Therapeutic Target For Bladder Cancer Research Articles

N6-Methyladenosine enhances the translation of ENO1 to promote the progression of bladder cancer by inhibiting PCNA ubiquitination

The mechanism underlying N6-methyladenosine (m6A) modification in bladder cancer (BC) remains elusive. We identified that the RBM15/METTL3 complex enhances m6A modification and promotes the ENO1 protein translation efficiency through its 359A site by depending on YTHDF1 in BC cells. In the tumor microenvironment, TGF-β effectively stimulates RBM15/METTL3 expression to improve ENO1 mRNA m6A modification through the Smad2/3 pathway. Reduced ENO1 m6A levels hamper tumor proliferation both in vitro and in vivo. Mechanistically, ENO1 augments PCNA protein stability by reducing its K48-linked ubiquitination and thus prevents protein degradation through the endoplasmic reticulum-associated degradation pathway. According to the subsequent experiments, the ENO1 inhibitor significantly reduced tumor proliferation both in vitro and in vivo. Our study highlights the significance of RBM15/METTL3 complex-mediated ENO1 mRNA m6A modification in ENO1 expression. It also reveals a novel mechanism by which ENO1 promotes BC progression, thereby suggesting that ENO1 can be a therapeutic target for BC.

Abstract B009: Targeting bladder cancer-macrophage interactions by SPARC peptides

Abstract Bladder cancer is one of the privileged cancers treated with two modalities of immune therapeutics: intravesical Bacille Calmette-Guérin (BCG) for non-muscle invasive disease and immune checkpoint inhibitors for advanced metastatic muscle invasive disease. However, recurrence, intolerance and adverse events are encountered in both modalities with limited second and third line treatment options. Clinical data indicated that pretreatment infiltration of bladder tumors with tumor associated macrophages (TAMs) is associated with BCG failure, resistance to standard of care chemo- and immune-therapy and poor disease outcome. Preclinical studies indicated that macrophage infiltration is the forerunner of carcinogen-induced preneoplastic bladder lesions and progresses throughout overt neoplastic and metastatic disease. Thus, TAMs represent a plausible therapeutic target for bladder cancer. Capitalizing on our earlier report that SPARC, as tumor suppressor of BCa, we developed peptides mapping the biologically active domains of SPARC and investigated their direct effect on tumor cell malignant phenotype. In tandem, we determined the effect of SPARC-peptides on macrophage differentiation, polarization, and anti-tumor effects. We identified a potent inhibitory effect of our peptides on a panel of bladder cancer cell survival and invasiveness in 2D and 3D cultures. Using macrophage cell lines as well as bone marrow-derived myeloid cells, we found these peptides induced an M1 phenotype in response to cytokine stimulation, with enhanced phagocytic activity and macrophage-induced tumor cell killing in vitro. Moreover, SPARC-peptides inhibited macrophage polarization to M2 phenotype. Our data strongly suggest that SPARC-peptides are promising therapeutic candidates for bladder cancer. Citation Format: Mariam Diab, Neveen Said. Targeting bladder cancer-macrophage interactions by SPARC peptides [abstract]. In: Proceedings of the AACR Special Conference on Bladder Cancer: Transforming the Field; 2024 May 17-20; Charlotte, NC. Philadelphia (PA): AACR; Clin Cancer Res 2024;30(10_Suppl):Abstract nr B009.

Abstract A026: PIN1 isomerase promotes the initiation and progression of bladder cancer through SREBP2-mediated cholesterol biosynthesis pathway

Abstract Bladder cancer, one of the most commonly diagnosed cancers worldwide, is associated with high morbidity, mortality, and a poor prognosis. The PIN1 phospho-dependent prolyl isomerase is frequently overexpressed in many human cancer types and affects tumor initiation and progression through disrupting the balance of oncoprotein and tumor suppressor protein signaling. However, the functional pSer/Thr.Pro protein substrates of PIN1 and its effects on downstream signaling in bladder carcinogenesis remain largely unknown. Phenotypically, we discovered that ablation of Pin1 in human/mouse bladder cancer cells and organoids formed from mouse primary urothelial cells resulted in decreased cell proliferation, stemness maintenance, cell invasion, migration, urothelium clearance capacity, and reduced free/total cholesterol levels in vitro. Re-expressing Pin1 in Pin1 knockout cells reversed the defects caused by lack of Pin1. Moreover, in vivo subcutaneous xenograft and allograft transplantation mouse models combined with an orthotopic implantation model were utilized to confirm a positive role for Pin1 in controlling tumor growth and potential metastasis. Therapeutically, a combination of the sulfopin PIN1 inhibitor and the simvastatin HMGCR inhibitor was shown to suppress cell proliferation in vitro and tumor growth in vivo synergistically. Mechanistically, we observed a negative enrichment of SREBP2-driven cholesterol metabolism pathways in PIN1 knockout cells via RNA-sequencing and used a suite of diverse molecular and biochemical approaches to show that PIN1 directly binds to JNK-dependent phosphorylated SREBP2 (Ser455) and vice versa. These results indicate that active cholesterol biosynthesis pathway in bladder cancer cells is regulated by PIN1-mediated isomerization and JNK-mediated phosphorylation of its key transcriptional factor SREBP2. These findings emphasize that PIN1 can act as a regulator and potential therapeutic target in bladder cancer. Citation Format: Xue Wang, Tony Hunter. PIN1 isomerase promotes the initiation and progression of bladder cancer through SREBP2-mediated cholesterol biosynthesis pathway [abstract]. In: Proceedings of the AACR Special Conference on Bladder Cancer: Transforming the Field; 2024 May 17-20; Charlotte, NC. Philadelphia (PA): AACR; Clin Cancer Res 2024;30(10_Suppl):Abstract nr A026.

CircACTR2 promotes bladder cancer progression through IKBKB-mediated NF-κB signaling pathway activation

BackgroundCircular RNAs (circRNAs) have significant roles in tumor progression. The role of circRNA derived from ARP2 actin-related protein 2 homolog (circACTR2) has been reported in various human diseases. However, the functions and regulatory mechanisms of circACTR2 in Bladder Cancer (BCa) remain unknown. ObjectivesThis study aims to explore the biological role and regulatory mechanism of circACTR2 in BCa. MethodsWe analyzed the effects of circACTR2 on BCa through bioinformatics analyses, RT-qPCR, and cell function assays. ResultsWe observed the upregulation of circACTR2 in BCa tissues and validated its circular structure. Loss-of-function assays demonstrated that silencing circACTR2 suppressed the proliferation, invasion, and migration of BCa cells. Mechanistic investigation revealed that circACTR2 sponges miR-219a-2-3p to elevate the expression of the inhibitor of nuclear factor kappa B kinase subunit beta (IKBKB). This induced upregulation of IKKβ protein promoted the nuclear translocation of p65, thereby activating the NF-κB signaling pathway. ConclusionsOur findings indicate that circACTR2 promotes BCa cell proliferation, migration, and invasion by activating the NF-κB signaling pathway via the miR-219a-2-3p/IKBKB axis, potentially unveiling a new therapeutic target for BCa.

Nucleoporin 93 Regulates Cancer Cell Growth and Stemness in Bladder Cancer via Wnt/β-Catenin Signaling.

Bladder cancer (BLCA) is a prevalent cancer type with an unmet need for new therapeutic strategies. Nucleoporin 93 (Nup93) is implicated in the pathophysiology of several cancers, but its relationship with bladder cancer remains unclear. Nup93 expression was analyzed in TCGA datasets and 88 BLCA patient samples. Survival analysis and Cox regression models evaluated the association between Nup93 levels and patient prognosis. BLCA cells were used to investigate the effects of Nup93 overexpression or knockdown on cell growth, invasion, stemness (sphere formation and ALDH2 + cancer stem cell marker), and Wnt/β-catenin signaling in vitro. The Wnt activator BML-284 was used to confirm the involvement of Wnt/β-catenin signaling pathway. A xenograft mouse model validated the in vitro findings. Nup93 was highly expressed in BLCA tissues and cell lines, and high Nup93 expression correlated with poor prognosis in BLCA patients. Nup93 silencing inhibited BLCA cell proliferation, Wnt/β-catenin activation, and cancer cell stemness. Conversely, Nup93 overexpression promoted these effects. BML-284 partially rescued the reduction in cell growth and stemness markers caused by Nup93 knockdown. Nup93 knockdown also suppressed the tumor formation of BLCA cells in vivo. Nup93 regulates BLCA cell growth and stemness via the Wnt/β-catenin pathway, suggesting its potential as a prognostic marker and therapeutic target in BLCA.

Integrative analysis of the bladder cancer from a cell cycle NCAM1 perspective at both single cell and bulk resolution.

Bladder cancer (BLCA) is a prevalent and deadly form of urinary cancer, and there is a need for effective therapies, particularly for muscle-invasive bladder cancer (MIBC). Cell cycle inhibitors show promise in restoring control of the cell cycle in BLCA cells, but their clinical prognosis evaluation is limited. Transcriptome and scRNA-seq data were collected from the Cancer Genome Atlas Program (TCGA)-BLCA and GSE190888 cohort, respectively. R software and the Seurat package were used for data analysis, including cell quality control, dimensionality reduction, and identification of differentially expressed genes. Genes related to the cell cycle were obtained from the genecards website, and a protein-protein interaction network analysis was performed using cytoscape software. Functional enrichment analysis, immune infiltration analysis, drug sensitivity analysis, and molecular docking were conducted using various tools and packages. BLCA cell lines were cultured and transfected for in vitro experimental assays, including RT-qPCR analysis, and CCK-8 cell viability assays. We identified 32 genes as independent risk or protective factors for BLCA prediction. Functional enrichment analysis revealed their involvement in cell cycle regulation, apoptosis, and various signaling pathways. Using these genes, we developed a nomogram for predicting BLCA survival, which displayed high prognosis stratification efficacy in BLCA patients. Four cell cycle associated key genes identified, including NCAM1, HBB, CKD6, and CTLA4. We also identified the main cell types in BLCA patients and investigated the functional differences between epithelial cells based on their expression levels of key genes. Furthermore, we observed a high positive correlative relationship between the infiltration of cancer-associated fibroblasts and the risk score value. Finally, we conducted in vitro experiments to demonstrate the suppressive role of NCAM1 in BLCA cell proliferation. These findings suggest that cell cycle associated genes could serve as potential biomarkers for predicting BLCA prognosis and may represent therapeutic targets for the development of more effective therapies. Hopefully, these findings provide valuable insights into the molecular mechanisms and potential therapeutic targets in BLCA from the perspective of cell cycle. Moreover, NCAM1 was a novel cell proliferation suppressor in the BLCA carcinogenesis.

Whole transcriptome sequencing identifies a competitive endogenous RNA network that regulates the immunity of bladder cancer

Several types of non-coding RNAs such as circRNAs, lncRNAs, and miRNAs have been identified to regulate mRNAs through the mechanism known as the competitive endogenous RNA (ceRNA) network. To explore the role of the ceRNA regulatory network in the immune microenvironment of bladder cancer, whole-transcriptome sequencing of bladder tumor and its peritumoral tissues from 38 bladder cancer patients, with a total of 63 samples, was performed to screen differentially expressed circ-, lnc-, mi-, and mRNAs to construct a circ/lnc-mi-mRNA regulatory network with pruning algorithms. We excavated a key immune-related gene BDNF to build the final ceRNA network as hsa-miR-107 sponged by hsa-circ-000211, AC108488.1, and LINC00163. Finally, a meta-analysis of 7 public datasets demonstrated that low expression of BDNF and high expression of hsa-miR-107 were associated with longer survival. Our study identified a ceRNA regulatory network as a potentially new prognostic marker and molecular therapeutic target of bladder cancer.

USP15 facilitates the progression of bladder cancer by amplifying the activation of the NF-κB signaling pathway.

USP15, a pivotal member of the deubiquitinase family, plays a crucial role in orchestrating numerous vital biological processes, including the regulation of NF-κB signaling pathway and deubiquitination of proto-oncogenes. In various cancers, USP15 has been validated to exhibit up-regulated expression, impacting the initiation and progression of cancer. However, its precise mechanism in bladder cancer remains elusive. Our study shed light on the significant overexpression of USP15 in bladder cancer cells compared to normal bladder cells, correlating with a poorer prognosis for bladder cancer patients. Strikingly, attenuation of USP15 expression greatly attenuated the proliferation, migration, and invasion of bladder cancer cells. Moreover, upregulation of USP15 was found to drive cancer progression through the activation of the NF-κB signaling pathway. Notably, USP15 directly deubiquitinates BRCC3, heightening its expression level, and subsequent overexpression of BRCC3 counteracted the antitumoral efficacy of USP15 downregulation. Overall, our findings elucidated the carcinogenic effects of USP15 in bladder cancer, primarily mediated by the excessive activation of the NF-κB signaling pathway, thereby promoting tumor development. These results underscore the potential of USP15 as a promising therapeutic target for bladder cancer in the future.

CircITGA7 regulates malignant phenotypes in bladder cancer cells via targeting miR-330-3p/KLF10 axis.

Bladder cancer (BCa) is one of the common malignancies. Circular RNAs (circRNAs) play regulatory roles in cancer progression. CircITGA7 is a circRNA generated from several exons of ITGA7. The potential role of circITGA7 in BCa remains unknown and needs to be explored. Quantitative real time polymerase chain reaction (qRT-PCR) was used to assess circITGA7 and miR-330-3p expression in BCa tissues and cell lines. Kaplan-Meier analysis was used to evaluate the overall survival of these BCa patients. The biological function of circITGA7 was examined by overexpression of circITGA7 using CCK-8, EdU, wound-healing, and Transwell assays. Xenograft assay was performed to further validate the in vitro results. To explore the mechanism of circITGA7, luciferase reporter, RNA pull-down, fluorescence in situ hybridization (FISH) assays were employed to examine the binding interaction among circITGA7, miR-330-3p and kruppel-like factor 10 (KLF10). Western blot was used to study the protein levels of KLF10.CircITGA7 was downregulated in BCa tissues and cell lines and indicated longer overall survival. Moreover, circITGA7 restricted cell proliferation, migration and invasion of BCa through negatively regulating miR-330-3p. The in vivo model showed that circITGA7 influenced the tumor growth. Besides, the overexpression of miR-330-3p promoted cell progression by directly targeting KLF10. Mechanistically, circITGA7 inhibited BCa progression by activating KLF10 via targeting miR-330-3p.CircITGA7 alleviates BCa cell progression via circITGA7/hsa-miR-330-3p/KLF10 axis, which may provide novel therapeutic targets for BCa.

The mechanism of the contribution of ICAM-1 to epithelial-mesenchymal transition (EMT) in bladder cancer.

Bladder cancer is one of the most prevalent cancers worldwide. Moreover, if not optimally treated, bladder cancer is a significant burden on healthcare systems due to multiple recurrences which often require more aggressive therapies. Therefore, targeted anti-cancer therapies, developed based on an in-depth understanding of specific proteins and molecular mechanisms, are promising in cancer treatment. Here, for the first time, we presented the new approaches indicating that intracellular adhesion molecule-1 (ICAM-1) may play a potential role in enhancing therapeutic effectiveness for bladder cancer. In the present study, we presented that ICAM-1 expression as well as its regulation in bladder cancer is strongly correlated with the high expression of N-cadherin. Importantly, the presence of N-cadherin and its regulator-TWIST-1 was abolished when ICAM-1 was silenced. We identified also that ICAM-1 is capable of regulating cellular migration, proliferation, and EMT progression in bladder cancer cells via the N-cadherin/SRC/AKT/GSK-3β/β-catenin signaling axis. Therefore, we propose ICAM-1 as a novel metastatic marker for EMT progression, which may also be used as a therapeutic target in bladder cancer.

Abstract 506: TRIP13 is a promising therapeutic target for bladder cancer

Abstract Bladder cancer (BCa) is the second most common genitourinary cancer. High recurrence rate, progression to MIBC and poor prognosis in metastatic disease are major challenges for this disease management. Therapeutic options for advanced stage BCa are very limited, hence developing agents to treat this disease progression are highly needed. Thyroid receptor-interacting protein 13 (TRIP13), a member of the AAA-ATPase family, regulates various cellular processes such as DNA repair and spindle assembly. TCGA data analysis showed significant overexpression of TRIP13 that correlates with disease stage and mortality. Recent studies have shown that TRIP13 amplification occurs in several types of human cancers, including BCa, and it regulates cell proliferation, tumor growth, and drug resistance. However, the role of TRIP13 in BCa and its potential as a therapeutic target remain elusive. Therefore, in the present study we performed a preclinical investigation of the TRIP13 role in BCa progression, underlying molecular mechanisms, and develop promising therapeutics against BCa. Initially we profiled TRIP13 expression in 10 different bladder cancer cell lines from a panel of human and rodent origin and found elevated protein expression in multiple BCa cell lines. TRIP13 levels were also significantly elevated in the BBN induced rat- and SV40T driven mouse- bladder tumor tissues compared to normal bladder tissues. Pharmacological inhibition of TRIP13 with a potent drug DCZ0415 significantly suppressed cell proliferation (IC50 ~7μM) in J82, NBT-II and MB49 cell lines and induced cell death. DCZ0415 treatment led to cell cycle arrest at G1 phase (P<0.001) in BCa cells. In mechanistic investigations on its effect on cell cycle progression by DCZ0415 treatment decreased the expression of CDK4, CDK6, Cyclin D1, and PCNA protein expression. TRIP13 inhibition also suppressed the migration and invasion potential of BCa cells. Annexin V and γ-H2AX staining suggested that DCZ0415 induced apoptosis in BCa cells and increased DNA damage. These findings were also complemented by an increase in cleaved caspase-3 and PARP proteins. We also observed modulation of cGAS-STING signaling pathway in BCa cells with DCZ0415 treatment, indicating potential for its combination with cGAS-STING stimulators. Importantly, we found that TRIP13 inhibition with DCZ0415 significantly suppressed the growth of 3D BCa models as well BBN-induced rat BCa tumoroids and spheroids. Overall, these results suggest that TRIP13 could be a potential therapeutic target for prevention of bladder cancer and warrants further validation in in vivo models (Funding supported by P30CA225520 and ACS-IRG grant). Citation Format: Surya Pratap Singh, Krishnendu Goswami, Adam S. Asch, Chinthalapally V. Rao, Venkateshwar Madka. TRIP13 is a promising therapeutic target for bladder cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 506.

Unveiling the role of YARS1 in bladder cancer: A prognostic biomarker and therapeutic target.

YARS is responsible for catalysing the binding of tyrosine to its cognate tRNA and plays a crucial role in basic biosynthesis. However, its biological functions in bladder cancer remains to be proven. We analysed variations in YARS1 expression and survival in bladder cancer using multiple data sets, including TCGA-BLCA, GSE13507 and bladder cancer-specific tissue microarrays. Furthermore, we explored the biological functions of YARS1 using transcriptome data. Our findings revealed a noteworthy correlation between YARS1 and immune infiltration in bladder cancer, as determined using the XCELL algorithm and single-cell analysis. In addition, we employed the TIDE algorithm to evaluate the responsiveness of different cohorts to immune checkpoint therapy. We investigated the regulatory associations between YARS1 and various aspects of bladder cancer, including senescence, ferroptosis and stemness. Finally, we established a ceRNA network that is directly linked to the overall prognosis, YARS1 can serve as a prognostic biomarker for bladder cancer; its interaction with MYC has implications for bladder cancer cell senescence, ferroptosis and stemness. Moreover, the identified ceRNA network has potential as a therapeutic target in bladder cancer.

CircRNA LOC729852 promotes bladder cancer progression by regulating macrophage polarization and recruitment via the miR-769-5p/IL-10 axis.

Circular RNAs (circRNAs) function as tumour promoters or suppressors in bladder cancer (BLCA) by regulating genes involved in macrophage recruitment and polarization. However, the underlying mechanisms are largely unknown. The aim of this study was to determine the biological role of circLOC729852 in BLCA. CircLOC729852 was upregulated in BLCA tissues and correlated with increased proliferation, migration and epithelial mesenchymal transition (EMT) of BCLA cells. MiR-769-5p was identified as a target for circLOC729852, which can upregulate IL-10 expression by directly binding to and suppressing miR-769-5p. Furthermore, our results indicated that the circLOC729852/miR-769-5p/IL-10 axis modulates autophagy signalling in BLCA cells and promotes the recruitment and M2 polarization of TAMs by activating the JAK2/STAT3 signalling pathway. In addition, circLOC729852 also promoted the growth of BLCA xenografts and M2 macrophage infiltration invivo. Thus, circLOC729852 functions as an oncogene in BLCA by inducing secretion of IL-10 by the M2 TAMs, which then facilitates tumour cell growth and migration. Taken together, circLOC729852 is a potential diagnostic biomarker and therapeutic target for BLCA.

Parkin inhibits proliferation and migration of bladder cancer via ubiquitinating Catalase

PRKN is a key gene involved in mitophagy in Parkinson’s disease. However, recent studies have demonstrated that it also plays a role in the development and metastasis of several types of cancers, both in a mitophagy-dependent and mitophagy-independent manner. Despite this, the potential effects and underlying mechanisms of Parkin on bladder cancer (BLCA) remain unknown. Therefore, in this study, we investigated the expression of Parkin in various BLCA cohorts derived from human. Here we show that PRKN expression was low and that PRKN acts as a tumor suppressor by inhibiting the proliferation and migration of BLCA cells in a mitophagy-independent manner. We further identified Catalase as a binding partner and substrate of Parkin, which is an important antioxidant enzyme that regulates intracellular ROS levels during cancer progression. Our data showed that knockdown of CAT led to increased intracellular ROS levels, which suppressed cell proliferation and migration. Conversely, upregulation of Catalase decreased intracellular ROS levels, promoting cell growth and migration. Importantly, we found that Parkin upregulation partially restored these effects. Moreover, we discovered that USP30, a known Parkin substrate, could deubiquitinate and stabilize Catalase. Overall, our study reveals a novel function of Parkin and identifies a potential therapeutic target in BLCA.

TFRC, associated with hypoxia and immune, is a prognostic factor and potential therapeutic target for bladder cancer

BackgroundBladder cancer is a common malignancy of the urinary system, and the survival rate and recurrence rate of patients with muscular aggressive (MIBC) bladder cancer are not ideal. Hypoxia is a pathological process in which cells acquire special characteristics to adapt to anoxic environment, which can directly affect the proliferation, invasion and immune response of bladder cancer cells. Understanding the exact effects of hypoxia and immune-related genes in BLCA is helpful for early assessment of the prognosis of BLCA. However, the prognostic model of BLCA based on hypoxia and immune-related genes has not been reported.PurposeHypoxia and immune cell have important role in the prognosis of bladder cancer (BLCA). The aim of this study was to investigate whether hypoxia and immune related genes could be a novel tools to predict the overall survival and immunotherapy of BLCA patients.MethodsFirst, we downloaded transcriptomic data and clinical information of BLCA patients from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases. A combined hypoxia and immune signature was then constructed on the basis of the training cohort via least absolute shrinkage and selection operator (LASSO) analysis and validated in test cohort. Afterwards, Kaplan–Meier curves, univariate and multivariate Cox and subgroup analysis were employed to assess the accuracy of our signature. Immune cell infiltration, checkpoint and the Tumor Immune Dysfunction and Exclusion (TIDE) algorithm were used to investigate the immune environment and immunotherapy of BLCA patients. Furthermore, we confirmed the role of TFRC in bladder cancer cell lines T24 and UMUC-3 through cell experiments.ResultsA combined hypoxia and immune signature containing 8 genes were successfully established. High-risk group in both training and test cohorts had significantly poorer OS than low-risk group. Univariate and multivariate Cox analysis indicated our signature could be regarded as an independent prognostic factor. Different checkpoint was differently expressed between two groups, including CTLA4, HAVCR2, LAG3, PD-L1 and PDCD1. TIDE analysis indicated high-risk patients had poor response to immunotherapy and easier to have immune escape. The drug sensitivity analysis showed that high-risk group patients were more potentially sensitive to many drugs. Meanwhile, TFRC could inhibit the proliferation and invasion ability of T24 and UMUC-3 cells.ConclusionA combined hypoxia and immune-related gene could be a novel predictive model for OS and immunotherapy estimation of BLCA patients and TFRC could be used as a potential therapeutic target in the future.

A study of blood myeloid-derived suppressor cells for predicting pembrolizumab efficacy in patients with metastatic urothelial carcinoma.

658 Background: Immune checkpoint inhibitors (ICIs) are crucial for treating metastatic urothelial carcinoma (mUC) patients, yet current prognostic biomarkers fail to predict ICI efficacy. Previous preclinical models have suggested myeloid-derived suppressor cells (MDSCs) as a potential therapeutic target in bladder cancer. This study aims to evaluate the predictive value of blood MDSCs in mUC patients undergoing pembrolizumab treatment. Methods: Ninety-eight mUC patients who progressed with platinum-based chemotherapy at Osaka Metropolitan University received pembrolizumab. Among them, 30 patients were assessed for blood MDSCs. Peripheral blood samples were collected from healthy individuals (n = 6) and mUC patients (n = 30) before pembrolizumab initiation. The mononuclear cell layer was isolated using density gradient centrifugation for MDSC analysis through flow cytometry. Monocytic and polymorphonuclear MDSCs were defined as CD11b+ HLADR – CD33 high CD14 + CD15 – and CD11b+ HLADR – CD33 low CD14 – CD15 +, respectively. The neutrophil-lymphocyte ratio (NLR) in blood served as a control. Treatment response was assessed via CT images using RECICT v1.1. Progression-free survival (PFS) and overall survival (OS) were calculated using the Kaplan-Meier method, with log-rank test for between-group comparison. Results: First, blood MDSCs were significantly elevated in mUC patients compared to healthy controls (p = 0.0024, Mann Whitney test). Secondly, patients with MDSClow (≤ median, n = 15) had significantly longer PFS than those with MDSChigh (> median, n = 15) (17.6 vs. 1.9 months, p = 0.0126). However, no significant difference was observed in OS. In contrast, when the NLR cutoff was set at 3.5, no significant association was found for PFS and OS. Conclusions: Blood MDSCs show promise as a predictive marker for pembrolizumab therapy efficacy.

Potential clinical application of microRNAs in bladder cancer.

Bladder cancer (BC) is the tenth most prevalent malignancy globally, presenting significant clinical and societal challenges because of its high incidence, rapid progression, and frequent recurrence. Presently, cystoscopy and urine cytology serve as the established diagnostic methods for BC. However, their efficacy is limited by invasive nature and low sensitivity. Therefore, the development of highly specific biomarkers and effective non-invasive detection strategies is imperative for achieving a precise and timely diagnosis of BC, as well as for facilitating an optimal tumor treatment and an improved prognosis. microRNAs (miRNAs), short noncoding RNA molecules spanning around 20-25 nucleotides, are implicated in the regulation of diverse carcinogenic pathways. Substantially altered miRNAs form robust functional regulatory networks that exert a notable influence on the tumorigenesis and progression of BC. Investigations into aberrant miRNAs derived from blood, urine, or extracellular vesicles indicate their potential roles as diagnostic biomarkers and prognostic indicators in BC, enabling miRNAs to monitor the progression and predict the recurrence of the disease. Simultaneously, the investigation centered on miRNA as a potential therapeutic agent presents a novel approach for the treatment of BC. This review provides a comprehensive analysis of the biological roles of miRNAs in tumorigenesis and progression, and systematically summarizes the potential as diagnosis and prognosis biomarkers as well as therapeutic targets for BC. Additionally, we evaluate the progress made in laboratory techniques within this field and discuss the prospects.

USP5 facilitates bladder cancer progression by stabilizing the c-Jun protein

BackgroundBladder cancer is the second most common genitourinary malignancy worldwide. The death rate of bladder cancer has increased every year. However, the molecular mechanism of bladder cancer is not sufficiently studied. Deubiquitinating enzymes (DUBs) play an important role in carcinogenesis. Several studies have demonstrated that USP5 associated with malignancy and pathological progression in hepatocellular carcinoma, colorectal and non-small cell lung cancer. However, the role of USP5 in bladder cancer need to be explored.MethodsThe USP5 expression was analysed using the web server GEPIA. To explore USP5 function in bladder cancer, we constructed USP5-knockout cell lines in T24 cells. A FLAG-USP5 (WT USP5) plasmid and a plasmid FLAG-USP5 C335A (catalytic-inactive mutant) used to overexpress USP5 in EJ cells. CCK8, colony formation, transwell and scratch assays were used to assess cell viability, proliferation and migration. RNA sequencing (RNA-seq) and dual-luciferase reporter assays were performed to screen the pathway. Coimmunoprecipitation and immunofluorescence were used to explore the interaction between USP5 and c-Jun. Cycloheximide (CHX) chase assays were performed to establish the effect of USP5 on c-Jun stability. Xenograft mouse model was used to study the role of USP5 in bladder cancer.ResultsUSP5 expression is increased in bladder cancer patients. Genetic ablation of USP5 markedly inhibited bladder cancer cell proliferation, viability, and migration both in vitro and in vivo. RNA-seq and luciferase pathway screening showed that USP5 activated JNK signalling, and we identified the interaction between USP5 and c-Jun. USP5 was found to activate c-Jun by inhibiting its ubiquitination.ConclusionsOur results show that high USP5 expression promotes bladder cancer progression by stabilizing c-Jun and that USP5 is a potential therapeutic target in bladder cancer.

CLIC3 interacts with NAT10 to inhibit N4-acetylcytidine modification of p21 mRNA and promote bladder cancer progression

Chromatin accessibility plays important roles in revealing the regulatory networks of gene expression, while its application in bladder cancer is yet to be fully elucidated. Chloride intracellular channel 3 (CLIC3) protein has been reported to be associated with the progression of some tumors, whereas the specific mechanism of CLIC3 in tumor remains unclear. Here, we screened for key genes in bladder cancer through the identification of transcription factor binding site clustered region (TFCR) on the basis of chromatin accessibility and TF motif. CLIC3 was identified by joint profiling of chromatin accessibility data with TCGA database. Clinically, CLIC3 expression was significantly elevated in bladder cancer and was negatively correlated with patient survival. CLIC3 promoted the proliferation of bladder cancer cells by reducing p21 expression in vitro and in vivo. Mechanistically, CLIC3 interacted with NAT10 and inhibited the function of NAT10, resulting in the downregulation of ac4C modification and stability of p21 mRNA. Overall, these findings uncover an novel mechanism of mRNA ac4C modification and CLIC3 may act as a potential therapeutic target for bladder cancer.

Melanoma Antigen Family A (MAGE A) as Promising Biomarkers and Therapeutic Targets in Bladder Cancer.

The Melanoma Antigen Gene (MAGE) is a large family of highly conserved proteins that share a common MAGE homology domain. Interestingly, many MAGE family members exhibit restricted expression in reproductive tissues but are abnormally expressed in various human malignancies, including bladder cancer, which is a common urinary malignancy associated with high morbidity and mortality rates. The recent literature suggests a more prominent role for MAGEA family members in driving bladder tumorigenesis. This review highlights the role of MAGEA proteins, the potential for them to serve as diagnostic or prognostic biomarker(s), and as therapeutic targets for bladder cancer.