Comprehensive Analysis of Genomic and Phenomic Data Reveals Context-Dependent Function of A20 (TNFAIP3) in Renal Cell Carcinoma

Background: A20, encoded by tumor necrosis factor alpha-induced protein 3 (TNFAIP3), is a key negative regulator of NF-κB signaling with context-dependent functions in cancer. Its role in renal cell carcinoma (RCC), particularly clear-cell RCC (ccRCC), remains incompletely defined. Methods: Public GEO transcriptomic data from 23 controls and 32 ccRCC samples were analyzed using gene set enrichment analysis (GSEA). A20-overexpressing HEK293 and 786-O cells were assessed by functional assays and transcriptomic profiling. Eight ccRCC FFPE samples were profiled to compare A20-associated transcriptional patterns with cell-line data. Targeted DNA sequencing was performed in 11 ccRCC and 8 papillary RCC samples, and whole-exome sequencing was conducted in A20-overexpressing 786-O cells. Key genes were further evaluated using Kaplan-Meier survival analysis in 530 ccRCC patients and TCGA-KIRC data comprising 533 primary tumors and 72 normal samples. Results: GEO analysis showed significant TNFAIP3/A20 upregulation in ccRCC (p = 3.96 × 10−5) and enrichment of NF-κB-related gene sets (p = 0.01). A20 overexpression produced distinct phenotypes in HEK293 and 786-O cells. In HEK293 cells, A20 increased BIK and ARHGAP6 expression and was associated with increased apoptosis and reduced wound closure. In 786-O cells, A20 suppressed ARHGAP6 and APAF1 and was associated with increased proliferation, enhanced wound closure, and reduced apoptosis relative to EV controls. A20-high ccRCC samples showed enrichment of NF-κB, TGF-β, DNA repair, mTOR/metabolic, hypoxia, and proteasome-related pathways. Genomic analyses identified alterations in NF-κB-related genes, including CARD10 and IRAK1. Conclusions: A20/TNFAIP3 may exert cell-context-dependent effects in RCC and is associated with tumor-relevant transcriptional and genomic alterations requiring further validation.