Identification of hub genes for the diagnosis and prognosis in triple negative breast cancer using transcriptome and differential methylation integration analysis.

Abstract

Introduction: Triple-negative breast cancer (TNBC) is characterized by the absence of estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2) expression. It is highly invasive and aggressive, making it the subtype of breast cancer with the poorest prognosis. Currently, systemic chemotherapy is the primary treatment option, but targeted therapies remain unavailable. Therefore, there is an urgent need to identify novel biomarkers for the early diagnosis and treatment of TNBC. Methods: We conducted an integrated analysis of transcriptome and methylation data to identify methylation-regulated differentially expressed genes (MDEGs). Gene Ontology (GO) analysis, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, and protein-protein interaction (PPI) network analysis were performed on MDEGs to investigate the impact of hub genes on the diagnosis and prognosis of TNBC. Subsequently, the expression levels and DNA methylation patterns of key genes were validated in the TNBC cell line MDA-MB-231 and the normal breast epithelial cell line MCF-10A using reverse transcription quantitative PCR (RT-qPCR) and quantitative methylation-specific PCR (qMSP). Results: A total of 98 upregulated and 87 downregulated genes were identified through transcriptomic profiling integration analysis. By incorporating methylation data, we further identified 22 genes with high expression of hypomethylation (hypo-MDEGs) and 32 genes with low expression of hypermethylation (hyper-MDEGs). The hypo-MDEGs were primarily involved in nuclear division, organelle fission, spindle formation, chromosome and kinetochore development, and protein binding. KEGG pathway analysis revealed that these genes were enriched in progesterone-mediated oocyte maturation, cell cycle regulation, and oocyte meiosis. Hyper-MDEGs were associated with cell proliferation, hormone response, pain, extracellular matrix composition, and binding to sulfur compounds, heparin, and glycosaminoglycans. PPI network analysis identified seven hub genes-EXO1, KIF11, FOXM1, CENPF, CCNB1, PLK1, and KIF23-which were all significantly overexpressed in TNBC tissues and positively correlated with each other (p < 0.05). Receiver operating characteristic curve analysis showed that the area under the curve (AUC) for all seven genes exceeded 0.9 (p < 0.05), suggesting strong diagnostic potential. Kaplan-Meier survival analysis indicated that KIF11, CCNB1, and PLK1 were associated with a higher hazard ratio (HR > 1, p < 0.05) in TNBC. In vitro validation experiments demonstrated that, compared to MCF-10A cells, MDA-MB-231 cells exhibited higher mRNA expression levels of KIF11, CCNB1, and PLK1, while their DNA methylation levels were lower. Conclusions: This study identified seven hypo-MDEGs, including EXO1, KIF11, FOXM1, CENPF, CCNB1, PLK1, and KIF23, which are involved in the regulation of the cell cycle and mitotic processes and have significant potential as diagnostic biomarkers for TNBC. Notably, elevated expression of KIF11, CCNB1, and PLK1 is associated with poor prognosis in patients with TNBC. These findings contribute to an improved understanding of the epigenetic molecular mechanisms underlying TNBC progression and highlight novel biomarkers that may enhance the accuracy of TNBC diagnosis and provide potential targets for therapeutic intervention.