[Pan-cancer analysis of ubiquitin-specific protease 7 and its expression changes in the carcinogenesis of scar ulcer].

Objective: To explore the biological role and clinical significance of ubiquitin-specific protease 7 (USP7) in the carcinogenesis of scar ulcer. Methods: A retrospective observational study combined with bioinformatics analysis was used. The RNA expression profile data of USP7 in tumor and/or its corresponding paracancular normal tissue were obtained from The Cancer Genome Atlas (TCGA) database and the Gene Expression Omnibus database, and the RNA sequencing data were transformed by log₂. The variations of USP7 gene were analyzed by cBioPortal database. The USP7 mRNA expression in tumor and adjacent normal tissue in TCGA database were obtained by using the "Gene_DE" module in TIMER 2.0 database. The survival rates of patients with high and low USP7 expression in cutaneous melanoma (SKCM), cervical squamous cell carcinoma (CESC), lung squamous cell carcinoma (LUSC), and head and neck squamous cell carcinoma (HNSC) were analyzed using the Gene Expression Profile Interactive Analysis 2 (GEPIA2) database, and the Kaplan-Meier survival curves were drawn. Sangerbox database was used to analyze the correlation of USP7 expression in pan-cancer with microsatellite instability (MSI) or tumor mutation burden (TMB) pan-cancer. Through the "correlation analysis" module in the GEPIA2 database, the correlation of USP7 expression in pan-cancer with the expression levels of five DNA mismatch repair genes (MLH1, MSH2, MSH6, PMS2, and EPCAM) and three essential DNA methyltransferases (DNMT)--DNMT1, DNMT3A, and DNMT3B were evaluated. The USP7 expression in CESC, HNSC, LUSC, and SKCM and its correlation with infiltration of immune cells (B cells, CD4⁺ T cells, CD8⁺ T cells, neutrophils, macrophages, and dendritic cells) were analyzed by the "Immune-Gene" module in TIMER 2.0 database. The "Similar Genes Detection" module of GEPIA2 database was used to obtain the top 100 protein sets with similar expression patterns to USP7. Intersection analysis was performed between the aforementioned protein sets and the top 50 protein sets that were directly physically bound to USP7 obtained by using the STRING database. Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) enrichment analysis were performed for the two protein sets mentioned above using the DAVID database. The samples of normal skin, hypertrophic scar, scar ulcer, and scar carcinoma with corresponding clinicopathologic features were collected from the Department of Pathology of Tongren Hospital of Wuhan University & Wuhan Third Hospital from October 2018 to October 2022, and the USP7 expression in tissue was detected by immunohistochemical method, with the number of samples of 6. Data were statistically analyzed with Log-rank test, one-way analysis of variance, and Bonferroni test. Results: In pan-cancer, the main gene variations of USP7 were mutation and amplification, and the top 3 tumors with the highest variation frequency (>6%) were bladder urothelial carcinoma, SKCM, and endometrial carcinoma. The main mutation of USP7 gene in pan-cancer was missense mutation. In SKCM with the highest mutation frequency, the main type of mutation was missense mutation in USP7_ICP0_bdg domain. USP7 mRNA expression in breast invasive carcinoma, bile duct carcinoma, colon carcinoma, esophageal carcinoma, HNSC, renal chromophobe cell carcinoma, hepatocellular carcinoma, lung adenocarcinoma, LUSC, prostate carcinoma, and gastric carcinoma was significantly higher than that in corresponding paracancer normal tissue (P<0.05). USP7 mRNA expression in glioblastoma multiforme, renal clear cell carcinoma, renal papillary cell carcinoma, and thyroid carcinoma was significantly lower than that in corresponding paracancular normal tissue (P<0.05). In addition, USP7 mRNA expression in SKCM metastases was much higher than that in primary tumor tissue (P<0.05). Survival curves showed no significant difference in survival rate between patients with high USP7 expression and patients with low USP7 expression in CESC, HNSC, LUSC, and SKCM (Log-rank P>0.05, with hazard ratios of 1.00, 0.99, 1.00, and 1.30, respectively). USP7 expression in colon cancer, colorectal cancer, thymic cancer, and thyroid cancer was negatively correlated with TMB (with Pearson correlation coefficients of -0.26, -0.19, -0.19, and 0.11, respectively, P<0.05). USP7 expression in glioma, CESC, lung adenocarcinoma, mixed renal carcinoma, and LUSC was positively correlated with MSI expression (with Pearson correlation coefficients of 0.22, 0.14, 0.15, 0.08, and 0.14, respectively, P<0.05), and USP7 expression in colon cancer, colorectal cancer, invasive breast cancer, prostate cancer, HNSC, thyroid cancer, and diffuse large B-cell lymphoma were significantly negatively correlated with MSI expression (with Pearson correlation coefficients of -0.31, -0.27, -0.13, -0.19, -0.16, -0.18, and -0.53, respectively, P<0.05). The expression of USP7 in CESC was positively correlated with that of both MSH2 and MSH6 (with Spearman correlation coefficients of 0.51 and 0.44, respectively, P<0.05), and the expression of USP7 in HNSC was positively correlated with the expression of EPCAM, MLH1, MSH2, MSH6, and PMS2 (with Spearman correlation coefficients of 0.39, 0.14, 0.49, 0.54, and 0.41, respectively, P<0.05), and the expression of USP7 in LUSC was positively correlated with the expression of EPCAM, MSH2, MSH6, and PMS2 (with Spearman correlation coefficients of 0.20, 0.36, 0.40, and 0.34, respectively, P<0.05), and the expression of USP7 in SKCM was positively correlated with the expression of EPCAM, MLH1, MSH2, MSH6, and PMS2 (with Spearman correlation coefficients of 0.11, 0.33, 0.42, 0.55, and 0.34, respectively, P<0.05). The expression of USP7 in CESC, HNSC, LUSC, and SKCM was significantly positively correlated with the expression of DNMT1, DNMT3A, and DNMT3B (with Spearman correlation coefficients of 0.42, 0.34, 0.22, 0.45, 0.52, 0.22, 0.36, 0.36, 0.22, 0.38, 0.46, and 0.21, respectively, P<0.05). The expression of USP7 in CESC, HNSC, LUSC, and SKCM was positively correlated with CD4⁺ T cell infiltration (with Partial correlation coefficients of 0.14, 0.22, 0.13, and 0.16, respectively, P<0.05). Being similar to the pattern of USP7 expression and ranked among top 100 protein sets, the top 5 proteins were C16orf72, BCLAF1, UBN, GSPT1, ERI2 (with Spearman correlation coefficients of 0.83, 0.74, 0.73, and 0.72, respectively, all P values<0.05). The top 50 protein sets that directly physically bind to USP7 overlapped with the aforementioned protein set by only one protein, thyroid hormone receptor interaction factor 12. KEGG enrichment analysis showed that USP7 related genes were involved in cell cycle, spliceosome, cell senescence, and p53 signal pathway. GO enrichment analysis showed that USP7 related genes were involved in transcriptional regulation, protein ubiquitination, DNA repair, and cytoplasmic pattern recognition receptor signal pathways. Analysis of clinical samples showed that USP7 expression was significantly higher in hypertrophic scars (0.35±0.05), scar ulcers (0.43±0.04), and scar cancers (0.61±0.03) than in normal skin (0.18±0.04), P<0.05. Conclusions: USP7 may be a clinical biomarker for the progression of cicatricial ulcer cancer.