Expression of collagen type IV alpha 4 chain in clear cell renal cell carcinoma and its relationship with prognosis and immune infiltration

Objective To explore the expression of collagen type Ⅳ alpha 4 chain (COL4A4) in clear cell renal cell carcinoma (ccRCC) and its relationship with prognosis and immune infiltration. Methods The differential expression of COL4A4 in ccRCC and adjacent normal tissues was analyzed using The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases. Western blot was used to examine the difference of COL4A4A expression in different human ccRCC cell lines including 786-O, OS-RC-2, Caki-1, SW839, and RCC4, and human normal renal epithelial cell line HK-2. Kaplan-Meier method was used to analyze the correlation between the expression of COL4A4 and the overall survival (OS), progression-free survival (PFS), and disease-specific survival of the patients. Receiver operating characteristic (ROC) curve was used to evaluate the diagnostic value of COL4A4 expression for ccRCC. The gene mutation landscape of COL4A4 was analyzed by the cBioPortal and COSMIC databases. The protein-protein interaction (PPI) network of COL4A4 was conducted by STRING. The co-expression network of COL4A4 in ccRCC was analyzed by the LinkedOmics database and used to perform the Gene Ontology (GO) annotations and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis by R software. The single sample gene set enrichment analysis (ssGSEA) was used to analyze the correlation of COL4A4 expression and immune infiltration in ccRCC based on TCGA database. Results The analysis in TCGA and GEO databases showed that COL4A4 expression was lower in ccRCC tissues (both P<0.05). Western blot confirmed the low expression of COL4A4 in all ccRCC cell lines (all P<0.05). Patients with high COL4A4 expression had better OS, PFS and disease-specific survival (all P<0.01). COL4A4 expression was an independent risk factor for ccRCC (HR=0.562, 95% CI: 0.352-0.898, P<0.05). The analysis in the COSMIC database found that COL4A4 had a low mutation frequency and was stably expressed in ccRCC. The PPI network results showed that COL4A4 was closely related to the integrin family. The GO annotations showed that COL4A4 was mainly involved in the biological processes of the regulation of GTPase activity, nuclear division, the regulation of cell cycle phase transition, autophagy, and T cell activation. The KEGG pathway enrichment analysis showed that COL4A4 mainly participated in the salmonella infection and tight junction pathways. The ssGSEA results showed that COL4A4 expression was neg-atively associated with regulatory T cell infiltration (r=-0.428, P<0.05). Conclusions COL4A4 may serve as a novel prognostic marker and a potential therapeutic target for ccRCC.