RESUMEN
Responses to anti-PD-1 immunotherapy occur but are infrequent in bladder cancer. The specific T cells that mediate tumor rejection are unknown. T cells from human bladder tumors and non-malignant tissue were assessed with single-cell RNA and paired T cell receptor (TCR) sequencing of 30,604 T cells from 7 patients. We find that the states and repertoires of CD8+ T cells are not distinct in tumors compared with non-malignant tissues. In contrast, single-cell analysis of CD4+ T cells demonstrates several tumor-specific states, including multiple distinct states of regulatory T cells. Surprisingly, we also find multiple cytotoxic CD4+ T cell states that are clonally expanded. These CD4+ T cells can kill autologous tumors in an MHC class II-dependent fashion and are suppressed by regulatory T cells. Further, a gene signature of cytotoxic CD4+ T cells in tumors predicts a clinical response in 244 metastatic bladder cancer patients treated with anti-PD-L1.
Asunto(s)
Linfocitos T CD4-Positivos/metabolismo , Neoplasias de la Vejiga Urinaria/genética , Neoplasias de la Vejiga Urinaria/metabolismo , Antígeno B7-H1/genética , Antígeno B7-H1/inmunología , Biomarcadores Farmacológicos/análisis , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/metabolismo , Regulación Neoplásica de la Expresión Génica/genética , Genes MHC Clase II , Humanos , Inhibidores de Puntos de Control Inmunológico/farmacología , Inmunoterapia , Linfocitos Infiltrantes de Tumor , Receptor de Muerte Celular Programada 1/genética , Receptores de Antígenos de Linfocitos T/genética , Análisis de la Célula Individual/métodos , Linfocitos T Reguladores , Neoplasias de la Vejiga Urinaria/inmunologíaRESUMEN
Heterotrimeric G-proteins (Gαßγ) are the main transducers of signals from GPCRs, mediating the action of countless natural stimuli and therapeutic agents. However, there are currently no robust approaches to directly measure the activity of endogenous G-proteins in cells. Here, we describe a suite of optical biosensors that detect endogenous active G-proteins with sub-second resolution in live cells. Using a modular design principle, we developed genetically encoded, unimolecular biosensors for endogenous Gα-GTP and free Gßγ: the two active species of heterotrimeric G-proteins. This design was leveraged to generate biosensors with specificity for different heterotrimeric G-proteins or for other G-proteins, such as Rho GTPases. Versatility was further validated by implementing the biosensors in multiple contexts, from characterizing cancer-associated G-protein mutants to neurotransmitter signaling in primary neurons. Overall, the versatile biosensor design introduced here enables studying the activity of endogenous G-proteins in live cells with high fidelity, temporal resolution, and convenience.
Asunto(s)
Transferencia de Energía por Resonancia de Bioluminiscencia/instrumentación , Transferencia de Energía por Resonancia de Bioluminiscencia/métodos , Técnicas Biosensibles/instrumentación , Técnicas Biosensibles/métodos , Guanosina Trifosfato/metabolismo , Proteínas de Unión al GTP Heterotriméricas/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Secuencias de Aminoácidos , Animales , Células Cultivadas , Subunidades alfa de la Proteína de Unión al GTP Gq-G11/química , Subunidades alfa de la Proteína de Unión al GTP Gq-G11/genética , Factores de Intercambio de Guanina Nucleótido/antagonistas & inhibidores , Guanosina Trifosfato/química , Células HEK293 , Células HeLa , Humanos , Ratones , Ratones Endogámicos C57BL , Mutación , Neoplasias/genética , Neoplasias/metabolismo , Neuronas/química , Neuronas/metabolismo , Neuronas/fisiología , Transducción de Señal , Neoplasias de la Vejiga Urinaria/genética , Neoplasias de la Vejiga Urinaria/metabolismoRESUMEN
We report a comprehensive analysis of 412 muscle-invasive bladder cancers characterized by multiple TCGA analytical platforms. Fifty-eight genes were significantly mutated, and the overall mutational load was associated with APOBEC-signature mutagenesis. Clustering by mutation signature identified a high-mutation subset with 75% 5-year survival. mRNA expression clustering refined prior clustering analyses and identified a poor-survival "neuronal" subtype in which the majority of tumors lacked small cell or neuroendocrine histology. Clustering by mRNA, long non-coding RNA (lncRNA), and miRNA expression converged to identify subsets with differential epithelial-mesenchymal transition status, carcinoma in situ scores, histologic features, and survival. Our analyses identified 5 expression subtypes that may stratify response to different treatments.
Asunto(s)
Neoplasias de la Vejiga Urinaria/genética , Neoplasias de la Vejiga Urinaria/patología , Anciano , Análisis por Conglomerados , Metilación de ADN , Humanos , MicroARNs/genética , Persona de Mediana Edad , Músculo Liso/patología , ARN Largo no Codificante/genética , Análisis de Supervivencia , Vejiga Urinaria/patología , Neoplasias de la Vejiga Urinaria/epidemiología , Neoplasias de la Vejiga Urinaria/terapiaRESUMEN
Loss of the Y chromosome (LOY) is observed in multiple cancer types, including 10-40% of bladder cancers1-6, but its clinical and biological significance is unknown. Here, using genomic and transcriptomic studies, we report that LOY correlates with poor prognoses in patients with bladder cancer. We performed in-depth studies of naturally occurring LOY mutant bladder cancer cells as well as those with targeted deletion of Y chromosome by CRISPR-Cas9. Y-positive (Y+) and Y-negative (Y-) tumours grew similarly in vitro, whereas Y- tumours were more aggressive than Y+ tumours in immune-competent hosts in a T cell-dependent manner. High-dimensional flow cytometric analyses demonstrated that Y- tumours promote striking dysfunction or exhaustion of CD8+ T cells in the tumour microenvironment. These findings were validated using single-nuclei RNA sequencing and spatial proteomic evaluation of human bladder cancers. Of note, compared with Y+ tumours, Y- tumours exhibited an increased response to anti-PD-1 immune checkpoint blockade therapy in both mice and patients with cancer. Together, these results demonstrate that cancer cells with LOY mutations alter T cell function, promoting T cell exhaustion and sensitizing them to PD-1-targeted immunotherapy. This work provides insights into the basic biology of LOY mutation and potential biomarkers for improving cancer immunotherapy.
Asunto(s)
Linfocitos T CD8-positivos , Deleción Cromosómica , Cromosomas Humanos Y , Escape del Tumor , Neoplasias de la Vejiga Urinaria , Animales , Humanos , Ratones , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/patología , Cromosomas Humanos Y/genética , Proteómica , Microambiente Tumoral/inmunología , Neoplasias de la Vejiga Urinaria/genética , Neoplasias de la Vejiga Urinaria/inmunología , Neoplasias de la Vejiga Urinaria/patología , Neoplasias de la Vejiga Urinaria/terapia , Escape del Tumor/genética , Escape del Tumor/inmunología , Perfilación de la Expresión Génica , Genómica , Pronóstico , Sistemas CRISPR-Cas , Edición Génica , Técnicas In Vitro , Inhibidores de Puntos de Control Inmunológico/farmacología , Inhibidores de Puntos de Control Inmunológico/uso terapéutico , Citometría de Flujo , InmunoterapiaRESUMEN
Drug resistance contributes to poor therapeutic response in urothelial carcinoma (UC). Metabolomic analysis suggested metabolic reprogramming in gemcitabine-resistant urothelial carcinoma cells, whereby increased aerobic glycolysis and metabolic stimulation of the pentose phosphate pathway (PPP) promoted pyrimidine biosynthesis to increase the production of the gemcitabine competitor deoxycytidine triphosphate (dCTP) that diminishes its therapeutic effect. Furthermore, we observed that gain-of-function of isocitrate dehydrogenase 2 (IDH2) induced reductive glutamine metabolism to stabilize Hif-1α expression and consequently stimulate aerobic glycolysis and PPP bypass in gemcitabine-resistant UC cells. Interestingly, IDH2-mediated metabolic reprogramming also caused cross resistance to CDDP, by elevating the antioxidant defense via increased NADPH and glutathione production. Downregulation or pharmacological suppression of IDH2 restored chemosensitivity. Since the expression of key metabolic enzymes, such as TIGAR, TKT, and CTPS1, were affected by IDH2-mediated metabolic reprogramming and related to poor prognosis in patients, IDH2 might become a new therapeutic target for restoring chemosensitivity in chemo-resistant urothelial carcinoma.
Asunto(s)
Carcinoma de Células Transicionales , Neoplasias de la Vejiga Urinaria , Humanos , Línea Celular Tumoral , Resistencia a Antineoplásicos/genética , Gemcitabina , Glucólisis , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Vía de Pentosa Fosfato , Neoplasias de la Vejiga Urinaria/tratamiento farmacológico , Neoplasias de la Vejiga Urinaria/genéticaRESUMEN
Minimally invasive approaches to detect residual disease after surgery are needed to identify patients with cancer who are at risk for metastatic relapse. Circulating tumour DNA (ctDNA) holds promise as a biomarker for molecular residual disease and relapse1. We evaluated outcomes in 581 patients who had undergone surgery and were evaluable for ctDNA from a randomized phase III trial of adjuvant atezolizumab versus observation in operable urothelial cancer. This trial did not reach its efficacy end point in the intention-to-treat population. Here we show that ctDNA testing at the start of therapy (cycle 1 day 1) identified 214 (37%) patients who were positive for ctDNA and who had poor prognosis (observation arm hazard ratio = 6.3 (95% confidence interval: 4.45-8.92); P < 0.0001). Notably, patients who were positive for ctDNA had improved disease-free survival and overall survival in the atezolizumab arm versus the observation arm (disease-free survival hazard ratio = 0.58 (95% confidence interval: 0.43-0.79); P = 0.0024, overall survival hazard ratio = 0.59 (95% confidence interval: 0.41-0.86)). No difference in disease-free survival or overall survival between treatment arms was noted for patients who were negative for ctDNA. The rate of ctDNA clearance at week 6 was higher in the atezolizumab arm (18%) than in the observation arm (4%) (P = 0.0204). Transcriptomic analysis of tumours from patients who were positive for ctDNA revealed higher expression levels of cell-cycle and keratin genes. For patients who were positive for ctDNA and who were treated with atezolizumab, non-relapse was associated with immune response signatures and basal-squamous gene features, whereas relapse was associated with angiogenesis and fibroblast TGFß signatures. These data suggest that adjuvant atezolizumab may be associated with improved outcomes compared with observation in patients who are positive for ctDNA and who are at a high risk of relapse. These findings, if validated in other settings, would shift approaches to postoperative cancer care.
Asunto(s)
Adyuvantes Farmacéuticos/uso terapéutico , Anticuerpos Monoclonales Humanizados/uso terapéutico , ADN Tumoral Circulante/sangre , Inmunoterapia , Neoplasias de la Vejiga Urinaria/diagnóstico , Neoplasias de la Vejiga Urinaria/tratamiento farmacológico , Biomarcadores de Tumor/sangre , Biomarcadores de Tumor/genética , ADN Tumoral Circulante/genética , Regulación Neoplásica de la Expresión Génica , Humanos , Estimación de Kaplan-Meier , Recurrencia Local de Neoplasia/sangre , Recurrencia Local de Neoplasia/genética , Cuidados Posoperatorios , Pronóstico , Recurrencia , Análisis de Supervivencia , Neoplasias de la Vejiga Urinaria/genética , Neoplasias de la Vejiga Urinaria/inmunologíaRESUMEN
Inferring gene regulatory network (GRN) is one of the important challenges in systems biology, and many outstanding computational methods have been proposed; however there remains some challenges especially in real datasets. In this study, we propose Directed Graph Convolutional neural network-based method for GRN inference (DGCGRN). To better understand and process the directed graph structure data of GRN, a directed graph convolutional neural network is conducted which retains the structural information of the directed graph while also making full use of neighbor node features. The local augmentation strategy is adopted in graph neural network to solve the problem of poor prediction accuracy caused by a large number of low-degree nodes in GRN. In addition, for real data such as E.coli, sequence features are obtained by extracting hidden features using Bi-GRU and calculating the statistical physicochemical characteristics of gene sequence. At the training stage, a dynamic update strategy is used to convert the obtained edge prediction scores into edge weights to guide the subsequent training process of the model. The results on synthetic benchmark datasets and real datasets show that the prediction performance of DGCGRN is significantly better than existing models. Furthermore, the case studies on bladder uroepithelial carcinoma and lung cancer cells also illustrate the performance of the proposed model.
Asunto(s)
Biología Computacional , Redes Reguladoras de Genes , Redes Neurales de la Computación , Humanos , Biología Computacional/métodos , Algoritmos , Neoplasias de la Vejiga Urinaria/genética , Neoplasias de la Vejiga Urinaria/patología , Escherichia coli/genéticaRESUMEN
Micro ribonucleic acids (miRNAs) play a pivotal role in governing the human transcriptome in various biological phenomena. Hence, the accumulation of miRNA expression dysregulation frequently assumes a noteworthy role in the initiation and progression of complex diseases. However, accurate identification of dysregulated miRNAs still faces challenges at the current stage. Several bioinformatics tools have recently emerged for forecasting the associations between miRNAs and diseases. Nonetheless, the existing reference tools mainly identify the miRNA-disease associations in a general state and fall short of pinpointing dysregulated miRNAs within a specific disease state. Additionally, no studies adequately consider miRNA-miRNA interactions (MMIs) when analyzing the miRNA-disease associations. Here, we introduced a systematic approach, called IDMIR, which enabled the identification of expression dysregulated miRNAs through an MMI network under the gene expression context, where the network's architecture was designed to implicitly connect miRNAs based on their shared biological functions within a particular disease context. The advantage of IDMIR is that it uses gene expression data for the identification of dysregulated miRNAs by analyzing variations in MMIs. We illustrated the excellent predictive power for dysregulated miRNAs of the IDMIR approach through data analysis on breast cancer and bladder urothelial cancer. IDMIR could surpass several existing miRNA-disease association prediction approaches through comparison. We believe the approach complements the deficiencies in predicting miRNA-disease association and may provide new insights and possibilities for diagnosing and treating diseases. The IDMIR approach is now available as a free R package on CRAN (https://CRAN.R-project.org/package=IDMIR).
Asunto(s)
Biología Computacional , Redes Reguladoras de Genes , MicroARNs , Neoplasias de la Vejiga Urinaria , Humanos , MicroARNs/genética , MicroARNs/metabolismo , Biología Computacional/métodos , Neoplasias de la Vejiga Urinaria/genética , Neoplasias de la Vejiga Urinaria/metabolismo , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Perfilación de la Expresión Génica , Femenino , Regulación Neoplásica de la Expresión GénicaRESUMEN
Molecular profiling studies have shown that 85% of canine urothelial carcinomas (UC) harbor an activating BRAF V595E mutation, which is orthologous to the V600E variant found in several human cancer subtypes. In dogs, this mutation provides both a powerful diagnostic marker and a potential therapeutic target; however, due to their relative infrequency, the remaining 15% of cases remain understudied at the molecular level. We performed whole exome sequencing analysis of 28 canine urine sediments exhibiting the characteristic DNA copy number signatures of canine UC, in which the BRAF V595E mutation was undetected (UDV595E specimens). Among these we identified 13 specimens (46%) harboring short in-frame deletions within either BRAF exon 12 (7/28 cases) or MAP2K1 exons 2 or 3 (6/28 cases). Orthologous variants occur in several human cancer subtypes and confer structural changes to the protein product that are predictive of response to different classes of small molecule MAPK pathway inhibitors. DNA damage response and repair genes, and chromatin modifiers were also recurrently mutated in UDV595E specimens, as were genes that are positive predictors of immunotherapy response in human cancers. Our findings suggest that short in-frame deletions within BRAF exon 12 and MAP2K1 exons 2 and 3 in UDV595E cases are alternative MAPK-pathway activating events that may have significant therapeutic implications for selecting first-line treatment for canine UC. We developed a simple, cost-effective capillary electrophoresis genotyping assay for detection of these deletions in parallel with the BRAF V595E mutation. The identification of these deletion events in dogs offers a compelling cross-species platform in which to study the relationship between somatic alteration, protein conformation, and therapeutic sensitivity.
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Carcinoma de Células Transicionales , MAP Quinasa Quinasa 1 , Proteínas Proto-Oncogénicas B-raf , Neoplasias de la Vejiga Urinaria , Animales , Perros , Secuenciación del Exoma , Proteínas Proto-Oncogénicas B-raf/genética , MAP Quinasa Quinasa 1/genética , Neoplasias de la Vejiga Urinaria/genética , Neoplasias de la Vejiga Urinaria/veterinaria , Carcinoma de Células Transicionales/genética , Carcinoma de Células Transicionales/veterinaria , Sistema de Señalización de MAP Quinasas , Variaciones en el Número de Copia de ADN , Eliminación de Secuencia , Masculino , FemeninoRESUMEN
Bladder cancer is a urothelial cancer and effective therapeutic strategies for its advanced stages are limited. Here, we report that CD271, a neurotrophin receptor, promotes the proliferation and migration of bladder cancer cells. CD271 knockdown decreased proliferation in both adherent and spheroid cultures, and vice versa when CD271 was overexpressed in bladder cancer cell lines. CD271 depletion impaired tumorigenicity in vivo. Migration activity was reduced by CD271 knockdown and TAT-Pep5, a known CD271-Rho GDI-binding inhibitor. Apoptosis was induced by CD271 knockdown. Comprehensive gene expression analysis revealed alterations in E2F- and Myc-related pathways upon CD271 expression. In clinical cases, patients with high CD271 expression showed significantly shortened overall survival. In surgically resected specimens, pERK, a known player in proliferation signaling, colocalizes with CD271. These data indicate that CD271 is involved in bladder cancer malignancy by promoting cell proliferation and migration, resulting in poor prognosis.
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Receptores de Factor de Crecimiento Nervioso , Neoplasias de la Vejiga Urinaria , Humanos , Adapaleno , Receptores de Factor de Crecimiento Nervioso/genética , Proliferación Celular , Transducción de Señal , Neoplasias de la Vejiga Urinaria/genética , Movimiento Celular , Línea Celular Tumoral , Regulación Neoplásica de la Expresión GénicaRESUMEN
Interactions between Tumor microenvironment (TME) cells shape the unique growth environment, sustaining tumor growth and causing the immune escape of tumor cells. Nonetheless, no studies have reported a systematic analysis of cellular interactions in the identification of cancer-related TME cells. Here, we proposed a novel network-based computational method, named as iATMEcell, to identify the abnormal TME cells associated with the biological outcome of interest based on a cell-cell crosstalk network. In the method, iATMEcell first manually collected TME cell types from multiple published studies and obtained their corresponding gene signatures. Then, a weighted cell-cell crosstalk network was constructed in the context of a specific cancer bulk tissue transcriptome data, where the weight between cells reflects both their biological function similarity and the transcriptional dysregulated activities of gene signatures shared by them. Finally, it used a network propagation algorithm to identify significantly dysregulated TME cells. Using the cancer genome atlas (TCGA) Bladder Urothelial Carcinoma training set and two independent validation sets, we illustrated that iATMEcell could identify significant abnormal cells associated with patient survival and immunotherapy response. iATMEcell was further applied to a pan-cancer analysis, which revealed that four common abnormal immune cells play important roles in the patient prognosis across multiple cancer types. Collectively, we demonstrated that iATMEcell could identify potentially abnormal TME cells based on a cell-cell crosstalk network, which provided a new insight into understanding the effect of TME cells in cancer. iATMEcell is developed as an R package, which is freely available on GitHub (https://github.com/hanjunwei-lab/iATMEcell).
Asunto(s)
Carcinoma de Células Transicionales , Neoplasias de la Vejiga Urinaria , Humanos , Neoplasias de la Vejiga Urinaria/genética , Microambiente Tumoral , Fenómenos Fisiológicos Celulares , Comunicación CelularRESUMEN
The emergence of mutational signatures in cancer research represents a paradigm shift, offering profound insights into tumor biology, therapeutic strategies, and risk assessment. A recent article published in The Journal of Pathology delves into the significance of mutational signatures, assessing their role as molecular fingerprints that illuminate the etiology of individual cancer cases. By deciphering the diverse patterns of DNA alterations, in the context of urachal and nonurachal bladder adenocarcinomas, researchers can unravel intrinsic and environmental risk factors shaping cancer development. The study highlighted a predominance of environmental risk factors in Chinese populations affected by bladder adenocarcinomas. In this context, in this commentary, we highlight some of the potential applications of molecular signatures in differential diagnosis and therapy prediction. As the field continues to evolve, a deeper understanding of mutational signatures promises to revolutionize precision oncology by offering personalized approaches informed by comprehensive mutational patterns of tumors. © 2024 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.
Asunto(s)
Adenocarcinoma , Mutación , Neoplasias de la Vejiga Urinaria , Humanos , Neoplasias de la Vejiga Urinaria/genética , Neoplasias de la Vejiga Urinaria/patología , Adenocarcinoma/genética , Adenocarcinoma/patología , Biomarcadores de Tumor/genética , Análisis Mutacional de ADN , Predisposición Genética a la EnfermedadRESUMEN
TP53 mutation is one of the most common genetic alterations in urothelial carcinoma (UrCa), and heterogeneity of TP53 mutants leads to heterogeneous clinical outcomes. This study aimed to investigate the clinical relevance of specific TP53 mutations in UrCa. In this study, a total of eight cohorts were enrolled, along with matched clinical annotation. TP53 mutations were classified as disruptive and nondisruptive according to the degree of disturbance of p53 protein function and structure. We evaluated the clinical significance of TP53 mutations in our local datasets and publicly available datasets. The co-occurring events of TP53 mutations in UrCa, along with their therapeutic indications, functional effects, and the tumor immune microenvironment, were also investigated. TP53 mutations were identified in 49.7% of the UrCa patients. Within this group, 25.1% of patients carried TP53Disruptive mutations, a genetic alteration correlated with a significantly poorer overall survival (OS) when compared to individuals with TP53Nondisruptive mutations and those with wild-type TP53. Significantly, patients with TP53Disruptive mutations exhibit an increased probability of responding favorably to PD-1/PD-L1 blockade and chemoimmunotherapy. Meanwhile, there was no noteworthy distinction in OS among patients with varying TP53 mutation status who underwent chemotherapy. Samples with TP53Disruptive mutations showed an enriched APOBEC- and POLE-related mutational signature, as well as an elevated tumor mutation burden. The sensitivity to immunotherapy in tumors carrying TP53Disruptive mutation may be attributed to the inflamed tumor microenvironment characterized by increased CD8+T cell infiltration and interferon-gamma signaling activation. In conclusion, UrCa patients with TP53Disruptive mutations have shown reduced survival rates, yet they may respond well to PD-1/PD-L1 blockade therapy and chemoimmunotherapy. By distinguishing specific TP53 mutations, we can improve risk stratification and offer personalized genomics-guided therapy to UrCa patients. © 2024 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.
Asunto(s)
Antígeno B7-H1 , Inhibidores de Puntos de Control Inmunológico , Mutación , Receptor de Muerte Celular Programada 1 , Microambiente Tumoral , Proteína p53 Supresora de Tumor , Neoplasias de la Vejiga Urinaria , Humanos , Proteína p53 Supresora de Tumor/genética , Inhibidores de Puntos de Control Inmunológico/uso terapéutico , Antígeno B7-H1/antagonistas & inhibidores , Antígeno B7-H1/genética , Receptor de Muerte Celular Programada 1/antagonistas & inhibidores , Receptor de Muerte Celular Programada 1/genética , Neoplasias de la Vejiga Urinaria/genética , Neoplasias de la Vejiga Urinaria/tratamiento farmacológico , Neoplasias de la Vejiga Urinaria/patología , Neoplasias de la Vejiga Urinaria/inmunología , Carcinoma de Células Transicionales/tratamiento farmacológico , Carcinoma de Células Transicionales/genética , Carcinoma de Células Transicionales/inmunología , Carcinoma de Células Transicionales/patología , Carcinoma de Células Transicionales/mortalidad , Biomarcadores de Tumor/genética , Masculino , Femenino , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Anciano , Persona de Mediana EdadRESUMEN
Bladder cancer(BC) is one of the most prevalent cancers in the urinary tract, with high recurrence and fatality rates. Research indicates that go-ichi-ni-san complex subunit 1 (GINS1) crucially influences cancer progression by regulating DNA replication through cell cycle modulation. Thus, suppressing the active proliferation of cells in tumor tissues may require silencing GINS1. However, the consequences of GINS1 in bladder cancer aren't to be determined. In this paper, we examine the role and mechanism of GINS1 in the development of bladder cancer. GINS1 expression levels and prognostic relevance in bladder cancer were validated using Western blotting, immunohistochemistry, and Kaplan-Meier survival analysis. The influence of GINS1 on bladder cancer was investigated using a variety of approaches, including cell transfection, cell counts, transwell migrations, colony formation, and flow cytometry. Immunohistochemistry studies demonstrate that GINS1 expression is increased in bladder cancer tissues. GINS1 silencing resulted in an arrest of the cell cycle at the phase of G0/G1, which inhibited BC cell growth both in vitro and in vivo. GINS1 knockdown also hindered the AKT/mTOR pathway. Furthermore, increased GINS1 expression affects the cell cycle and stimulates the AKT/mTOR pathway, allowing BC to develop more quickly. Consequently, GINS1 occurs as a latent therapeutic target, particularly for individuals with BC.
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Proliferación Celular , Proteínas Cromosómicas no Histona , Proteínas Proto-Oncogénicas c-akt , Transducción de Señal , Serina-Treonina Quinasas TOR , Neoplasias de la Vejiga Urinaria , Neoplasias de la Vejiga Urinaria/patología , Neoplasias de la Vejiga Urinaria/genética , Neoplasias de la Vejiga Urinaria/metabolismo , Humanos , Proteínas Proto-Oncogénicas c-akt/metabolismo , Proteínas Proto-Oncogénicas c-akt/genética , Serina-Treonina Quinasas TOR/metabolismo , Serina-Treonina Quinasas TOR/genética , Proteínas Cromosómicas no Histona/metabolismo , Proteínas Cromosómicas no Histona/genética , Proliferación Celular/genética , Animales , Línea Celular Tumoral , Proteínas Proto-Oncogénicas c-myc/metabolismo , Proteínas Proto-Oncogénicas c-myc/genética , Regulación Neoplásica de la Expresión Génica , Ratones , Progresión de la Enfermedad , Ratones Desnudos , Masculino , Femenino , Pronóstico , Ratones Endogámicos BALB C , Proteínas de Unión al ADNRESUMEN
Bladder cancer is a common tumor that impacts the urinary system and marked by a significant fatality rate and an unfavorable prognosis. Promising antineoplastic properties are exhibited by brusatol, which is obtained from the dried ripe fruit of Brucea javanica. The present study aimed to evaluate the influence of brusatol on the progression of bladder cancer and uncover the molecular mechanism involved. We used Cell Counting Kit-8, colony formation and EdU assays to detect cell numbers, viability and proliferation. We used transwell migration assay to detect cell migration ability. The mechanism of brusatol inhibition of bladder cancer proliferation was studied by flow cytometry and western blotting. It was revealed that brusatol could reduce the viability and proliferation of T24 and 5637 cells. The transwell migration assay revealed that brusatol was able to attenuate the migration of T24 and 5637 cells. We found that treatment with brusatol increased the levels of reactive oxygen species, malondialdehyde and Fe2+, thereby further promoting ferroptosis in T24 and 5637 cells. In addition, treatment with RSL3 (an agonistor of ferroptosis) ferrostatin-1 (a selective inhibitor of ferroptosis) enhanced or reversed the brusatol-induced inhibition. In vivo, treatment with brusatol significantly suppressed the tumor growth in nude mice. Mechanistically, brusatol induced ferroptosis by upregulating the expression of ChaC glutathione-specific gamma-glutamylcyclotransferase (Chac1) and decreasing the expression of SLC7A11 and Nrf2 in T24 and 5637 cells. To summarize, the findings of this research demonstrated that brusatol hindered the growth of bladder cancer and triggered ferroptosis via the Chac1/Nrf2/SLC7A11 pathway.
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Sistema de Transporte de Aminoácidos y+ , Movimiento Celular , Proliferación Celular , Factor 2 Relacionado con NF-E2 , Cuassinas , Neoplasias de la Vejiga Urinaria , Neoplasias de la Vejiga Urinaria/patología , Neoplasias de la Vejiga Urinaria/tratamiento farmacológico , Neoplasias de la Vejiga Urinaria/metabolismo , Neoplasias de la Vejiga Urinaria/genética , Cuassinas/farmacología , Humanos , Factor 2 Relacionado con NF-E2/metabolismo , Factor 2 Relacionado con NF-E2/genética , Animales , Ratones , Proliferación Celular/efectos de los fármacos , Movimiento Celular/efectos de los fármacos , Línea Celular Tumoral , Sistema de Transporte de Aminoácidos y+/metabolismo , Sistema de Transporte de Aminoácidos y+/genética , Ratones Desnudos , Transducción de Señal/efectos de los fármacos , Ferroptosis/efectos de los fármacos , Ensayos Antitumor por Modelo de Xenoinjerto , Especies Reactivas de Oxígeno/metabolismo , Progresión de la Enfermedad , Ratones Endogámicos BALB C , Regulación Neoplásica de la Expresión Génica/efectos de los fármacosRESUMEN
Cisplatin resistance is a major challenge for systemic therapy against advanced bladder cancer (BC). Little information is available on the regulation of cisplatin resistance and the underlying mechanisms require elucidation. Here, we detected that downregulation of the tumor suppressor, PPP2R2B (a serine/threonine protein phosphatase 2 A regulatory subunit), in BC promoted cell proliferation and migration. What's more, low PPP2R2B expression was correlated with cisplatin resistance. In vitro and in vivo experiments verified that PPP2R2B could promote BC sensitivity to cisplatin. In terms of mechanism, we identified a novel function of PPP2R2B as a nucleocytoplasmic transport molecule. PPP2R2B promoted ISG15 entry into the nucleus by mediating binding of IPO5 with ISG15. Nuclear translocation of ISG15 inhibited DNA repair, further increasing ISG15 expression through activation of the STING pathway. Besides, PPP2R2B was down-regulated by SUV39H1-mediated histone 3 lysine 9 trimethylation, which could be restored by the SUV39H1-specific inhibitor, chaetocin. Our data suggest that PPP2R2B expression level is a potential biomarker for chemotherapy response and that chemotherapy in combination with chaetocin may be a feasible treatment strategy for patients with BC.
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Cisplatino , Citocinas , Resistencia a Antineoplásicos , Proteína Fosfatasa 2 , Ubiquitinas , Neoplasias de la Vejiga Urinaria , Neoplasias de la Vejiga Urinaria/tratamiento farmacológico , Neoplasias de la Vejiga Urinaria/metabolismo , Neoplasias de la Vejiga Urinaria/genética , Neoplasias de la Vejiga Urinaria/patología , Proteína Fosfatasa 2/metabolismo , Proteína Fosfatasa 2/genética , Humanos , Cisplatino/farmacología , Resistencia a Antineoplásicos/genética , Resistencia a Antineoplásicos/efectos de los fármacos , Ubiquitinas/metabolismo , Ubiquitinas/genética , Citocinas/metabolismo , Animales , Línea Celular Tumoral , Ratones , Proliferación Celular/efectos de los fármacos , Ratones Desnudos , Núcleo Celular/metabolismo , Antineoplásicos/farmacología , Ratones Endogámicos BALB C , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Movimiento Celular/efectos de los fármacos , Femenino , Proteínas del Tejido NerviosoRESUMEN
Patients with bladder cancer (BCa) frequently acquires resistance to platinum-based chemotherapy, particularly cisplatin. This study centered on the mechanism of cisplatin resistance in BCa and highlighted the pivotal role of lactylation in driving this phenomenon. Utilizing single-cell RNA sequencing, we delineated the single-cell landscape of Bca, pinpointing a distinctive subset of BCa cells that exhibit marked resistance to cisplatin with association with glycolysis metabolism. Notably, we observed that H3 lysine 18 lactylation (H3K18la) plays a crucial role in activating the transcription of target genes by enriching in their promoter regions. Targeted inhibition of H3K18la effectively restored cisplatin sensitivity in these cisplatin-resistant epithelial cells. Furthermore, H3K18la-driven key transcription factors YBX1 and YY1 promote cisplatin resistance in BCa. These findings enhance our understanding of the mechanisms underlying cisplatin resistance, offering valuable insights for identifying novel intervention targets to overcome drug resistance in Bca.
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Cisplatino , Neoplasias de la Vejiga Urinaria , Humanos , Cisplatino/farmacología , Cisplatino/uso terapéutico , Histonas/genética , Histonas/metabolismo , Análisis de Expresión Génica de una Sola Célula , Resistencia a Antineoplásicos/genética , Línea Celular Tumoral , Neoplasias de la Vejiga Urinaria/tratamiento farmacológico , Neoplasias de la Vejiga Urinaria/genética , Neoplasias de la Vejiga Urinaria/metabolismoRESUMEN
Drug resistance and tumor recurrence remain clinical challenges in the treatment of urothelial carcinoma (UC). However, the underlying mechanism is not fully understood. Here, we performed single-cell RNA sequencing and identified a subset of urothelial cells with epithelial-mesenchymal transition (EMT) features (EMT-UC), which is significantly correlated with chemotherapy resistance and cancer recurrence. To validate the clinical significance of EMT-UC, we constructed EMT-UC like cells by introducing overexpression of two markers, Zinc Finger E-Box Binding Homeobox 1 (ZEB1) and Desmin (DES), and examined their histological distribution characteristics and malignant phenotypes. EMT-UC like cells were mainly enriched in UC tissues from patients with adverse prognosis and exhibited significantly elevated EMT, migration and gemcitabine tolerance in vitro. However, EMT-UC was not specifically identified from tumorous tissues, certain proportion of them were also identified in adjacent normal tissues. Tumorous EMT-UC highly expressed genes involved in malignant behaviors and exhibited adverse prognosis. Additionally, tumorous EMT-UC was associated with remodeled tumor microenvironment (TME), which exhibited high angiogenic and immunosuppressive potentials compared with the normal counterparts. Furthermore, a specific interaction of COL4A1 and ITGB1 was identified to be highly enriched in tumorous EMT-UC, and in the endothelial component. Targeting the interaction of COL4A1 and ITGB1 with specific antibodies significantly suppressed tumorous angiogenesis and alleviated gemcitabine resistance of UC. Overall, our findings demonstrated that the driven force of chemotherapy resistance and recurrence of UC was EMT-UC mediated COL4A1-ITGB1 interaction, providing a potential target for future UC treatment.
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Resistencia a Antineoplásicos , Transición Epitelial-Mesenquimal , Integrina beta1 , Recurrencia Local de Neoplasia , Neovascularización Patológica , Microambiente Tumoral , Humanos , Transición Epitelial-Mesenquimal/efectos de los fármacos , Recurrencia Local de Neoplasia/tratamiento farmacológico , Recurrencia Local de Neoplasia/patología , Integrina beta1/metabolismo , Integrina beta1/genética , Neovascularización Patológica/tratamiento farmacológico , Neovascularización Patológica/patología , Neovascularización Patológica/genética , Microambiente Tumoral/efectos de los fármacos , Neoplasias de la Vejiga Urinaria/tratamiento farmacológico , Neoplasias de la Vejiga Urinaria/patología , Neoplasias de la Vejiga Urinaria/genética , Gemcitabina , Urotelio/patología , Urotelio/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Línea Celular Tumoral , Desoxicitidina/análogos & derivados , Desoxicitidina/farmacología , Desoxicitidina/uso terapéutico , Pronóstico , Movimiento Celular/efectos de los fármacos , Homeobox 1 de Unión a la E-Box con Dedos de Zinc/genética , Homeobox 1 de Unión a la E-Box con Dedos de Zinc/metabolismo , Angiogénesis , Colágeno Tipo IVRESUMEN
Urinary cell-free DNA (ucfDNA) is a potential biomarker for bladder cancer detection. However, the biological characteristics of ucfDNA are not well understood. We explored the roles of deoxyribonuclease 1 (DNASE1) and deoxyribonuclease 1-like 3 (DNASE1L3) in the fragmentation of ucfDNA using mouse models. The deletion of Dnase1 in mice (Dnase1-/-) caused aberrations in ucfDNA fragmentation, including a 24-fold increase in DNA concentration, and a 3-fold enrichment of long DNA molecules, with a relative decrease of fragments with thymine ends and reduction of jaggedness (i.e., the presence of single-stranded protruding ends). In contrast, such changes were not observed in mice with Dnase1l3 deletion (Dnase1l3-/-). These results suggested that DNASE1 was an important nuclease contributing to the ucfDNA fragmentation. Western blot analysis revealed that the concentration of DNASE1 protein was higher in urine than DNASE1L3. The native-polyacrylamide gel electrophoresis zymogram showed that DNASE1 activity in urine was higher than that in plasma. Furthermore, the proportion of ucfDNA fragment ends within DNase I hypersensitive sites (DHSs) was significantly increased in Dnase1-deficient mice. In humans, patients with bladder cancer had lower proportions of ucfDNA fragment ends within the DHSs when compared with participants without bladder cancer. The area under the curve (AUC) for differentiating patients with and without bladder cancer was 0.83, suggesting the analysis of ucfDNA fragmentation in the DHSs may have potential for bladder cancer detection. This work revealed the intrinsic links between the nucleases in urine and ucfDNA fragmentomics.
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Ácidos Nucleicos Libres de Células , Neoplasias de la Vejiga Urinaria , Animales , Ácidos Nucleicos Libres de Células/genética , ADN/genética , Desoxirribonucleasa I/genética , Desoxirribonucleasa I/metabolismo , Endodesoxirribonucleasas/genética , Endonucleasas , Humanos , Ratones , Ratones Noqueados , Neoplasias de la Vejiga Urinaria/genéticaRESUMEN
Metastasis contributes to the dismal prognosis of bladder cancer (BLCA). The mechanical status of the cell membrane is expected to mirror the ability of cell migration to promote cancer metastasis. However, the mechanical characteristics and underlying molecular profile associated with BLCA metastasis remain obscure. To study the unique cellular architecture and traits associated with cell migration, using a process called cell-based systematic evolution of ligands by exponential enrichment (cell-SELEX) we generated an aptamer-based molecular probe, termed spl3c, which identified cytoskeleton-associated protein 4 (CKAP4). CKAP4 was associated with tumor metastasis in BLCA, but we also found it to be a mechanical regulator of BLCA cells through the maintenance of a central-to-peripheral gradient of stiffness on the cell membrane. Notably, such mechanical traits were transportable through exosome-mediated intercellular CKAP4 trafficking, leading to significant enhancement of migration in recipient cells and, consequently, aggravating metastatic potential in vivo. Taken together, our study shows the robustness of this aptamer-based molecular tool for biomarker discovery, revealing the dominance of a CKAP4-induced central-to-peripheral gradient of membrane stiffness that benefits cell migration and delineating the role of exosomes in mediating mechanical signaling in BLCA metastasis.