ANO4 Expression Is a Potential Prognostic Biomarker in Non-Metastasized Clear Cell Renal Cell Carcinoma

Background: Over the past decade, transcriptome profiling has elucidated many pivotal pathways involved in oncogenesis. However, a detailed comprehensive map of tumorigenesis remains an enigma to solve. Propelled research has been devoted to investigating the molecular drivers of clear cell renal cell carcinoma (ccRCC). To add another piece to the puzzle, we evaluated the role of anoctamin 4 (ANO4) expression as a potential prognostic biomarker in non-metastasized ccRCC. Methods: A total of 422 ccRCC patients with the corresponding ANO4 expression and clinicopathological data were obtained from The Cancer Genome Atlas Program (TCGA). Differential expression across several clinicopathological variables was performed. The Kaplan-Meier method was used to assess the impact of ANO4 expression on the overall survival (OS), progression-free interval (PFI), disease-free interval (DFI), and disease-specific survival (DSS). Univariate and multivariate Cox logistic regression analyses were conducted to identify independent factors modulating the aforementioned outcomes. Gene set enrichment analysis (GSEA) was used to discern a set of molecular mechanisms involved in the prognostic signature. Tumor immune microenvironment was estimated using xCell. Results: ANO4 expression was upregulated in tumor samples compared to normal kidney tissue. Albeit the latter finding, low ANO4 expression is associated with advanced clinicopathological variables such as tumor grade, stage, and pT. In addition, low ANO4 expression is linked to shorter OS, PFI, and DSS. Multivariate Cox logistic regression analysis identified ANO4 expression as an independent prognostic variable in OS (HR: 1.686, 95% CI: 1.120-2.540, p = 0.012), PFI (HR: 1.727, 95% CI: 1.103-2.704, p = 0.017), and DSS (HR: 2.688, 95% CI: 1.465-4.934, p = 0.001). GSEA identified the following pathways to be enriched within the low ANO4 expression group: epithelial-mesenchymal transition, G2-M checkpoint, E2F targets, estrogen response, apical junction, glycolysis, hypoxia, coagulation, KRAS, complement, p53, myogenesis, and TNF-α signaling via NF-κB pathways. ANO4 expression correlates significantly with monocyte (ρ = -0.1429, p = 0.0033) and mast cell (ρ = 0.1598, p = 0.001) infiltration. Conclusions: In the presented work, low ANO4 expression is portrayed as a potential poor prognostic factor in non-metastasized ccRCC. Further experimental studies should be directed to shed new light on the exact molecular mechanisms involved.