The potential mechanism of ursolic acid in the treatment of bladder cancer based on network pharmacology and molecular docking.

OBJECTIVE: This study explored the potential molecular mechanisms of ursolic acid (UA) in bladder cancer treatment using network pharmacology and molecular docking. METHODS: The Traditional Chinese Medicine Systems Pharmacology and UniProt databases were used to screen potential targets of UA. Relevant bladder cancer target genes were extracted using the GeneCards database. All data were pooled and intercrossed to obtain common target genes of UA and bladder cancer. Gene Ontology functional annotation and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses were performed. Molecular docking was conducted to verify the possible binding conformation between UA and bladder cancer cells. Then, in vitro experiments were performed to further validate the predicted results. RESULTS: UA exerts anti-tumor effects on bladder cancer through multiple targets and pathways. Molecular docking indicated that UA undergoes stable binding with the proteins encoded by the top six core genes (STAT3, VEGFA, CASP3, TP53, IL1B, and CCND1). The in vitro experiments verified that UA can induce bladder cancer cell apoptosis by regulating the PI3K/Akt signaling pathway. CONCLUSIONS: Our study illustrated the potential mechanism of UA in bladder cancer based on network pharmacology and molecular docking. The results will provide scientific references for follow-up studies and clinical treatment.

Unraveling neoantigen-associated genes in bladder cancer: An in-depth analysis employing 101 machine learning algorithms.

The therapeutic outcomes for bladder cancer (BLCA) remain suboptimal. Concurrently, there is a growing appreciation for the role of neoantigens in tumors. In this study, we explored the mechanisms underlying the involvement of neoantigen-associated genes in BLCA and their impact on prognosis. Our analysis incorporated both single-cell sequencing and bulk sequencing data sourced from publicly available databases. By employing a comprehensive set of 10 machine learning algorithms, we generated 101 algorithm combinations. The optimal combination, determined based on consistency indices, was utilized to construct a prognostic model comprising nine genes (CAPG, ACTA2, PDIA6, AKNA, PTMS, SNAP23, ID2, CD3G, SP140). Subsequently, we validated this model in an independent cohort, demonstrating its robust testing efficacy. Moreover, we explored the correlations between various clinical traits, model scores, and genes. Leveraging extensive public data resources, we conducted a drug sensitivity analysis to provide insights for targeted drug screening. Additionally, consensus clustering analysis and immune infiltration analysis were performed on bulk sequencing datasets and immunotherapy cohorts. These analyses yield valuable insights into the role of neoantigens in BLCA, guiding future research endeavors.

Combinatorial Gene Expression Profiling of Serum HULC, HOTAIR, and UCA1 lncRNAs to Differentiate Hepatocellular Carcinoma from Liver Diseases: A Systematic Review and Meta-Analysis.

Hepatocellular carcinoma (HCC) presents a significant global health challenge due to limited early detection methods, primarily relying on conventional approaches like imaging and alpha-fetoprotein (AFP). Although non-coding RNAs (ncRNAs) show promise as potential biomarkers in HCC, their true utility remains uncertain. We conducted a comprehensive review of 76 articles, analyzing 88 circulating lncRNAs in 6426 HCC patients. However, the lack of a standardized workflow protocol has hampered holistic comparisons across the literature. Consequently, we herein confined our meta-analysis to only a subset of these lncRNAs. The combined analysis of serum highly upregulated in liver cancer (HULC) gene expression with homeobox transcript antisense intergenic RNA (HOTAIR) and urothelial carcinoma-associated 1 (UCA1) demonstrated markedly enhanced sensitivity and specificity in diagnostic capability compared to traditional biomarkers or other ncRNAs. These findings could have substantial implications for the early diagnosis and tailored treatment of HCC.

Unveiling nature's potential weapon: Magnolol's role in combating bladder cancer by upregulating the miR-124 and inactivating PKC-δ/ERK axis.

BACKGROUND: Bladder cancer (BC) is a challenging disease to manage. Researchers have been investigating the potential of magnolol, a compound derived from Magnolia officinalis, as an anti-cancer agent. However, the exact regulatory mechanism of magnolol and its impact on the NF-κB signaling pathway in BC remain unclear. MATERIALS: To comprehensively evaluate its therapeutic potential, the researchers conducted a series of experiments using BC cell lines (TSGH8301, T24, and MB49) and in vivo animal models. RESULTS: The results of the study demonstrated that magnolol exhibits cytotoxic effects on BC cells by activating both the extrinsic and intrinsic apoptosis signaling pathways. Additionally, the expression of anti-apoptotic genes was downregulated by magnolol treatment. The researchers also uncovered the regulatory role of PKCδ/ERK and miR-124-3p in the NF-κB pathway, which may be influenced by magnolol. Treatment with magnolol led to the inactivation of PKCδ/ERK and an increase in miR-124-3p expression, effectively inhibiting NF-κB-mediated progression of BC. Importantly, the administration of magnolol did not result in significant toxicity in normal tissues, highlighting its potential as a safe adjunctive therapy with minimal adverse effects. CONCLUSION: These findings position magnolol as a promising therapeutic agent for the treatment of BC. By activating apoptosis signaling pathways and inhibiting NF-κB pathway through the upregulation of miR-124-3p and downregulation of PKCδ/ERK activation, magnolol holds promise for suppressing tumor progression and improving patient outcomes in BC. Further research and clinical trials are warranted to explore the full potential of magnolol in the future.

Comparative transcriptomic analysis and mechanistic characterization revealed the use of formononetin for bladder cancer treatment.

Bladder cancer is one of the most common cancers worldwide, with 213 000 deaths reported in 2020. Patients with a progression from non-muscle-invasive bladder cancer to muscle-invasive disease have poorer prognosis and survival rates. Therefore, there is an urgent need to identify novel drugs to prevent the recurrence and metastasis of bladder cancer. Formononetin is an active compound extracted from the herb Astragalus membranaceus that possesses anticancer properties. Few studies have demonstrated the anti-bladder cancer effects of formononetin; however, the detailed mechanism remains unknown. In this study, we used two bladder cancer cell lines, TM4 and 5637, to investigate the potential role of formononetin in bladder cancer treatment. Comparative transcriptomic analysis was conducted to delineate the molecular mechanisms underlying the anti-bladder cancer effects of formononetin. Our results showed that formononetin treatment inhibited the proliferation and colony-forming abilities of bladder cancer cells. Additionally, formononetin reduced the migration and invasion of bladder cancer cells. Transcriptomic analysis further highlighted the involvement of formononetin-mediated two clusters of genes involved in endothelial cell migration (FGFBP1, LCN2, and STC1) and angiogenesis (SERPINB2, STC1, TNFRSF11B, and THBS2). Taken together, our results suggest the potential use of formononetin to inhibit the recurrence and metastasis of bladder cancer through the regulation of different oncogenes.

Competition between p53 and YY1 determines PHGDH expression and malignancy in bladder cancer.

PURPOSE: Serine metabolism is frequently dysregulated in many types of cancers and the tumor suppressor p53 is recently emerging as a key regulator of serine metabolism. However, the detailed mechanism remains unknown. Here, we investigate the role and underlying mechanisms of how p53 regulates the serine synthesis pathway (SSP) in bladder cancer (BLCA). METHODS: Two BLCA cell lines RT-4 (WT p53) and RT-112 (p53 R248Q) were manipulated by applying CRISPR/Cas9 to examine metabolic differences under WT and mutant p53 status. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) and non-targeted metabolomics analysis were adopted to identify metabolomes changes between WT and p53 mutant BLCA cells. Bioinformatics analysis using the cancer genome atlas and Gene Expression Omnibus datasets and immunohistochemistry (IHC) staining was used to investigate PHGDH expression. Loss-of-function of PHGDH and subcutaneous xenograft model was adopted to investigate the function of PHGDH in mice BLCA. Chromatin immunoprecipitation (Ch-IP) assay was performed to analyze the relationships between YY1, p53, SIRT1 and PHGDH expression. RESULTS: SSP is one of the most prominent dysregulated metabolic pathways by comparing the metabolomes changes between wild-type (WT) p53 and mutant p53 of BLCA cells. TP53 gene mutation shows a positive correlation with PHGDH expression in TCGA-BLCA database. PHGDH depletion disturbs the reactive oxygen species homeostasis and attenuates the xenograft growth in the mouse model. Further, we demonstrate WT p53 inhibits PHGDH expression by recruiting SIRT1 to the PHGDH promoter. Interestingly, the DNA binding motifs of YY1 and p53 in the PHGDH promoter are partially overlapped which causes competition between the two transcription factors. This competitive regulation of PHGDH is functionally linked to the xenograft growth in mice. CONCLUSION: YY1 drives PHGDH expression in the context of mutant p53 and promotes bladder tumorigenesis, which preliminarily explains the relationship between high-frequency mutations of p53 and dysfunctional serine metabolism in bladder cancer.

Over-expression of Long Non-coding RNA Urothelial Cancer-associated 1 as a Predictive Marker for Prostate Cancer.

BACKGROUND/AIM: To determine the expression of long non-coding RNA urothelial cancer-associated 1 (UCA1) by performing array-based quantitative polymerase chain reaction (PCR) and to identify the clinicopathological significance of UCA1 expression in prostate cancer using in situ hybridization (ISH) of surgically resected specimens. MATERIALS AND METHODS: Array-based quantitative PCR was performed using 10 pairs of fresh malignant (prostate cancer) and normal tissue samples to determine UCA1 expression. Single-color RNA ISH of surgically resected prostate cancer specimens was performed using 70 formalin-fixed, paraffin-embedded tissue specimens to examine the clinicopathological significance of UCA1. RESULTS: Prostate cancer tissues exhibited higher levels of UCA1 expression than paired benign tissues. Furthermore, a correlation between high UCA1 expression and unfavourable clinicopathological characteristics, including advanced pathologic T stage, extraprostatic extension, presence of Gleason pattern 5, and involvement of the resection margins was observed. Notably, increased UCA1 expression significantly correlated with high- or very-high-risk patients, as defined by the 2023 National Comprehensive Cancer Network guidelines. CONCLUSION: UCA1 could be used as a novel diagnostic and prognostic biomarker for establishing an effective treatment protocol for prostate cancer.

Using chitosan-stabilized, hyaluronic acid-modified selenium nanoparticles to deliver CD44-targeted PLK1 siRNAs for treating bladder cancer.

Aims: Achieving an effective biocompatible system for siRNAs delivery to the tumor site remains a significant challenge. Materials & methods: Selenium nanoparticles (SeNPs) modified by chitosan (CS) and hyaluronic acid (HA) were fabricated for PLK1 siRNAs (siPLK1) delivery to the bladder cancer cells. The HA-CS-SeNP@siPLK1 efficacy was evaluated using in vitro and in vivo models. Results: HA-CS-SeNP@siPLK1 was selectively internalized into T24 cells through clathrin-mediated endocytosis. Treatment with HA-CS-SeNP@siPLK1 successfully silenced the PLK1 gene, inhibited cell proliferation and induced cell cycle arrest in vitro. HA-CS-SeNP@siPLK1 could also inhibit tumor growth in vivo without causing systemic toxicity. Conclusion: Our results suggest that HA-CS-SeNPs may provide a good vehicle for delivering siPLK1 to the bladder tumor site.

siRNAs are small biomolecules shown as novel insights in cancer gene therapy because of their capability to silence target genes. However, achieving an effective biocompatible system for siRNA delivery to the tumor site remains a significant challenge. This work aimed to develop a nanoparticle-based delivery system consisting of selenium nanoparticles modified by chitosan and hyaluronic acid to sustain the release of siRNAs to bladder cancer cells. The results of this study demonstrated that this nanosystem successfully silenced the PLK1 gene and reduced the proliferation in vitro and in vivo. These findings suggest that hyaluronic acid-chitosan-selenium nanoparticles may open a new insight for targeted gene therapy for bladder cancer.

Bladder cancer organoids as a functional system to model different disease stages and therapy response.

Bladder Cancer (BLCa) inter-patient heterogeneity is the primary cause of treatment failure, suggesting that patients could benefit from a more personalized treatment approach. Patient-derived organoids (PDOs) have been successfully used as a functional model for predicting drug response in different cancers. In our study, we establish PDO cultures from different BLCa stages and grades. PDOs preserve the histological and molecular heterogeneity of the parental tumors, including their multiclonal genetic landscapes, and consistently share key genetic alterations, mirroring tumor evolution in longitudinal sampling. Our drug screening pipeline is implemented using PDOs, testing standard-of-care and FDA-approved compounds for other tumors. Integrative analysis of drug response profiles with matched PDO genomic analysis is used to determine enrichment thresholds for candidate markers of therapy response and resistance. Finally, by assessing the clinical history of longitudinally sampled cases, we can determine whether the disease clonal evolution matched with drug response.

Long non­coding RNA BLACAT2/miR­378a­3p/YY1 feedback loop promotes the proliferation, migration and invasion of uterine corpus endometrial carcinoma.

Uterine corpus endometrial carcinoma (UCEC) is a common gynecological malignancy with high rates of mortality and morbidity. The expression of long non­coding RNA bladder cancer­associated transcript 2 (BLACAT2) has been previously found to be aberrantly upregulated in UCEC. However, the regulatory consequences of this in UCEC progression remain poorly understood. In the present study, human UCEC cell lines AN3CA and HEC­1­A were infected with lentiviruses to overexpress BLACAT2 (Lv­BLACAT2) or knock down BLACAT2 using short hairpin RNA (Lv­shBLACAT2). BLACAT2 overexpression was found to promote the G1/S transition of cell cycle progression and UCEC cell proliferation. In addition, BLACAT2 overexpression was observed to facilitate UCEC cell migration and invasion. By contrast, BLACAT2 knockdown resulted in inhibitory effects in UCEC cell physiology. BLACAT2 overexpression also contributed to the activation of the MEK/ERK pathway. Subsequently, BLACAT2 was demonstrated to bind to microRNA (miR)­378a­3p according to dual­luciferase assays, where it appeared to function as a sponge of miR­378a­3p in 293T cells. miR­378a­3p overexpression was found to suppress UCEC cell proliferation, invasion, and ERK activation. Lentivirus­mediated knockdown of its target, the transcription factor Yin Yang­1 (YY1), was observed to reverse the oncogenic effects of BLACAT2 overexpression. Furthermore, YY1 was found to bind to the promoter of BLACAT2, suggesting that YY1 can regulate BLACAT2 expression. To conclude, results from the present study suggest that BLACAT2, miR­378a­3p and YY1 can form a feedback loop instead of an unidirectional axis, which can in turn regulate UCEC tumorigenesis through the MEK/ERK pathway. The present study furthered the understanding of UCEC tumorigenesis and may provide novel therapeutic targets for UCEC treatment.

Inhibition of SND1 overcomes chemoresistance in bladder cancer cells by promoting ferroptosis.

Chemotherapy remains one of the most important adjuvant treatments for bladder cancer (BC). However, similar to other malignancies, BC is prone to chemotherapy resistance and only approximately half of muscle­invasive patients with BC respond to chemotherapy. The present study aimed to reveal the mechanisms underlying chemoresistance in BC cells. Cell viabilities were assessed by CCK­8 assay. The differentiated expression of genes in chemoresistant and their parental BC cells were examined by RNA sequencing. Cell death was determined by flow cytometry. Different cell death inhibitors were used to determine the types of cell death. Levels of reactive oxygen species, iron, glutathione and malondialdehyde were assessed using the corresponding commercial kits. ChIP and dual luciferase activity assays were performed to investigate the interaction between staphylococcal nuclease and tumour domain containing 1 (SND1) and glutathione peroxidase 4 (GPX4) mRNA. RNAi was used to knockdown SND1 or GPX4. The results revealed that SND1 in BC cells were insensitive to cisplatin, and inhibition of SND1 overcame this resistance. Silencing of SND1 enhanced cell death induced by cisplatin by promoting ferroptosis in BC cells. Mechanistically, SND1 was revealed to bind to the 3'UTR region of GPX4 mRNA and stabilise it. Knockdown of GPX4 could also overcome chemoresistance, and overexpressing GPX4 reversed the effects of silencing of GPX4 on the chemosensitivity of BC cells. Thus, targeting the SND1­GPX4 axis may be a potential strategy to overcome chemoresistance in BC cells.

Integrative tumour mutation burden with CD39 and PD-L1 for the prediction of response to PD-L1 blockade and adjuvant chemotherapy in muscle-invasive bladder cancer patients.

BACKGROUND: CD39, a rate-limiting enzyme to convert extracellular ATP (eATP) to adenosine, has been reported to be a key modulator of immune response, but its correlation with therapeutic sensitivity remains obscure. We conducted this study to determine whether the integration of CD39 and traditional biomarkers could improve the prediction of responsiveness to PD-L1 blockade and platinum-based chemotherapy. METHODS: We retrospectively enrolled a total of 760 patients from IMvigor210 trial, TCGA database and Zhongshan Hospital in this study. We constructed the CPT scoring system based on CD39, PD-L1 and tumour mutation burden (TMB) and validated its efficacy in predicting therapeutic responsiveness in MIBC patients. Kaplan-Meier survival and Cox regression analyses were applied to assess clinical outcomes of patients. RESULTS: The CPT scoring system could predict the response to PD-L1 blockade and platinum-based chemotherapy. The CPT score was positively correlated with APOBEC mutational signature and SNV neoantigens enrichment, antigen presentation, and TCR signalling. High CPT score also indicated the inflamed immune phenotype and basal/squamous molecular subtype. CONCLUSIONS: CD39 expression is closely correlated with the immunogenic contexture of MIBC. Integrating CD39 with PD-L1 and TMB could stratify the sensitivity of patients with MIBC to PD-L1 blockade and platinum-based chemotherapy.

Omega-3 polyunsaturated fatty acids: a modified approach for chemo-prevention of bladder cancer in a rat model and molecular studies of antineoplastic mechanisms.

OBJECTIVES: To test the chemo-preventative effects of omega-3 against bladder cancer (BC) induction in a rat model and its potential antineoplastic mechanisms. MATERIAL AND METHODS: Ninety male Fisher rats were divided into three groups during a 22-week protocol: group 1 (control), group 2 (Placebo + N-butyl-N-4- hydroxybutyl nitrosamine (BBN) for induction of BC and group 3 received omega-3 (1200 mg/kg/day) + BBN. At the end, blood samples and bladder tissues were collected and checked for the presence of malignancy, markers of angiogenesis (VEGF relative gene expression), inflammation (IL-6), proliferation (KI-67 expressions), oxidative stress (serum MDA and serum SOD) and epigenetic control (miRNA-145 level). RESULTS: At the end of the study, 60% and 86.6% rats survived in group 2 and 3 with significant weight loss among rats in group 2 when compared with other groups. In group 2, all rats developed visible bladder lesions of which five and 13 developed squamous cell carcinoma (SCC) and transitional cell carcinoma (TCC). In omega3-treated group, only one developed low grade SCC and one developed high grade non- invasive TCC. Bladders from omega-3-treated rats showed lower expression ofKI-67 (p < 0.05), VEGF (p < 0.001) and IL-6 (p < 0.001) and significant higher expression of mi-RNA (p < 0.001). Also, omega-3-treated group showed statistically significant lower MDA level (p < 0.001). CONCLUSION: Omega-3 inhibits bladder tumor growth in the BBN-induced BC rat model, due to anti-inflammatory, antioxidant, anti-proliferative, and anti-angiogenic properties together with epigenetic control.

Mechanisms of Compound Kushen Injection for the treatment of bladder cancer based on bioinformatics and network pharmacology with experimental validation.

Bladder cancer is the most common malignancy of the urinary system. Compound Kushen Injection (CKI) is a Chinese medicinal preparation that has been widely used in the treatment of various types of cancers in the past two decades. However, the pharmacological effect of CKI on bladder cancer is not still completely understood. In the current study, network pharmacology combined with bioinformatics was used to elucidate the therapeutic mechanism and potential targets of CKI in bladder cancer. The mechanism by which CKI was effective against bladder cancer was further verified in vitro using human bladder cancer cell line T24. Network pharmacology analysis identified 35 active compounds and 268 target genes of CKI. Bioinformatics data indicated 5500 differentially expressed genes associated with bladder cancer. Common genes of CKI and bladder cancer suggested that CKI exerted anti-bladder cancer effects by regulating genes such as MMP-9, JUN, EGFR, and ERK1. Functional enrichment analysis indicated that CKI exerted therapeutic effects on bladder cancer by regulating certain biological processes, including cell proliferation, cell migration, and cell apoptosis. In addition, Kyoto Encyclopedia of Genes and Genomes enrichment analysis implicated pathways related to cancer, bladder cancer, and the PI3K-Akt signaling pathway. Consistently, cell experiments indicated that CKI inhibited the proliferation and migration of T24 cells, and induced their apoptosis. Moreover, RT-qPCR and Western blot results demonstrated that CKI was likely to treat bladder cancer by down-regulating the gene and protein expression of MMP-9, JUN, EGFR, and ERK1. CKI inhibited the proliferation and migration, and induced the apoptosis of T24 bladder cancer cells through multiple biological pathways and targets. CKI also exhibited significant effects on the regulation of key genes and proteins associated with bladder cancer. Overall, our findings provide solid evidence and deepen current understanding of the therapeutic effects of CKI for bladder cancer, and further support its clinical use.

High ORAI3 expression correlates with good prognosis in human muscle-invasive bladder cancer.

The involvement of store-operated calcium channels (SOCCs) in tumor initiation and metastatic dissemination has been extensively studied, but how its member ORAI3 influences tumor progression is still elusive. The present study aimed to evaluate the prognostic value of ORAI3 expression and examine the correlation between ORAI3 expression and immune cell infiltration within the tumor microenvironment (TME) in human muscle-invasive bladder cancer (MIBC). We examined the expression profile of ORAI3 in MIBC using data from two databases; analyzed the correlation between ORAI3 expression and patient survival; explored cellular pathways related to ORAI3 expression by Gene Set Enrichment Analysis (GSEA); and predicted potential drugs using Connectivity Map (CMap). ORAI3 was significantly lower expressed in tumor mass compared to normal samples in MIBC, with a higher level of methylation at the promoter region in tumor than in normal tissue, indicating that ORAI3 is suppressed during cancer progression. Survival analysis showed that higher expression of ORAI3 correlated with good prognosis in MIBC. GSEA demonstrated that ORAI3 expression inversely correlated with cell differentiation, development and gene silencing, with differential expression of genes involved in epidermal and keratinocyte differentiation pathways and inflammatory responses. RNA sequencing of an ORAI3-silenced human bladder cancer cell line (T24 cells) corroborated enhancement of pro-neoplastic pathways in absence of ORAI3. Western blottingMoreover, ORAI3 facilitated the recruitment of Th17 cells and natural killer cells, whereas hampered Th2 and macrophage infiltration. Our results revealed 4 molecules with potential to be beneficial as adjuvant drugs in MIBC treatment. We concluded that high ORAI3 expression correlates with increased survival in human MIBC.

[Expression pattern of the histone lysine demethylase family and its potential role in bladder cancer: a multi-omics analysis].

OBJECTIVE: To investigate the expression patterns of 19 histone lysine demethylases (KDMs) and their role in bladder cancer. METHODS: In this study, UALCAN and GSCALite were used to analyze the transcriptional expression, methylation level and somatic variation of KDMs in bladder cancer samples from TCGA. Kaplan Meier-Plotter and Assistant for clinical bioinformatics were used to investigate the effect of KDMs expression on the prognosis of BLCA samples. The immune infiltration and drug sensitivity of KDMs in bladder cancer were analyzed by Timer and GSCALite. RESULTS: The KDMs did not show consistent expressions patterns in bladder cancer, where the expressions of KDM1A/1B/2B/4A/4B/5B/5C were significantly upregulated while those of KDM3B/6B/7C were significantly downregulated. Methylation data analysis showed that methylation levels of KDM1A/3B/4A/4B/4C/5A/5B/5C/7B were significantly downregulated and that of KDM7C was upregulated. The transcription levels of 14 KDMs had significant negative correlations with their methylation levels, and among them KDM1A showed the strongest correlation. Mutation analysis revealed that KDM6A had the highest frequency of nonsynonymous mutations with the largest variety, and these mutations were complementary to nonsynonymous mutations of the other KDMs. Survival analysis showed that KDM3A/4C/5D/6A/7B were protective for OS while KDM3B/5B/5C adversely affected RFS of BLCA patients. Further comprehensive prognostic modeling confirmed that KDM4C/6A/7B were potential prognostic biomarkers of bladder cancer, and their expressions were positively correlated with immune infiltration in BLCA patients. KDM2B/3B/4B/4C/5A were negatively correlated with the sensitivity to most anticancer drugs, while KDM2B/4B were positively correlated with the sensitivity to 4 anticancer drugs. CONCLUSION: The expression patterns of the KDMs in bladder cancer highlight a high mutation complementarity and a negative correlation between over-expression and hypomethylation level closely related with the prognosis, immune infiltration and drug sensitivity.

Mechanisms of Compound Kushen Injection for the treatment of bladder cancer based on bioinformatics and network pharmacology with experimental validation / 中国天然药物

Bladder cancer is the most common malignancy of the urinary system. Compound Kushen Injection (CKI) is a Chinese medicinal preparation that has been widely used in the treatment of various types of cancers in the past two decades. However, the pharmacological effect of CKI on bladder cancer is not still completely understood. In the current study, network pharmacology combined with bioinformatics was used to elucidate the therapeutic mechanism and potential targets of CKI in bladder cancer. The mechanism by which CKI was effective against bladder cancer was further verified in vitro using human bladder cancer cell line T24. Network pharmacology analysis identified 35 active compounds and 268 target genes of CKI. Bioinformatics data indicated 5500 differentially expressed genes associated with bladder cancer. Common genes of CKI and bladder cancer suggested that CKI exerted anti-bladder cancer effects by regulating genes such as MMP-9, JUN, EGFR, and ERK1. Functional enrichment analysis indicated that CKI exerted therapeutic effects on bladder cancer by regulating certain biological processes, including cell proliferation, cell migration, and cell apoptosis. In addition, Kyoto Encyclopedia of Genes and Genomes enrichment analysis implicated pathways related to cancer, bladder cancer, and the PI3K-Akt signaling pathway. Consistently, cell experiments indicated that CKI inhibited the proliferation and migration of T24 cells, and induced their apoptosis. Moreover, RT-qPCR and Western blot results demonstrated that CKI was likely to treat bladder cancer by down-regulating the gene and protein expression of MMP-9, JUN, EGFR, and ERK1. CKI inhibited the proliferation and migration, and induced the apoptosis of T24 bladder cancer cells through multiple biological pathways and targets. CKI also exhibited significant effects on the regulation of key genes and proteins associated with bladder cancer. Overall, our findings provide solid evidence and deepen current understanding of the therapeutic effects of CKI for bladder cancer, and further support its clinical use.

CRTAC1 (Cartilage acidic protein 1) inhibits cell proliferation, migration, invasion and epithelial-mesenchymal transition (EMT) process in bladder cancer by downregulating Yin Yang 1 (YY1) to inactivate the TGF-ß pathway.

Cartilage acidic protein 1 (CRTAC1) is predicted to be aberrantly expressed in bladder cancer based on bioinformatics analysis. However, its functions and molecular mechanism in bladder cancer remain elusive. This study aimed to explore the role of CRTAC1 in bladder cancer. The mRNA and protein levels of CRTAC1 and Yin Yang 1 (YY1) were detected by reverse transcription quantitative polymerase chain reaction and western blotting. We found that CRTAC1 was downregulated in bladder cancer tissues and cells. Cell Counting Kit-8 assays, colony formation assays, wound healing assays and Transwell assays and western blotting revealed that CRTAC1 overexpression inhibited cell viability, proliferation, migration, invasion and epithelial-mesenchymal transition (EMT) process in bladder cancer, while CRTAC1 knockdown exerted opposite effects on these malignant behaviors. Mechanistically, CRTAC1 targeted YY1 in bladder cancer cells. YY1 was upregulated in bladder cancer tissues and cells. CRTAC1 negatively modulated the mRNA and protein expression of YY1 in bladder cancer cells. Co-localization of CRTAC1 and YY1 expression was assessed using immunofluorescence staining and Co-Immunoprecipitation assays. The interaction between CRTAC1 and YY1 was explored by Chromatin immunoprecipitation and luciferase reporter assays. Moreover, CRTAC1 inactivated the TGF-ß pathway by downregulating YY1 expression. Protein levels of factors associated with the TGF-ß pathway were examined by western blotting. Rescue assays indicated that CRTAC1 inhibited malignant behaviors of bladder cancer cells by targeting YY1. Overall, CRTAC1 inhibited malignant phenotypes of bladder cancer cells by targeting YY1 to inactivate the TGF-ß pathway.

YY1-induced long non-coding RNA PSMA3 antisense RNA 1 functions as a competing endogenous RNA for microRNA 214-5p to expedite the viability and restrict the apoptosis of bladder cancer cells via regulating programmed cell death-ligand 1.

Bladder cancer (BC) is one of the most common malignant tumors in the urinary system. Our research aimed to explore the function and underlying mechanisms of long noncoding RNA (lncRNA) PSMA3-AS1 in BC. RT-qPCR was utilized to detect the levels of PSMA3-AS1, miR-214-5p, and PD-L1. ChIP assay was employed to confirm the transcription factor of PSMA3-AS1. Luciferase reporter assay was carried out to demonstrate the relationships between miR-214-5p and PSMA3-AS1 or PD-L1. The diagnostic value of PSMA3-AS1 was evaluated by the ROC curve. CCK-8, wound healing, transwell, and flow cytometry assays were applied to analyze cell viability, migration, invasion, and apoptosis. Western blotting was used to confirm the expression of cleaved caspase-3. The present study revealed that BC tissues and cells exhibited an increased expression in PSMA3-AS1. High expression of PSMA3-AS1 was related to poor prognosis in BC patients. Then, the area under the ROC curve for PSMA3-AS1 was up to 0.8954. Moreover, ChIP assay elaborated that YY1 could bind to the PSMA3-AS1 promoter region. Furthermore, it was found that that PSMA3-AS1 knockdown repressed BC cell viability and metastasis, and promoted apoptosis. In addition, miR-214-5p was inversely correlated with PSMA3-AS1 or PD-L1 levels. MiR-214-5p deletion reversed the impacts of PSMA3-AS1 deletion on BC progression, and PD-L1 inhibition also abrogated the influence of miR-214-5p deletion in BC development. In conclusion, YY1-induced PSMA3-AS1 exerted an oncogenic function in BC cells via targeting miR-214-5p and enhancing PD-L1, providing potential biomarkers for BC therapy.

Grape Seed Proanthocyanidins Inhibit Migration and Invasion of Bladder Cancer Cells by Reversing EMT through Suppression of TGF-ß Signaling Pathway.

Bladder cancer (BC) is the most common cancer of the urinary system. Despite advances in diagnosis and therapy, the prognosis is still poor because of recurrence and metastasis. Epithelial-mesenchymal transition (EMT) is considered to play an important role in the invasion and metastasis of BC. Grape seed proanthocyanidins (GSPs) exhibit chemopreventive and chemotherapeutic activities against several types of cancer. However, their effects and underlying mechanisms on the invasive potential of BC remain unclear. In this study, we found that GSPs inhibited migration, invasion, and MMP-2/-9 secretion of both T24 and 5637 bladder cancer cells at noncytotoxic concentrations. We also discovered that 5637 cells were more suitable than T24 cells for the EMT study. Further study showed that GSPs inhibited EMT by reversing the TGF-ß-induced morphological change and upregulation of mesenchymal markers N-cadherin, vimentin, and Slug as well as downregulation of epithelial markers E-cadherin and ZO-1 in 5637 cells. GSPs also inhibited TGF-ß-induced phosphorylation of Smad2/3, Akt, Erk, and p38 in 5637 cells without affecting the expression of total Smad2/3, Akt, Erk, and p38. Taken together, the results of the present study demonstrate that GSPs effectively inhibit the migration and invasion of BC cells by reversing EMT through suppression of the TGF-ß signaling pathway, which indicates that GSPs could be developed as a potential chemopreventive and therapeutic agent against bladder cancer.