Integrative multi-omics analysis identifies FAM3D as a prognostic tumor suppressor in cervical cancer

Cervical cancer (CC) remains a leading malignancy among women worldwide. Epigenetic and transcriptional dysregulation complicate the identification of causal genes with prognostic and therapeutic relevance. We performed a multi-omics integrative analysis combining summary data-based Mendelian randomization (SMR) using eQTL and pQTL datasets with genome-wide association study (GWAS) summary statistics (ID: ukb-b-8777) to prioritize candidate genes associated with CC. Differential expression analyses were conducted using multiple GEO cohorts, and protein expression was validated by immunohistochemistry (IHC) in clinical specimens. DNA methylation profiling and correlation analyses were performed using GEO and TCGA datasets to investigate epigenetic regulation. Kaplan-Meier and combined expression-methylation survival analyses were used to evaluate prognostic significance. Functional validation was conducted in HeLa cells using wound-healing and CCK-8 assays following FAM3D knockdown or overexpression. RNA sequencing was further performed to explore the downstream molecular pathways regulated by FAM3D. Integrative SMR analysis identified FAM3D as a protective gene for CC. FAM3D expression was significantly reduced in CC tissues and exhibited a progressive decline from normal cervical tissues to cervical intraepithelial neoplasia (CIN) and invasive CC, which was further confirmed by IHC. Promoter hypermethylation of FAM3D (cg26334888) was associated with its transcriptional downregulation and negatively correlated with gene expression. Survival analyses demonstrated that high FAM3D expression (p = 0.016) and low promoter methylation (p = 0.048) were associated with favorable overall survival, while combined expression-methylation analysis further improved prognostic stratification (p = 0.006). Functional assays showed that FAM3D suppressed CC cell proliferation and migration. Transcriptomic profiling revealed that FAM3D restoration induced extensive transcriptional reprogramming and was associated with inflammatory signaling, cytokine-mediated pathways, and extracellular matrix remodeling. Key hub genes identified downstream of FAM3D included CXCL8, MMP1, EREG, and LCN2. FAM3D is a potential tumor suppressor and prognostic biomarker in CC, whose expression is partially regulated by promoter methylation. Integrative multi-omics analyses and functional studies suggest that FAM3D may inhibit CC progression through modulation of inflammatory signaling and extracellular matrix-associated pathways. These findings provide new insights into the molecular mechanisms underlying CC and support the potential clinical utility of FAM3D as a prognostic biomarker and therapeutic target.