Progression Of Bladder Urothelial Carcinoma Research Articles

The Oncogenic Potential of IQ Motif-Containing GTPase-Activating Protein3 in Human Bladder Cancer: An In-Silico Analysis

Introduction: IQ motif-containing GTPase-activating protein3 (IQGAP3) contributes to the progression of bladder urothelial carcinoma (BLCA), but its mechanisms are not systematically specified. Due to the oncogenic potential of IQGAP3, the current in-silico study intended to elucidate IQGAP3's role in BLCA progression. Materials and Methods: Many bioinformatics tools, including UALCAN, Kaplan–Meier plotter, TNMplot, cBioPortal, GeneMania, Enrichr, TIMER2, muTarget, and UCSC Xena, were applied in the current study. Results: The IQGAP3 level was more pronouncedly raised in BLCA tissues than in normal bladder tissues, and its increased expression was related to the advanced stage and higher grade. Enhanced IQGAP3 expression could result from its genetic alteration. Moreover, the mutation in P53 and RB1 genes was robustly associated with increased IQGAP3 expression. Besides, IQGAP3 correlative genes were dominantly involved in the cell cycle. On the other hand, IQGAP3 upregulation influenced immune checkpoint levels in the tumor microenvironment. Conclusion: The in-silico findings suggested that IQGAP3 overexpression could be a crucial biomarker in BLCA.

Vitexin Induces Apoptosis and Ferroptosis and Suppresses Malignant Proliferation and Invasion of Bladder Urothelial Carcinoma through PI3K/AKT-Nrf2 Axis

Background: Bladder urothelial carcinoma (BUC) is a type of malignant urinary system. Although several strategies have been applied in the treatment of BUC, its survival remains unsatisfactory, especially in the patients with advanced BUC. Vitexin, a natural flavonoid has exhibited the inhibitory effect on various tumors, however, its effect on BUC is still unclear. Objective: This study aimed to explore the effect of vitexin on the progression of BUC. Methods: The toxicity of vitexin on T24 and 5637 cells was detected by cell counting kit-8 (CCK-8). The effects of vitexin on proliferation, apoptosis, invasion, epithelial-mesenchymal transition (EMT) and ferroptosis in BUC cells were determined by CCK-8, flow cytometry, western blot, transwell and immunofluorescence assays. Additionally, the related mechanism was explored by examining the expression of the phosphatidylinositol-3 kinase (PI3K)/protein kinase B (AKT)-nuclear factor-erythroid 2 related factor 2 (Nrf2) pathway. Besides, in vivo validation was performed in the xenografted mice. Results: Vitexin reduced the BUC cell viability and enhanced the apoptosis rate and the relative protein expression of p53 and cleaved-caspase3. Also, vitexin decreased the invasion number, and increased the relative protein expression of E-cadherin with the decreased N-cadherin protein level in T24 and 5637 cells. Besides, vitexin promoted the levels of ROS and MDA, while reduced the GSH level. Vitexin also increased the level of iron, but decreased the relative protein expression of xCT and GPX4. Erastin further increased the vitexin-induced iron levels, whereas inverse outcomes were observed in the application of ferrostatin-1. Additionally, vitexin decreased the relative protein levels of PI3K, p-AKT/AKT, and nuclear Nrf2, while increased the relative protein level of cytoplasmic Nrf2. Overexpression of PI3K notably inverted the effect of vitexin on cell viability, apoptosis, invasion, level of ROS and iron. Furthermore, vitexin reduced the tumor volume and weight of xenografted mice. Vitexin decreased the protein level of N-cadherin, while increased apoptosis rate of xenografted mice. In addition, vitexin reduced the relative protein levels of PI3K, p-AKT/AKT, and nuclear Nrf2 with the enhanced relative protein expression of cytoplasmic Nrf2 in xenografted mice. Moreover, vitexin decreased the relative protein expression of xCT and GPX4 and the GSH level, whereas increased the MDA level in xenografted mice. Conclusion: Vitexin suppressed malignant proliferation and invasion and induced apoptosis and ferroptosis of BUC involving in PI3K/AKT-Nrf2 pathway.

CircPVT1 Inhibits the Proliferation and Aids in Prediction of the Prognosis of Bladder Cancer.

Circular RNA PVT1 (circPVT1) is aberrantly expressed in several cancers, but its functional role and clinical relevance in bladder urothelial carcinoma (BLCA) remain unknown. This study aimed to identify the expression level of circPVT1 in BLCA and investigated its functional relevance with BLCA progression both in vitro and in vivo. GEPIA, UALCAN, and OncoLnc were referred to presented data. Quantitative real-time PCR (qPCR) was used for the measurement of transnational expression of genes in BLCA specimens and cell lines. Immunohistochemistry (IHC) and fluorescence in situ hybridization analysis (FISH) assays were performedto detect HER2 amplification, Pearson's correlation analysis to analyze the correlation between circPVT1 expression and clinical characteristics, Cox regression and K-M survival analyses to analyze prognostic factors. A nomogram was constructed for predicting prognosis. The proliferation of cells was measured by CCK-8 and colony formation assay, and the proliferation in vivo was evaluated using nude mouse models. qPCR was used to detect the expression of proliferation-related genes. circPVT1 was but mRNA PVT1 was not significantly overexpressed in BLCA. A high circPVT1 expression was associated with a better survival and negative HER2, but not with age, gender, and T stage. circPVT1 was an independent prognostic factor for the overall survival of BLCA patients. Knocking down circPVT1 promoted BLCA proliferation in vitro and in vivo. Knocking down circPVT1 upregulated ERBB2, MKI67, and PCNA expression and downregulated TP53 expression, but exerted no influence on CCND1 and CCNB1 expression. circPVT1 is a tumor suppressor and novel prognostic biomarker for BLCA.

TUBA1C is a potential new prognostic biomarker and promotes bladder urothelial carcinoma progression by regulating the cell cycle

BackgroundTUBA1C is an α-tubulin isoform involved in mitosis, and its dysregulation has been implicated in tumor progression. There is still no clear understanding of its role in bladder urothelial carcinoma (BLCA).MethodsThis study examined the differential expression of TUBA1C and its prognostic significance in bladder cancer based on data from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) and also assessed the correlation of TUBA1C expression level with immune cell infiltration and immune checkpoint gene expression levels and the half-inhibitory concentration (IC50) of different chemotherapeutic agents. Immunotherapy response was estimated using the Tumor Immune Dysfunction and Exclusion (TIDE) algorithm. We detected TUBA1C expression in BLCA cells using PCR and Western blotting. Functional assays, including CCK-8, colony formation, transwell, apoptosis and cell cycle assays, were also performed to assess the oncogenic role of TUBA1C in BLCA.ResultIn three independent public cohorts, TUBA1C was significantly upregulated in bladder tumor tissues, and high TUBA1C expression in bladder cancer was associated with a poorer outcome than low expression. TUBA1C was an independent prognostic risk factor for bladder cancer, and numerous immune checkpoint genes and infiltrating immune cells were associated with TUBA1C. TIDE analysis revealed that TUBA1C showed great potential for predicting the immunotherapy response in bladder cancer patients. In addition, drug sensitivity analysis revealed that high TUBA1C expression indicated sensitivity to multiple chemotherapeutic agents. Functional assays revealed that silencing TUBA1C significantly inhibited the proliferation, migration and invasion of BLCA cells and induced apoptosis and cell cycle arrest.ConclusionThe overexpression of TUBA1C in bladder cancer predicts a poor prognosis and may also be a potential immunotherapeutic target. As a prognostic marker, TUBA1C influences tumor progression by regulating the cell cycle.

Prognostic and immunological role of Asporin across cancers and exploration in bladder cancer

BackgroundAsporin (ASPN) has been identified as a player in tumorigenesis, but its precise roles and modulatory function are largely unknown. Methods: In the present study, ASPN expression was first explored, followed by a prognostic evaluation of ASPN and a comprehensive investigation of the connections between ASPN and immunomodulation, immune cell infiltration and potential compounds on a pancancer level. Finally, ASPN expression was validated in bladder urothelial carcinoma (BLCA) tissues, and the potential function of ASPN, including its effects on migration and invasion capabilities, was investigated in tumor cells. Results: The expression of ASPN exhibited significant variation across cancers and was found to be associated with patient prognosis. In addition, the expression level of APSN was markedly correlated with the abundances of infiltrating immune cells and cancer-associated fibroblasts and the expression levels of immunomodulatory genes based on the results of pancancer analysis. Metastasis and immune-associated signaling pathways were identified in enrichment analysis based on ASPN expression. Finally, we confirmed that ASPN expression increased with the degree of malignancy in BLCA tissues and cell lines and that low expression of ASPN hindered the migration and invasion of cells. Conclusions: ASPN has the potential to be a biomarker of cancer prognosis and a therapeutic target, and it also has predictive capability for the progression of BLCA.

CXCL12, a potential modulator of tumor immune microenvironment (TIME) of bladder cancer: From a comprehensive analysis of TCGA database.

Tumor immune microenvironment (TIME) plays a significant role in the initiation and progression of bladder urothelial carcinoma (BLCA). However, there are only a few researches regarding the association between immune-related genes and tumor-infiltrating immune cells (TICs) in TIME of BLCA. We calculated the proportion of immune/stromal component and TICs of 414 BLCA samples and 19 normal samples downloaded from TCGA database with the help of ESTIMATE and CIBERSORT algorithms. Differentially expressed genes (DEGs) were obtained from the comparison between Stromal and Immune Score and further analyzed by GO and KEGG enrichment analysis, as well as PPI network and COX regression analysis. CXCL12 was overlapping among the above analyses. Single gene analysis of CXCL12 was carried out through difference analysis, paired analysis and GSEA. The association between CXCL12 and TICs was assessed by difference analysis and correlation analysis. Immune and stromal component in TIME of BLCA were associated with patients' clinicopathological characteristics. 284 DEGs were primarily enriched in immune-associated activities, among which CXCL12 was the most significant gene sharing the leading nodes in PPI network and being closely related with patients' survival. Single gene analysis and immunohistochemistry revealed that CXCL12 was down-regulated in BLCA samples and significantly related with the clinicopathological characteristics of patients. Further analysis suggested that CXCL12 was involved in the immune-associated activities probably through its close cross-talk with TICs. CXCL12 down-regulation could be a potential biomarker to predict the unbalanced immune status of TIME of BLCA, which might provide an extra insight for the immunotherapy of BLCA.

KLK6 Functions as an Oncogene and Unfavorable Prognostic Factor in Bladder Urothelial Carcinoma.

Background Kallikrein-related peptidase 6 (KLK6) has been substantiated as a diagnostic, prognostic, and therapeutic molecular in several cancer types. In our study, we attempt to explore the biological functions of KLK6 in bladder urothelial carcinoma (BLCA). Methods KLK6 gene expression prognostic, gene ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), gene set enrichment analysis (GSEA), and immune infiltration were analyzed using The Cancer Genome Atlas (TCGA) database. In vitro and in vivo experimental measurements, including CCK8, transwell migration, TUNEL, and nude mouse transplanted tumor model, were used to evaluate the antineoplastic activities of KLK6 loss-of-function. Results The combination of bioinformatics analyses and experimental measurements demonstrate that KLK6 expression is aberrantly upregulated in human specimens and cell lines of BLCA. GO and GSEA enrichment analyses exhibited that KLK6 is implicated in the inflammatory response and immune infiltration, suggesting that upregulation of KLK6 may be associated with the progression of BLCA. Knockdown of KLK6 is able to inhibit the growth and migration and trigger apoptosis of RT4 and T24 cells. Moreover, the TCGA database indicates that KLK6 high expression in BLCA patients showed a poorer prognosis than those patients with KLK6 low expression. Univariate and multivariate regression analyses suggest KLK6 as an independent prognostic factor to predict unfavorable OS in patients with BLCA. Conclusion KLK6 is an independent prognostic factor and an antitumor target of BLCA. KLK6 expression positively correlates with several immune cells infiltration, indicating that inhibition of KLK6 may contribute to immunotherapy of BLCA.

Clinical Eosinophil-Associated Genes can Serve as a Reliable Predictor of Bladder Urothelial Cancer.

Background: Numerous studies have shown that infiltrating eosinophils play a key role in the tumor progression of bladder urothelial carcinoma (BLCA). However, the roles of eosinophils and associated hub genes in clinical outcomes and immunotherapy are not well known. Methods: BLCA patient data were extracted from the TCGA database. The tumor immune microenvironment (TIME) was revealed by the CIBERSORT algorithm. Candidate modules and hub genes associated with eosinophils were identified by weighted gene co-expression network analysis (WGCNA). The external GEO database was applied to validate the above results. TIME-related genes with prognostic significance were screened by univariate Cox regression analysis, lasso regression, and multivariate Cox regression analysis. The patient’s risk score (RS) was calculated and divided subjects into high-risk group (HRG) and low-risk group (LRG). The nomogram was developed based on the risk signature. Models were validated via receiver operating characteristic (ROC) curves and calibration curves. Differences between HRG and LRG in clinical features and tumor mutational burden (TMB) were compared. The Immune Phenomenon Score (IPS) was calculated to estimate the immunotherapeutic significance of RS. Half-maximal inhibitory concentrations (IC50s) of chemotherapeutic drugs were predicted by the pRRophetic algorithm. Results: 313 eosinophil-related genes were identified by WGCNA. Subsequently, a risk signature containing 9 eosinophil-related genes (AGXT, B3GALT2, CCDC62, CLEC1B, CLEC2D, CYP19A1, DNM3, SLC5A9, SLC26A8) was finally developed via multiplex analysis and screening. Age (p < 0.001), grade (p < 0.001), and RS (p < 0.001) were independent predictors of survival in BLCA patients. Based on the calibration curve, our risk signature nomogram was confirmed as a good predictor of BLCA patients’ prognosis at 1, 3, and 5 years. The association analysis of RS and immunotherapy indicated that low-risk patients were more credible for novel immune checkpoint inhibitors (ICI) immunotherapy. The chemotherapeutic drug model suggests that RS has an effect on the drug sensitivity of patients. Conclusions: In conclusion, the eosinophil-based RS can be used as a reliable clinical predictor and provide insights into the precise treatment of BLCA.

Downregulation of PTCD1 in Bladder Urothelial Carcinoma Predicts Poor Prognosis and Levels of Immune Infiltration.

Pentatricopeptide repeat domain 1 (PTCD1) was reported to regulate mitochondrial metabolism and oxidative phosphorylation. However, the effect and mechanism of PTCD1 in the development of bladder urothelial carcinoma (BLCA) remain unclear. The databases from The Cancer Genome Atlas (TCGA) and Human Protein Atlas (HPA) were used to analyze the expression changes, clinical features, and prognostic values of PTCD1. A nomogram was built to predict the prognostic outcomes of BLCA cases. The potential genes interacting with PTCD1 were explored by Weighted Gene Coexpression Network Analysis (WGCNA). The estimation of associations between PTCD1 and tumor mutations, tumor immunities, and m6A methylations was performed. The study found that the gradual decrease of PTCD1 expression was observed with the increase of stage and grade. Low PTCD1 expression was greatly correlated with higher pathological stage, N stage, and poor prognosis in TCGA cohorts; interestingly, low-grade BLCA cases all exhibited high expression of PTCD1. HPA database analysis implied that the expression of PTCD1 protein in BLCA was lower than that in normal bladder tissue, and the protein expression of PTCD1 in high-grade BLCA was lower than that in low-grade BLCA. Multivariate Cox regression analysis indicated that PTCD1 may serve as an independent factor influencing prognosis of BLCA. Mechanistically, PTCD1 played a regulatory role in BLCA progression through multiple tumor-related pathways containing PI3K-Akt signaling, ECM-receptor interaction, oxidative phosphorylation, and extracellular matrix organization. WGCNA reported that PTCD1 had a strong positive correlation with POLR2J, ZNHT1, ATP5MF, PDAP1, BUD31, and COPS6. Besides, the mRNA expression of PTCD1 was negatively associated with immune cells' infiltrations, immune functions, and checkpoints, especially with some m6A methylation regulators in BLCA. In sum, downregulation of PTCD1 expression may be involved in the development of BLCA and remarkably correlated with poor prognosis. Meantime, it showed an influence in immune cell infiltration and may serve as an agreeable prognostic indicator in BLCA.

ANLN Serves as an Oncogene in Bladder Urothelial Carcinoma via Activating JNK Signaling Pathway

Introduction: To understand the significance of ANLN (anillin, actin-binding protein)-mediated c-Jun N-terminal kinase (JNK) signal pathway on the progression of bladder urothelial carcinoma (BLCA). Methods: The Cancer Genome Atlas (TCGA) database was utilized to perform the clinical significance of ANLN in BLCA. Then, ANLN expression was determined in human normal primary bladder epithelial cells (BdEC) and BLCA cells. Later, ANLN knockdown was performed in BLCA cells, where the expression of MAPK8, MAPK9, and p-JNK/JNK was detected. BLCA cells were divided into the Mock, siNC, siANLN, SP600125 (a selective JNK inhibitor), and ANLN + SP600125 group, followed by measurements of real-time quantitative polymerase chain reaction, 3-4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide, Annexin V-FITC/PI, Wound-healing, Transwell, and immunofluorescence assays. Results: ANLN was upregulated in the BLCA tissues, which showed a relation with the stage of patients. Besides, BLCA patients with high expression of ANLN had a worse prognosis than those with low expression of ANLN. Besides, the expression of ANLN in the BLCA tissues was positively correlated with MAPK8 and MAPK9. SP600125 suppressed the JNK signal pathway, reduced the proliferation, and increased BLCA cell apoptosis, with the reductions in the invasion and migration and the upregulation of phospho-histone H3 Ser-10 (pHH3), which was abolished by the overexpression of ANLN. Conclusion: ANLN, as an oncogene of BLCA, may associate with the activation of JNK signal pathway. Inhibiting ANLN could deactivate the JNK signal pathway, thereby suppressing the proliferation, invasion, and migration while promoting the apoptosis of BLCA cells.

PGM1 and ENO1 Promote the Malignant Progression of Bladder Cancer via Comprehensive Analysis of the m6A Signature and Tumor Immune Infiltration.

Background While N6-methyladenosine (m6A) modification of RNA and the tumor immune microenvironment both influence the progression of cancer, little attention has been paid to interactions between these two factors. Thus, we systematically explored potential biomarkers in the malignant progression of bladder urothelial carcinoma (BLCA) via combining expression of m6A methylation regulators with tumor immune infiltration. Methods We extracted m6A regulators from published literature, downloaded BLCA RNA-seq and clinical information from the Cancer Genome Atlas database, and integrated three main bioinformatic methods and qPCR to explore the biological variations in the malignant progression of BLCA. Results FTO, IGF2BP3, and YTHDC1 have a significant difference in bladder cancer and prognosis. Two subgroups (clusters 1 and 2) were identified according to three key m6A regulators; cluster 1 was preferentially associated with poor prognosis and immune infiltration relative to cluster 2 significantly. We further identified PGM1 and ENO1 as potential prognostic biomarkers, as they were correlated with FTO and IGF2BP3 positively but with YTHDC1, negatively. M2 macrophage and TFH cells were highly infiltrated in BLCA and were associated with BLCA prognosis. Finally, PGM1 and ENO1 were correlated with M2 macrophage and TFH cells and their surface markers CD163and CXCR5. Conclusions PGM1 and ENO1 are highly correlated with the malignant progression of BLCA, and the expression of these genes may be new indicators for the diagnosis and prognosis of BLCA.

FAM83H and Nectin1 expression are related with survival and relapse of bladder urothelial carcinoma patients

BackgroundFAM83H was originally reported to be essential for dental enamel formation. However, FAM83H has recently been implicated in tumorigenesis and tumor progression. Analysis of a publicly available gene expression database revealed a significant correlation between FAM83H and Nectin1 mRNA expression and bladder urothelial carcinoma (BUC). Therefore, we investigated the association between FAM83H and Nectin1 expression levels and the survival and recurrence of BUC in BUC patients using a tissue microarray.MethodsWe performed immunohistochemical staining of FAM83H and Nectin1 in 165 human BUC tissue sections, and analyzed the prognostic significance of FAM83H and Nectin1 expression.ResultsBoth FAM83H and Nectin1 were mainly expressed in the cytoplasm, and their expression was significantly associated. FAM83H expression was significantly correlated with higher histologic grade, higher T stage, higher TNM stage, and recurrence. Nectin1 expression was significantly associated with higher histologic grade and recurrence. Univariate analysis showed FAM83H expression and Nectin1 expression were significantly associated with worse overall survival (OS) and shorter relapse-free survival (RFS) of BUC patients. In multivariate analysis, levels of FAM83H and Nectin1 were independent indicators of shorter survival of BUC patients.ConclusionsOur results suggest that FAM83H and Nectin1 are important in the progression of BUC, and that expression patterns of these two proteins can be used as prognostic indicators of survival in BUC patients.

Role of NRP1 in Bladder Cancer Pathogenesis and Progression.

Bladder urothelial carcinoma (BC) is a fatal invasive malignancy and the most common malignancy of the urinary system. In the current study, we investigated the function and mechanisms of Neuropilin-1 (NRP1), the co-receptor for vascular endothelial growth factor, in BC pathogenesis and progression. The expression of NRP1 was evaluated using data extracted from GEO and HPA databases and examined in BC cell lines. The effect on proliferation, apoptosis, angiogenesis, migration, and invasion of BC cells were validated after NRP1 knockdown. After identifying differentially expressed genes (DEGs) induced by NRP1 silencing, GO/KEGG and IPA® bioinformatics analyses were performed and specific predicted pathways and targets were confirmed in vitro. Additionally, the co-expressed genes and ceRNA network were predicted using data downloaded from CCLE and TCGA databases, respectively. High expression of NRP1 was observed in BC tissues and cells. NRP1 knockdown promoted apoptosis and suppressed proliferation, angiogenesis, migration, and invasion of BC cells. Additionally, after NRP1 silencing the activity of MAPK signaling and molecular mechanisms of cancer pathways were predicted by KEGG and IPA® pathway analysis and validated using western blot in BC cells. NRP1 knockdown also affected various biological functions, including antiviral response, immune response, cell cycle, proliferation and migration of cells, and neovascularisation. Furthermore, the main upstream molecule of the DEGs induced by NRP1 knockdown may be NUPR1, and NRP1 was also the downstream target of NUPR1 and essential for regulation of FOXP3 expression to activate neovascularisation. DCBLD2 was positively regulated by NRP1, and PPAR signaling was significantly associated with low NRP1 expression. We also found that NRP1 was a predicted target of miR-204, miR-143, miR-145, and miR-195 in BC development. Our data provide evidence for the biological function and molecular aetiology of NRP1 in BC and for the first time demonstrated an association between NRP1 and NUPR1, FOXP3, and DCBLD2. Specifically, downregulation of NRP1 contributes to BC progression, which is associated with activation of MAPK signaling and molecular mechanisms involved in cancer pathways. Therefore, NRP1 may serve as a target for new therapeutic strategies to treat BC and other cancers.

MiR-25 regulates cell proliferation and metastasis in bladder urothelial carcinoma.

Background: Bladder urothelial carcinoma (BC) is a common malignant tumor with a high incidence. This study aims to explore the role of miR-25 in BC tumorigenesis.Material and Methods: The expression of miR-25 and PTEN were detected in clinical BC tissues. BC cell lines T24 and 5637 were used to transfect miR-25 mimics or inhibitors. Luciferase reporter gene detection confirmed the correlation between miR-25 and PTEN. CCK-8 method and flow cytometry were used to detect cell viability and apoptosis. Cell migration and invasion ability were examined by transwell assays. Western blotting detects the protein levels of PTEN, β-catenin, GSK-3β and p-GSK-3β.Results: MiR-25 and PTEN expression are found to be negatively correlated in BC tissues. Further research confirmed that PTEN is a direct target of miR-25. In addition, the overexpression of miR-25 down-regulates the expression of PTEN, induces cell survival and inhibits apoptosis, while the knockout of miR-25 leads to the opposite result. miR-25 also inhibits the phosphorylation of GSK-3β and β-catenin without changing the total level of GSK-3β. In vivo experiments confirmed that miR-25 plays an oncogene's role by regulating the PTEN and Wnt/β-catenin signaling pathways.Conclusion: Our research shows that miR-25 has a negative regulatory effect on the expression of PTEN in clinical specimens and in vitro. miR-25 can promote the proliferation of BC cells and induce cell invasion. Therefore, miR-25 may be used as a biomarker to predict the progression of BC.

MAFG-AS1 promotes tumor progression via regulation of the HuR/PTBP1 axis in bladder urothelial carcinoma.

Long noncoding RNAs (lncRNAs) play a crucial role in progression of bladder urothelial carcinoma (BUC). However, the molecular mechanisms behind this role have not been elucidated yet. Here, we found that the lncRNA MAFG‐AS1, which is highly expressed in BUC, is correlated with aggressive characteristics and poor prognosis of BUC. We demonstrate that MAFG‐AS1 can promote BUC proliferation, invasion, metastasis, and epithelial‐mesenchymal transition in vitro and in vivo. Mechanistically, MAFG‐AS1 direct binding to Hu antigen R (HuR) could recruit ubiquitin‐specific proteinase 5 (USP5) to prevent HuR from degrading by ubiquitination. We further demonstrate that overexpression of MAFG‐AS1 can upregulate the expression of polypyrimidine tract‐binding protein 1 (PTBP1) through promoting its stability mediated by bound HuR. In conclusion, these findings indicate that MAFG‐AS1 promotes the progression of BUC via regulation of the HUR/PTBP1 axis. Targeting MAFG‐AS1 may provide a novel strategy for individualized therapy and a potential biomarker for prognosis of BUC.

Weighted gene co-expression network analysis can sort cancer-associated fibroblast-specific markers promoting bladder cancer progression.

The role of cancer-associated fibroblasts (CAFs) has been thoroughly investigated in tumour microenvironments but not in bladder urothelial carcinoma (BLCA). The cell fraction of CAFs gradually increased with BLCA progression. Weighted gene co-expression network analysis (WGCNA) revealed a specific gene expression module of CAFs that are relevant to cancer progression and survival status. Fifteen key genes of the module were consistent with a fibroblast signature in single-cell RNA sequencing, functionally related to the extracellular matrix, and significant in survival analysis and tumour staging. A comparison of the luminal-infiltrated versus luminal-papillary subtypes and fibroblast versus urothelial carcinoma cell lines and immunohistochemical data analysis demonstrated that the key genes were specifically expressed in CAFs. Moreover, these genes are highly correlated with previously reported CAF markers. In summary, CAFs play a major role in the progression of BLCA, and the 15 key genes act as BLCA-specific CAF markers and can predict CAF changes. WGCNA can, therefore, be used to sort CAF-specific gene set in cancer tissues.

SLC12A5 interacts and enhances SOX18 activity to promote bladder urothelial carcinoma progression via upregulating MMP7.

Solute carrier family 12 member 5 (SLC12A5) has an oncogenic role in bladder urothelial carcinoma. The present study aimed to characterize the molecular mechanisms of SLC12A5 in bladder urothelial carcinoma pathogenesis. Functional assays identified that in bladder urothelial carcinoma SLC12A5 interacts with and stabilizes SOX18, and then upregulates matrix metalloproteinase 7 (MMP7). In vivo and in vitro assays were performed to confirm the effect of SLC12A5’s interaction with SOX18 on MMP7‐mediated bladder urothelial carcinoma progression. SLC12A5 was upregulated in human bladder tumors, and correlated with the poor survival of patients with bladder urothelial carcinoma tumor invasion and metastasis, promoted by SLC12A5 overexpression. We demonstrated that SLC12A5 interacted with SOX18, and then upregulated MMP7, thus enhancing tumor progression. Importantly, SLC12A5 expression correlated positively with SOX18 and MMP7 expression in bladder urothelial carcinoma. Furthermore, SLC12A5 expression was suppressed by miR‐133a‐3p. Ectopic expression of SLC12A5 partly abolished miR‐133a‐3p‐mediated suppression of cell migration. SLC12A5‐SOX18 complex‐mediated upregulation on MMP7 was important in bladder urothelial carcinoma progression. The miR‐133a‐3p/SLC12A5/SOX18/MMP7 signaling axis was critical for progression, and provided an effective therapeutic approach against bladder urothelial carcinoma.

Screening and identification of potential prognostic biomarkers in bladder urothelial carcinoma: Evidence from bioinformatics analysis

Bladder urothelial carcinoma (BLCA) is a very much invasive urological malignant tumor that results in reduced patient survival. At current, the mechanism of BLCA metastasis is not clear. This study was aimed to explore the prognostic and diagnostic related biomarkers in BLCA and guide the therapy. First, we acquired data of gene expression in non-muscle invasive bladder cancer samples (N = 15) and muscle invasive bladder cancer samples (N = 15) from the Gene Expression Omnibus (GEO) database. Integrated analysis was used to explore differentially expressed genes (DEGs) (up and down regulated genes) between non invasive BLCA samples and invasive BLCA samples. Pathway enrichment analyses and Gene ontology (GO) of DEGs were performed. DEGs were used to construct a protein–protein interaction (PPI) network using Protein Interaction Network Analysis platform (PINA) database, and the modules from PPI network were verified by Cytoscape software and hub genes were extracted. Furthermore, gene regulatory networks were depicted based on miRNAs and TFs with interest target genes. The survival analysis, expression analysis, stage analysis, mutation analysis, immunohistochemical (IHC) analysis, ROC analysis and RT-PCR for the integrated expression score was applied to determine the diagnostic ability of the candidate biomarkers. In total, 900 DEGs that consisted of 455 up regulated and 445 down regulated genes were screened out. The differentially expressed genes were enriched in stearate biosynthesis, metabolic pathways, acetate conversion to acetyl-CoA, focal adhesion, oxoacid metabolic process, mitochondrial matrix, circulatory system development and extracellular matrix, of which pathways and GO terms were associated in the BLCA development. PPI network analysis, demonstrated the interactions between those DEGs, and 10 hub genes (P4HB, APEX1, SIRT5, SRP68, HIP1R, MYC, A2M, MAP1B, HIPK2 and TCF) were extracted from the network and verified by survival analysis, expression analysis, stage analysis, mutation analysis, immunohistochemical (IHC) analysis, ROC analysis, RT-PCR and immune infiltration analysis.. In this study, we highlighted mechanisms underlying the control of BLCA molecular signatures by TFs and miRNAs which their alteration reduce the chance of survival rate in BLCA. These results revealed that the DEGs may serve as candidate essential genes during BLCA metastasis. The 10 hub genes, including P4HB, APEX1, SIRT5, SRP68, HIP1R, MYC, A2M, MAP1B, HIPK2 and TCF, may serve as promising prognostic biomarkers in BLCA metastasis. In conclusion, the present study identifies DEGs and hub genes that may be involved in poor prognosis and early recurrence of BLCA. The expression levels of P4HB, APEX1, SIRT5, SRP68, HIP1R, MYC, A2M, MAP1B, HIPK2 and TCF are of high prognostic value, and may help us understand better the underlying carcinogenesis or progression of BLCA. Further studies are required to elucidate molecular pathogenesis and alteration in signaling pathways for these genes in BLCA.

Weighted Gene Co-Expression Network Analysis Can Sort Cancer-Associated Fibroblast-Specific Markers Promoting Bladder Cancer Progression

Background: Cancer-associated fibroblasts (CAFs) play an important role in cancer progression, but their roles in bladder urothelial carcinoma (BLCA) have not been thoroughly investigated. Methods: Estimate the Proportion of Immune and Cancer cells (EPIC) application was designed to calculate cell fractions in tissues. Weighted gene co-expression network analysis (WGCNA), by combining calculation principles, was used to identify genes expressed in specific cells in a tumour microenvironment (TME). Functional enrichment analysis and external data validation were conducted to assess the relationship between key genes and CAFs. Findings: Cell fraction of CAFs gradually increases with the progression of BLCA. Network construction revealed a specific gene module of CAFs that is the most relevant to cancer progression and survival status. Fifteen key genes of the CAF module were functionally related to the extracellular matrix and were significant in the analysis of survival and differentiation of tumour staging. A comparison of luminal-infiltrated subtypes with luminal-papillary subtypes and fibroblast cell lines with urothelial carcinoma cell lines, and the analysis of immunohistochemical data demonstrated that the key genes were specifically expressed in CAFs. Importantly, the key genes were highly correlated with some previously reported CAF markers. Interpretation: CAFs play a major role in the progression of BLCA. BLCA-specific CAF markers, the 15 key genes, can predict the changes in CAFs. The WGCNA can sort cell-specific gene sets in cancer tissues. Funding Statement: 345 Talent project - Shengjing Hospital of China Medical University. Declaration of Interests: The authors declare no conflict of interest. Ethics Approval Statement: Not required.

Enhanced CDR1as Promotes the Development of Bladder Urothelial Carcinoma.

The current study aimed to evaluate the expression of circular RNA (circRNA) CDR1as in bladder urothelial carcinoma and adjacent normal tissues, thereby laying the foundation for the research of bladder urothelial carcinoma. The level of CDR1as was evaluated in the tissues of bladder urothelial carcinoma patients. ROC analysis was carried out to explore the possible diagnostic value of CDR1as. In the current study, we showed novel data that the expression of CDR1as in bladder urothelial carcinoma tissues was significantly higher than that in adjacent tissues. Furthermore, the expression of CDR1as was significantly higher in non-muscle invasive bladder cancer (NMIBC) and high-grade cancer tissues than in muscle invasive bladder cancer (MIBC) and low-grade cancer tissues. In patients with lymph node metastasis, the expres-sion of CDR1as was also significantly higher. Moreover, the expression of CDR1as was higher in high-stage, high-grade bladder urothelial carcinoma than in low-stage and low-grade cancer tissues. Real time PCR analysis showed that there was no difference in the expression of CDR1as among the patients according to the grouping of gender (male vs. female), age (≤ 65 years vs. > 65 years), tumor size (≤ 3 cm vs. > 3 cm), and lymphatic invasion (LVI). The current study for the first time showed upregulation of CDR1as may be involved in the development and progression of bladder urothelial carcinoma.