Integrated Bioinformatics and Experimental Validation of the hsa_circ_0000378/miR-205-5p/RAD51 ceRNA Axis in Breast...

Breast cancer (BC) remains the leading cause of cancer-related mortality in women worldwide, primarily due to its high invasiveness and therapeutic resistance. This study explores the role of noncoding RNAs, circular RNAs (circRNAs), in BC progression through a competing endogenous RNA (ceRNA) network. Three GEO circRNA microarray datasets (GSE101123, GSE165884, GSE182471) were retrieved, normalized, and batch-corrected using ComBat. Differentially expressed circRNAs (DEcircRNAs) were identified via limma (|log₂FC| > 1, FDR < 0.05). Differentially expressed miRNAs (DEmiRNAs) and mRNAs (DEgenes) were derived from TCGA-BRCA RNA-Seq (1,091 tumors, 113 normals) and miRNA-Seq (1,078 tumors, 104 normals) data using DESeq2 (|log₂FC| > 1, FDR < 0.05). CircRNAs harboring miRNA response elements (MREs) were selected via CSCD, and miRNA-mRNA interactions predicted through TarBase, prioritizing upregulated DEgenes. A ceRNA network was constructed in Cytoscape based on expression concordance. The hsa_circ_0000378/hsa-miR-205-5p/RAD51 axis was validated in 48 paired BC and adjacent non-tumor tissues by RT-qPCR. Results indicated hsa_circ_0000378 upregulation (2.74-fold, p<0.001), hsa-miR-205-5p downregulation (0.64-fold, p=0.0022), and RAD51 upregulation (3.46-fold, p<0.001) in tumors. Spearman correlations showed negative associations between hsa_circ_0000378 and hsa-miR-205-5p (r = -0.474, p<0.001), hsa-miR-205-5p and RAD51 (r = -0.383, p<0.001), and positive between hsa_circ_0000378 and RAD51 (r = 0.497, p<0.001), supporting ceRNA regulation. ROC analysis revealed RAD51's diagnostic potential (AUC=0.83, 95% CI: 0.74-0.90, sensitivity=0.81, specificity=0.55), followed by hsa_circ_0000378 (AUC=0.75, 95% CI: 0.65-0.85, sensitivity=0.71, specificity=0.77), and hsa-miR-205-5p (AUC=0.66, 95% CI: 0.56-0.76, sensitivity=0.69, specificity=0.55). These results propose the hsa_circ_0000378/hsa_miR-205-5p/RAD51 axis as a potential biomarker; mechanistic validation and larger cohorts are needed for clinical application.