Identification of autophagy-related genes signature predicts chemotherapeutic and immunotherapeutic efficiency in bladder cancer (BLCA).

Autophagy maintains cellular homeostasis by degrading and recycling cytoplasmic components under stress conditions, which is identified to be involved in tumorigenesis and now has been recognized as novel target in cancer treatment. In present study, we gathered total autophagy-related genes and established an autophagy-related genes signature (ATGRS) through LASSO cox regression analysis in BLCA. Kaplan-Meier survival and multivariate cox regression analyses both showed the ATGRS was a robust independent prognostic factor with high accuracy. Subsequently, integrated analyses indicated that ATGRS had a strong correlation with molecular subtypes, clinicopathological characteristics and somatic mutation alteration. Moreover, ATGRS was found to be positively correlated with the infiltration of immune cells in tumour microenvironment (TME) and immune checkpoint expression, indicating the potent role of autophagy by regulating the TME. In addition, ATGRS was proved to be efficient in predicting the clinical benefit of immune checkpoint inhibitors (ICIs) based immunotherapy and chemotherapy in BLCA. Furthermore, we observed abnormal expression levels of autophagy-related genes and found the different behaviour of ATGRS in pancancer by LASSO cox regression analysis. Therefore, construction of ATGRS in BLCA could help us to interpret the underlying mechanism of autophagy and sheds a light on the clinical application for a combination of autophagy modification with targeted immunotherapy and chemotherapy in BLCA.

Identification of a prognostic gene signature based on an immunogenomic landscape analysis of bladder cancer.

Cancer immune plays a critical role in cancer progression. Tumour immunology and immunotherapy are one of the exciting areas in bladder cancer research. In this study, we aimed to develop an immune-related gene signature to improve the prognostic prediction of bladder cancer. Firstly, we identified 392 differentially expressed immune-related genes (IRGs) based on TCGA and ImmPort databases. Functional enrichment analysis revealed that these genes were enriched in inflammatory and immune-related pathways, including in 'regulation of signaling receptor activity', 'cytokine-cytokine receptor interaction' and 'GPCR ligand binding'. Then, we separated all samples in TCGA data set into the training cohort and the testing cohort in a ratio of 3:1 randomly. Data set GSE13507 was set as the validation cohort. We constructed a prognostic six-IRG signature with LASSO Cox regression in the training cohort, including AHNAK, OAS1, APOBEC3H, SCG2, CTSE and KIR2DS4. Six IRGs reflected the microenvironment of bladder cancer, especially immune cell infiltration. The prognostic value of six-IRG signature was further validated in the testing cohort and the validation cohort. The results of multivariable Cox regression and subgroup analysis revealed that six-IRG signature was a clinically independent prognostic factor for bladder cancer patients. Further, we constructed a nomogram based on six-IRG signature and other clinicopathological risk factors, and it performed well in predict patients' survival. Finally, we found six-IRG signature showed significant difference in different molecular subtypes of bladder cancer. In conclusions, our research provided a novel immune-related gene signature to estimate prognosis for patients' survival with bladder cancer.

An outcome model for human bladder cancer: A comprehensive study based on weighted gene co-expression network analysis.

The precision evaluation of prognosis is crucial for clinical treatment decision of bladder cancer (BCa). Therefore, establishing an effective prognostic model for BCa has significant clinical implications. We performed WGCNA and DEG screening to initially identify the candidate genes. The candidate genes were applied to construct a LASSO Cox regression analysis model. The effectiveness and accuracy of the prognostic model were tested by internal/external validation and pan-cancer validation and time-dependent ROC. Additionally, a nomogram based on the parameter selected from univariate and multivariate cox regression analysis was constructed. Eight genes were eventually screened out as progression-related differentially expressed candidates in BCa. LASSO Cox regression analysis identified 3 genes to build up the outcome model in E-MTAB-4321 and the outcome model had good performance in predicting patient progress free survival of BCa patients in discovery and test set. Subsequently, another three datasets also have a good predictive value for BCa patients' OS and DFS. Time-dependent ROC indicated an ideal predictive accuracy of the outcome model. Meanwhile, the nomogram showed a good performance and clinical utility. In addition, the prognostic model also exhibits good performance in pan-cancer patients. Our outcome model was the first prognosis model for human bladder cancer progression prediction via integrative bioinformatics analysis, which may aid in clinical decision-making.

Characterization of hypoxia response patterns identified prognosis and immunotherapy response in bladder cancer.

Intra-tumoral hypoxia and immunity are highly correlated with prognosis of tumor patients. Nonetheless, no studies have reported a systematic analysis of the relationship between hypoxia response and immunity in bladder cancer (BLCA). In this study, we comprehensively evaluated the hypoxia response patterns and their association with genomic and clinicopathological characteristics of 1,343 BLCA patients using unsupervised consensus clustering. Five hypoxia response patterns were defined, and the HPXscore was constructed using least absolute shrinkage and selection operator (LASSO)-Cox regression algorithms to represent the individual hypoxia response pattern. The low HPXscore group was characterized by immune activation and high DNA damage repair, which was referred to the immune-inflamed phenotype. However, activation of stromal-related pathways was observed in the high HPXscore group, which is recognized as T cell suppressive and more likely to be an immune-excluded phenotype. Furthermore, the HPXscore was an independent prognostic factor and could act as a good predictor for immunotherapeutic outcomes in BLCA. Thus, depicting a comprehensive landscape of the hypoxia characteristics may therefore help us to interpret the underlying mechanism of immune escape and shed light on the clinical application of hypoxia modification and immune checkpoints targeting immunotherapies for BLCA.

Wild-type IDH1 inhibits the tumor growth through degrading HIF-α in renal cell carcinoma.

The purpose of our study was to explore the effect and intrinsic mechanism of wild-type IDH1 and its substrate α-KG on renal cell carcinoma (RCC). IDH1 was observed lower expression in RCC cell lines. Phenotype experiment was carried out in the wild-type IDH1 and mutant IDH1R132H plasmid treated cell line. The results showed that the wild-type IDH1 could significantly inhibit the proliferation, migration and promote the apoptosis of RCC cell lines, which were consistent with the IDH1's substrate α-KG. The mutant IDH1R132H was found to lose this biological function of IDH1. Moreover, we verified the proliferation inhibition of IDH1 in vivo. In addition, we verified the correlation between IDH1 and hypoxia signal-related proteins in vitro and in vivo, specifically, IDH1 overexpression could significantly reduce the expression of HIF-1α and HIF-2α proteins and its downstream proteins (VEGF, TGF-α). Furthermore, we preliminarily verified the possibility of α-KG in the RCC's treatment by injecting α-KG into the xenograft model. α-KG significantly reduced tumor size and weight in tumor-bearing mice. This study provided a new therapeutic target and small molecule for the study of the treatment and mechanism of RCC.

KNSTRN promotes tumorigenesis and gemcitabine resistance by activating AKT in bladder cancer.

KNSTRN is a component of the mitotic spindle, which was rarely investigated in tumorigenesis. AKT plays an essential role in tumorigenesis by modulating the phosphorylation of various substrates. The activation of AKT is regulated by PTEN and PIP3. Here, we prove KNSTRN is positively correlated with malignancy of bladder cancer and KNSTRN activates AKT phosphorylation at Thr308 and Ser473. More importantly, our study reveals that both KNSTRN and PTEN interact with PH domain of AKT at cell membrane. The amount of KNSTRN interacted with AKT is negatively related to PTEN. Furthermore, PIP3 pull-down assay proves that KNSTRN promoted AKT movement to PIP3. These data suggest KNSTRN may activate AKT phosphorylation by promoting AKT movement to PIP3 and alleviating PTEN suppression. Based on the activation of AKT phosphorylation, our study demonstrates that KNSTRN promotes bladder cancer metastasis and gemcitabine resistance in vitro and in vivo. Meanwhile, the effect of KNSTRN on tumorigenesis and gemcitabine resistance could be restored by AKT specific inhibitor MK2206 or AKT overexpression. In conclusion, we identify an oncogene KNSTRN that promotes tumorigenesis and gemcitabine resistance by activating AKT phosphorylation and may serve as a therapeutic target in bladder cancer.

E3 ubiquitin ligase RNF126 affects bladder cancer progression through regulation of PTEN stability.

E3 ubiquitin ligase RNF126 (ring finger protein 126) is highly expressed in various cancers and strongly associated with tumorigenesis. However, its specific function in bladder cancer (BCa) is still debatable. Here, we found that RNF126 was significantly upregulated in BCa tissue by TCGA database, and our studies indicated that downregulation of RNF126 significantly inhibited cell proliferation and metastasis through the EGFR/PI3K/AKT signaling pathway in BCa cells. Furthermore, we identified PTEN, an inhibitor of the PI3K/AKT signaling pathway, as a novel substrate for RNF126. By co-immunoprecipitation assays, we proved that RNF126 directly interacts with PTEN. Predominantly, PTEN binds to the C-terminal containing the RING domain of RNF126. The in vivo ubiquitination assay showed that RNF126 specifically regulates PTEN stability through poly-ubiquitination. Furthermore, PTEN knockdown restored cell proliferation, metastasis, and tumor formation of BCa cells inhibited by RNF126 silencing in vitro and in vivo. In conclusion, these results identified RNF126 as an oncogene that functions through ubiquitination and degradation of PTEN in BCa.

An Epithelial-Mesenchymal Transition (EMT) Preoperative Nomogram for Prediction of Lymph Node Metastasis in Bladder Cancer (BLCA).

Lymph node (LN) metastasis is a lethal independent risk factor for patients with bladder cancer (BLCA). Accurate evaluation of LN metastasis is of vital importance for disease staging, treatment selection, and prognosis prediction. Several histopathologic parameters are available to predict LN metastasis postoperatively. To date, medical imaging techniques have made a great contribution to preoperatively diagnosis of LN metastasis, but it also exhibits substantial false positives. Therefore, a reliable and robust method to preoperatively predict LN metastasis is urgently needed. Here, we selected 19 candidate genes related to epithelial-mesenchymal transition (EMT) across the LN metastasis samples, which was previously reported to be responsible for the subtype transition and correlation with malignancy and prognosis of BLCA, to establish an EMT-LN signature through LASSO logistic regression analysis. The EMT-LN signature could significantly predict LN metastasis with high accuracy in the TCGA-BLCA cohort, as well as several independent cohorts. As integrating with C3orf70 mutation, we developed an individualized prediction nomogram based on the EMT-LN signature. The nomogram exhibited good discrimination on LN metastasis status, with AUC of 71.7% and 75.9% in training and testing datasets of the TCGA-BLCA cohort. Moreover, the EMT-LN nomogram displayed good calibration with p > 0.05 in the Hosmer-Lemeshow goodness of fit test. Decision curve analysis (DCA) revealed that the EMT-LN nomogram was of high potential for clinical utility. In summary, we established an EMT-LN nomogram integrating an EMT-LN signature and C3orf70 mutation status, which acted as an easy-to-use tool to facilitate preoperative prediction of LN metastasis in BLCA individuals.

WFDC2 suppresses prostate cancer metastasis by modulating EGFR signaling inactivation.

WAP four-disulfide core domain 2 (WFDC2) is a small secretory protein that has been widely studied in ovarian cancer. It has been proven that WFDC2 promotes proliferation and metastasis in ovarian cancer, and serves as a diagnostic biomarker. However, the specific function of WFDC2 in prostate cancer has not been reported. Here, we first screened the diagnostic marker and favorable prognostic factor WFDC2 in prostate cancer by bioinformatics. WFDC2 expression was negatively correlated with Gleason score and metastasis in prostate cancer. Then, we revealed that overexpression of WFDC2, and addition of recombinant protein HE4 can significantly inhibit prostate cancer metastasis in vivo and in vitro. By co-immunoprecipitation and co-localization assays, we proved that WFDC2 binds to the extracellular domain of epidermal growth factor receptor (EGFR). Immunoblot showed that WFDC2 overexpression and recombinant protein HE4 addition inactivated the EGFR/AKT/GSK3B/Snail signaling pathway, and then restrained the progression of epithelial-mesenchymal transition. In conclusion, our study identified that the tumor suppressor WFDC2 can suppress prostate cancer metastasis by inactivating EGFR signaling.

CTHRC1 Is a Prognostic Biomarker and Correlated With Immune Infiltrates in Kidney Renal Papillary Cell Carcinoma and Kidney Renal Clear Cell Carcinoma.

Kidney renal clear cell carcinoma (KIRC) and kidney renal papillary cell carcinoma (KIRP) are the most common RCC types. RCC has high immune infiltration levels, and immunotherapy is currently one of the most promising treatments for RCC. Collagen triple helix repeat containing 1 (CTHRC1) is an extracellular matrix protein that regulates tumor invasion and modulates the tumor microenvironment. However, the association of CTHRC1 with the prognosis and tumor-infiltrating lymphocytes of KIRP and KIRC has not been reported. We examined the CTHRC1 expression differences in multiple tumor tissues and normal tissues via exploring TIMER, Oncomine, and UALCAN databases. Then, we searched the Kaplan-Meier plotter database to evaluate the correlation of CTHRC1 mRNA level with clinical outcomes. Subsequently, the TIMER platform and TISIDB website were chosen to assess the correlation of CTHRC1 with tumor immune cell infiltration level. We further explored the causes of aberrant CTHRC1 expression in tumorigenesis. We found that CTHRC1 level was significantly elevated in KIRP and KIRC tissues relative to normal tissues. CTHRC1 expression associates with tumor stage, histology, lymph node metastasis, and poor clinical prognosis in KIRP. The CTHRC1 level correlates to tumor grade, stage, nodal metastasis, and worse survival prognosis. Additionally, CTHRC1 is positively related to different tumor-infiltrating immune cells in KIRP and KIRC. Moreover, CTHRC1 was closely correlated with the gene markers of diverse immune cells. Also, high CTHRC1 expression predicted a worse prognosis in KIRP and KIRC based on immune cells. Copy number variations (CNV) and DNA methylation might contribute to the abnormal upregulation of CTHRC1 in KIRP and KIRC. In conclusion, CTHRC1 can serve as a biomarker to predict the prognosis and immune infiltration in KIRP and KIRC.

The role and function of PPARγ in bladder cancer.

Peroxisome proliferator-activated receptor gamma (PPARγ), a member of the nuclear receptor superfamily, participates in multiple physiological and pathological processes. Extensive studies have revealed the relationship between PPARγ and various tumors. However, the expression and function of PPARγ in bladder cancer seem to be controversial. It has been demonstrated that PPARγ affects the occurrence and progression of bladder cancer by regulating proliferation, apoptosis, metastasis, and reactive oxygen species (ROS) and lipid metabolism, probably through PPARγ-SIRT1 feedback loops, the PI3K-Akt signaling pathway, and the WNT/ß-catenin signaling pathway. Considering the frequent relapses after chemotherapy, some researchers have focused on the relationship between PPARγ and chemotherapy sensitivity in bladder cancer. Moreover, the feasibility of PPARγ ligands as potential therapeutic targets for bladder cancer has been uncovered. Taken together, this review summarizes the relevant literature and our findings to explore the complicated role and function of PPARγ in bladder cancer.

TRPM8 Inhibition Regulates the Proliferation, Migration and ROS Metabolism of Bladder Cancer Cells.

INTRODUCTION: Based on accumulating evidence, transient receptor potential (TRP) ion channels may play important roles in the occurrence and the progression of cancer. TRP melastatin 8 (TRPM8), a member of the TRP family, functions as a Ca2+-permeable channel and regulates various physiological and pathological processes. However, the effects of TRPM8 on bladder cancer (BCa) and its underlying mechanisms have not been elucidated. METHODS: BCa tissues and matched noncancerous tissues were collected to examine the expression of the TRPM8 mRNA and protein using qRT-PCR, Western blotting and immunofluorescence staining. Meanwhile, the effect of knockdown or inhibition of the activity of the TRPM8 protein on the proliferation, migration and ROS metabolism of bladder cancer cells was detected using the MTT assay, clonogenic survival assay, Transwell chamber migration assay, and reactive oxygen species (ROS) detection, respectively. Furthermore, a mouse model transplanted with BCa cells was established to assess tumor growth after TRPM8 expression was inhibited in vivo. RESULTS: Compared with the noncancerous tissues, the levels of TRPM8 in BCa tissues were significantly increased. Knockdown or inhibition of the activity of the TRPM8 protein in BCa cells reduced cell proliferation and migration. Moreover, the production of ROS was increased in cells treated with siTRPM8, which was accompanied by increased levels of Catalase, HO-1 and SOD2. Furthermore, a mouse model transplanted with the stable TRPM8-deficient T24 cell line was established, demonstrating that knockdown of TRPM8 delayed tumor growth in vivo. DISCUSSION: TRPM8 might play an essential for BCa tumor progression and metastasis by interfering with BCa cell proliferation, motility, ROS metabolism and migration.

miR-601 inhibits proliferation, migration and invasion of prostate cancer stem cells by targeting KRT5 to inactivate the Wnt signaling pathway.

OBJECTIVE: This study aimed to verify the hypothesis that downregulation of miR-601 inhibits the proliferation, migration, and invasion of prostate cancer stem cells (PCSCs) by the Wnt signaling pathway through targeting keratin 5 (KRT5). METHODS: Bioinformatic tools were applied to predict miRNAs and genes potentially associated with prostate cancer, then miR-601 and KRT5 were selected. Subsequently, PCSCs were investigated with respect to miR-601 overexpression or inhibition, KRT5 overexpression, or treatment with a Wnt pathway inhibitor. A series of experiments including western blotting, RT-qPCR, wound healing experiment, transwell assay, MTT assay, annexin V-FITC/PI flow cytometric analysis, nude mice assay and immunohistochemistry were then carried out. RESULTS: Compared with negative control group, migration, invasion, and proliferation of PCSCs and Wnt-1 expression were all enhanced, but apoptosis was attenuated in the miR-601 mimic group. Furthermore, results identified in the other groups (KRT5, miR-601 inhibitor, miR-601 inhibitor + KRT5, Wnt signaling pathway inhibitor, PRI-724/PRI-724 + KRT5) were opposite to those identified with the miR-601 mimic group (all P<0.05). Compared with the miR-601 inhibitor + KRT5 group, migration, invasion, and proliferation of PCSCs and Wnt-1 expression were all increased, whereas apoptosis was suppressed in KRT5 or miR-601 inhibitor groups (all P<0.05). Compared with the PRI-724 + KRT5 group, migration, invasion, and proliferation of PCSCs and Wnt-1 expression were also enhanced, whereas apoptosis was inhibited in PRI-724 or KRT5 groups (all P<0.05). CONCLUSION: Results obtained from the present study have demonstrated that downregulation of miR-601 is able to inhibit the proliferation, migration, and invasion of PCSCs by activating KRT5, and subsequently inhibiting the Wnt pathway.

Identifying a Novel Biomarker TOP2A of Clear Cell Renal Cell Carcinoma (ccRCC) Associated with Smoking by Co-Expression Network Analysis.

Although it is well known that smoking is one of pathogenesis of clear cell renal cell carcinoma (ccRCC), the underlying molecular mechanism is still unclear. In our study, the microarray dataset GSE46699 is analyzed by weighted gene co-expression network analysis (WGCNA). Then we identify 15 co-expressed gene modules in which the lightcyan module (R2 = 0.30) is the most significant. Combined with the protein-protein interaction (PPI) network and WGCNA, two hub genes are identified. Meanwhile, linear regression analyses indicate that TOP2A has a higher connection with smoking in ccRCC, survival analysis proved that overexpression of TOP2A in ccRCC could lead to shorter survival time. Furthermore, bioinformatical analyses based on GSE46699 and GSE2109 as well as qRT-PCR experiment show similar results that TOP2A is significantly up-regulated in smoking ccRCC compared to non-smoking ccRCC samples. In addition, Functional analysis, pathway enrichment analysis and gene set enrichment analysis (GSEA) indicate that high expression of TOP2A is related to cell cycle and p53 signaling pathway in ccRCC samples. Moreover, in vitro experiments revealed that TOP2A induced cell cycle arrest at G2 phase and proliferation inhibition via p53 phosphorylation. Taken together, by using WGCNA, we have identified a novel biomarker named TOP2A, which could affect the development of smoking-related ccRCC by regulating cell cycle and p53 signaling pathway.