphorylation was assessed by western blot analysis. Next, bioinformatics analysis was performed to identify miRNA targets and enriched signaling pathways for selected circRNAs. RESULTS: Ionizing radiation induced distinct temporal and dose-dependent circRNA expression patterns. Six circRNAs were characterized by progressive upregulation, peaking at 24 h, with circTP53 expression paralleling p53 phosphorylation patterns. Four circRNAs showed early suppression with limited recovery. Cell line-specific responses reflected TNBC molecular heterogeneity, with MDA-MB-231 cells displaying unique recovery patterns. In addition, functional analysis for circTP53, circFBXW7, and circATAD2 revealed enrichment in critical pathways including p53 signaling, FOXO regulation, MAPK signaling, and ubiquitin-mediated proteolysis. CONCLUSION: The cell line-specific radiation-dependent circRNA expression patterns reflect the molecular diversity of TNBC, and pathway enrichments indicate the potential applications of these circRNAs as radiation sensitivity biomarkers and therapeutic targets to improve TNBC treatment efficacy.