Abstract: Objective To investigate the multi⁃omics characteristics of BEN domain containing 3(BEND3 ) in breast cancer and its effect on tumor cell proliferation, migration, and invasion. Methods BEND3 mRNA and protein expression were analyzed for prognostic value using the TCGA, GTEx, and GEO, UALCAN and HPA database, A total of 48 paired breast cancer and adjacent tissues samples from The First Affiliated Hospital of Xinjiang Medical University (August 2023 to July 2024) were collected for immunohistochemistry. Prognostic significance was assessed with univariable and multivariable Cox regression and Kaplan⁃Meier analyses, validated by PrognoScan and Kaplan⁃Meier Plotter. Genetic mutations, copy number variations, and methylation status were examined using the cBioPortal and GSCA. Immune cell infiltration was assessed via TIMER2.0, and a protein⁃protein interaction (PPI) network was constructed using STRING, followed by GO, KEGG, and GSEA enrichment analyses. BEND3 knockdown and overexpression in MCF⁃7 and MDA⁃MB⁃231 cell lines were verified by qRT⁃PCR and Western blot. Subsequently, cell proliferation, migration, and invasion were evaluated using CCK⁃8 assay, clolny formation assay wound healing assay, and Transwell assays, respectively. Results BEND3 mRNA and protein levels were significantly elevated in breast cancer (P<0.001), particularly in the Basal and triple⁃negative subtypes (all P<0.001). It exhibited moderate diagnostic value for breast cancer (AUC=0.820) and was linked to poor  overall survival, disease⁃specific survival and progression⁃free interval (all P<0.05), serving as an independent prognostic risk factor for breast cancer (P<0.05). BEND3 expression was influenced by gene amplification, copy number variation, and DNA and RNA methylation, and was correlated with immune cell infiltration, tumor mutational burden and neoantigen load. Functional enrichment analysis revealed that it was involved in cell cycle and Notch signaling, as well as metabolic reprogramming. In vitro, BEND3 overexpression enhanced breast cancer cell proliferation, migration, and invasion, while knockdown inhibited these effects (all P<0.05). Conclusions BEND3 is markedly overexpressed in  breast cancer and is modulated by genetic and epigenetic alterations, associated with poor prognosis and the tumor immune microenvironment. The elevated expression of BEND3 facilitates the proliferation, migration, and invasion in  breast cancer cell.

Key words: Breast cancer, BEN domain containing 3, Multi?omics, Prognosis, Proliferation, Metastasis