Unveiling the impact of tertiary lymphoid structures on immunotherapeutic responses of clear cell renal cell carcinoma.

Tertiary lymphoid structures (TLS) are organized aggregates of immune cells that form under pathological conditions. However, the predictive value of TLS in clear cell renal cell carcinoma (ccRCC) for immunotherapies remains unclear. We comprehensively assessed the implications for prognosis and immunological responses of the TLS spatial and maturation heterogeneity in 655 ccRCC patients. A higher proportion of early-TLS was found in peritumoral TLS, while intratumoral TLS mainly comprised secondary follicle-like TLS (SFL-TLS), indicating markedly better survival. Notably, presence of TLS, especially intratumoral TLS and SFL-TLS, significantly correlated with better survival and objective reflection rate for ccRCC patients receiving anti-Programmed Cell Death Protein-1 (PD-1)/Programmed Cell Death-Ligand-1 (PD-L1) immunotherapies. In peritumoral TLS cluster, primary follicle-like TLS, the proportion of tumor-associated macrophages, and Treg infiltration in the peritumoral regions increased prominently, suggesting an immunosuppressive tumor microenvironment. Interestingly, spatial transcriptome annotation and multispectral fluorescence showed that an abundance of mature plasma cells within mature TLS has the capacity to produce IgA and IgG, which demonstrate significantly higher objective response rates and a superior prognosis for ccRCC patients subjected to immunotherapy. In conclusion, this study revealed the implications of TLS spatial and maturation heterogeneity on the immunological status and clinical responses, allowing the improvement of precise immunotherapies of ccRCC.

Single-cell RNA-seq analysis decodes the kidney microenvironment induced by polystyrene microplastics in mice receiving a high-fat diet.

In recent years, the environmental health issue of microplastics has aroused an increasingly significant concern. Some studies suggested that exposure to polystyrene microplastics (PS-MPs) may lead to renal inflammation and oxidative stress in animals. However, little is known about the essential effects of PS-MPs with high-fat diet (HFD) on renal development and microenvironment. In this study, we provided the single-cell transcriptomic landscape of the kidney microenvironment induced by PS-MPs and HFD in mouse models by unbiased single-cell RNA sequencing (scRNA-seq). The kidney injury cell atlases in mice were evaluated after continued PS-MPs exposure, or HFD treated for 35 days. Results showed that PS-MPs plus HFD treatment aggravated the kidney injury and profibrotic microenvironment, reshaping mouse kidney cellular components. First, we found that PS-MPs plus HFD treatment acted on extracellular matrix organization of renal epithelial cells, specifically the proximal and distal convoluted tubule cells, to inhibit renal development and induce ROS-driven carcinogenesis. Second, PS-MPs plus HFD treatment induced activated PI3K-Akt, MAPK, and IL-17 signaling pathways in endothelial cells. Besides, PS-MPs plus HFD treatment markedly increased the proportions of CD8+ effector T cells and proliferating T cells. Notably, mononuclear phagocytes exhibited substantial remodeling and enriched in oxidative phosphorylation and chemical carcinogenesis pathways after PS-MPs plus HFD treatment, typified by alterations tissue-resident M2-like PF4+ macrophages. Multispectral immunofluorescence and immunohistochemistry identified PF4+ macrophages in clear cell renal cell carcinoma (ccRCC) and adjacent normal tissues, indicating that activate PF4+ macrophages might regulate the profibrotic and pro-tumorigenic microenvironment after renal injury. In conclusion, this study first systematically revealed molecular variation of renal cells and immune cells in mice kidney microenvironment induced by PS-MPs and HFD with the scRNA-seq approach, which provided a molecular basis for decoding the effects of PS-MPs on genitourinary injury and understanding their potential profibrotic and carcinogenesis in mammals.

Single-cell transcriptome analysis of liver immune microenvironment changes induced by microplastics in mice with non-alcoholic fatty liver.

Recent studies have discovered that tiny particles of microplastics (MPs) at the nano-scale level can enter the body of organisms from the environment, potentially causing metabolic ailments. However, further investigation is required to understand the alterations in the immune microenvironment associated with non-alcoholic fatty liver disease (NAFLD) occurrence following exposure to MPs. Experiments were performed using mice, which were given a normal chow or high-fat diet (NCD or HFD, respectively) plus free drinking of sterile water with or without MPs, respectively. Employing an impartial technique known as unbiased single-cell RNA-sequencing (scRNA-seq), the cellular (single-cell) pathology landscape of NAFLD and related changes in the identified immune cell populations induced following MPs plus HFD treatment were assessed. The results showed that mice in the HFD groups had remarkably greater NAFLD activity scores than those from the NCD groups. Moreover, administration of MPs plus HFD further worsened the histopathological changes in the mice's liver, leading to hepatic steatosis, inflammatory cell infiltrations and ballooning degeneration. Following the construction of a sing-cell resolution transcriptomic atlas of 43,480 cells in the mice's livers of the indicated groups, clear cellular heterogeneity and potential cell-to-cell cross-talk could be observed. Specifically, we observed that MPs exacerbated the pro-inflammatory response and influenced the stemness of hepatocytes during HFD feeding. Importantly, treatment with MPs significantly increase the infiltration of the infiltrating liver-protecting Vsig4+ macrophages in the liver of the NAFLD mouse model while remarkably decreasing the angiogenic S100A6+ macrophage subpopulation. Furthermore, mice treated with MPs plus HFD exhibited significantly increased recruitment of CD4+ cells and heightened exhaustion of CD8+ T cells than those from the control group, characteristics typically associated with the dysregulation of immune homeostasis and severe inflammatory damage. Overall, this study offers valuable perspectives into comprehending the potential underlying cellular mechanisms and regulatory aspects of the microenvironment regarding MPs in the development of NAFLD.

CCL19: a novel prognostic chemokine modulates the tumor immune microenvironment and outcomes of cancers.

BACKGROUND: CCL19 is a chemokine involved in cancer research due to its important role in the tumor microenvironment (TME) and clinical relevance in cancers. This study aimed to analyze transcription expression, genomic alteration, association with tumor immune microenvironment of CCL19 expression and its prediction value for prognosis and responses to immunotherapy for patients with cancers. METHODS: RNA sequencing data and corresponding clinicopathological information of a total of large-scale cancer patients were obtained from The Cancer Genome Atlas and Gene Expression Omnibus databases. Multiplex immunofluorescence (mIF) was implemented to identify differential infiltration of Treg, CD8+ T cells, and tumor-associated macrophages, while CCL19 immunohistochemistry was conducted on 182 breast cancer samples from a real-world cohort. RESULTS: Based on large-scale multi-center survival analysis of cancer patients, we found the prognosis of patients with high CCL19 expression was prominently better than those with low CCL19 expression. For patients from multiple independent cohorts, suppressed CCL19 expression exerts significant progressive phenotype and apoptosis activity of cancers, especially in breast and ovarian cancer. Interestingly, anti-tumor immune cells, specifically the CD8+ T cells and macrophages, were clustered from TME by elevated CCL19 expression. Additionally, higher CCL19 levels reflected heightened immune activity and substantial heterogeneity. CONCLUSIONS: In conclusion, our findings support the notion that elevated CCL19 expression is linked to favorable outcomes and enhanced anti-tumor immunity, characterized by increased CD8+ T cells within the TME. This suggests the potential of CCL19 as a prognostic marker, predictive biomarker for immunotherapy, therapeutic target of cancers.

Protein Arginine Methyltransferases Refine the Classification of Clear Cell Renal Cell Carcinoma with Distinct Prognosis and Tumor Microenvironment Characteristics.

Background: Clear cell renal cell carcinoma (ccRCC) is an aggressive urological cancer that originates from the proximal tubular epithelium. As one of the most common post-translational modification, protein arginine methylation plays a pivotal role in various cancer-associated biological functions, especially in cancer immunity. Therefore, constructing a protein arginine methylation-related prognostic signature would be beneficial in guiding better personalized clinical management for patients with ccRCC. Methods: Based on the multi-omics profiling of the expression levels of eight protein arginine methyltransferases (PRMTs) in 763 ccRCC samples (from TCGA, CPTAC, EMBL, and ICGC databases), we established a scoring system with machine-learning algorithms to quantify the modification patterns on clinical and immunological characterizations of individual ccRCC patient, which was termed as PRMTScore. Moreover, we utilized two external clinical cohorts receiving immunotherapy (n=302) to validate the reliability of the PRMTScore system. Multiplex immunohistochemistry (mIHC) was performed to characterize the cellular composition of 30 paired ccRCC samples. The proteomic profiling of 232 ccRCC samples obtained from Fudan University Shanghai Cancer Center (FUSCC) was analyzed to validate the protein expression of PRMT5 in ccRCC. Finally, CCK-8, transwell, and wound healing assays were conducted to elucidate the role of PRMT5 in ccRCC in vitro. Results: A total of 763 ccRCC patients with available multi-omics profiling were stratified into two clusters (PRMTCluster A and B) with distinctive prognosis, genomic alterations, tumor microenvironment (TME) characteristics, and fundamental biological mechanisms. Subsequently, protein arginine methylation-related prognostic signature (PRMTScore) was constructed and consisted of SLC16A12, HRH2, F2RL3, and SAA1. The PRMTScore showed remarkable differences in outcomes, immune and stromal fractions, expressions of immune checkpoints, the abundance of immune cells, and immunotherapy response in ccRCC patients. Additionally, preliminary insights unveiled the tumor-suppressive role of PRMT5 in ccRCC, and the signal of PRMT5low significantly predicted aggressive prognosis and the high abundance of PD1+ CD8+ cells in ccRCC. Conclusion: We constructed a PRMTScore system, which showed the potent ability to assess the prognosis, TME characteristics, and immunotherapy response for patients with ccRCC. Moreover, this is the first study to propose that PRMT5 acts as a cancer suppressor in ccRCC.

Integrated machine learning and bioinformatic analyses used to construct a copper-induced cell death-related classifier for prognosis and immunotherapeutic response of hepatocellular carcinoma patients.

Background: Copper as phytonutrient has powerful activity against health diseases. A newly discovered mechanism of cell death that affects energy metabolism by copper ("cuproptosis") can induce multiple cuproptosis-related genes. Hepatocellular carcinoma (HCC) is a poorly prognosed widespread cancer having danger of advanced metastasis. Therefore, earlier diagnosis followed by the specific targeted therapy are required for improved prognosis. The work herein constructed scoring system built on ten cuproptosis-related genes (CRGs) to predict progression of tumor and metastasis more accurately and test patient reaction toward immunotherapy. Methods: A comprehensive assessment of cuproptosis patterns in HCC samples from two databases and a real-world cohort was performed on ten CRGs, that were linked to immune cell infiltration signatures of TME (tumor microenvironment). Risk signatures were created for quantifying effect of cuproptosis on HCC, and the effects of related genes on cellular function of HCC were investigated, in addition to the effects of immunotherapy and targeted therapy drugs. Results: Two distinct cuproptosis-associated mutational patterns were identified, with distinct immune cell infiltration characteristics and survival likelihood. Studies have shown that assessment of cuproptosis-induced tumor mutational patterns can help predict tumor stage, phenotype, stromal activity, genetic diversity, and patient prognosis. High risk scores are characterized by lower survival and worse treatment with anti-PD-L1/CTAL4 immunotherapy and first-line targeted drugs. Cytological functional assays show that CDKN2A and GLS promote proliferation, migration and inhibit copper-dependent death of HCC cells. Conclusion: HCC patients with high-risk scores exhibit significant treatment disadvantage and survival rates. Cuproptosis plays a non-negligible role in the development of HCC. Quantifying cuproptosis-related designs of tumors will aid in phenotypic categorization, leading to efficient personalized and targeted therapeutics and precise prediction of prognosis and metastasis.

Integrating machine learning and bioinformatics analysis to m6A regulator-mediated methylation modification models for predicting glioblastoma patients' prognosis and immunotherapy response.

BACKGROUND: Epigenetic regulations of immune responses are essential for cancer development and growth. As a critical step, comprehensive and rigorous explorations of m6A methylation are important to determine its prognostic significance, tumor microenvironment (TME) infiltration characteristics and underlying relationship with glioblastoma (GBM). METHODS: To evaluate m6A modification patterns in GBM, we conducted unsupervised clustering to determine the expression levels of GBM-related m6A regulatory factors and performed differential analysis to obtain m6A-related genes. Consistent clustering was used to generate m6A regulators cluster A and B. Machine learning algorithms were implemented for identifying TME features and predicting the response of GBM patients receiving immunotherapy. RESULTS: It is found that the m6A regulatory factor significantly regulates the mutation of GBM and TME. Based on Europe, America, and China data, we established m6Ascore through the m6A model. The model accurately predicted the results of 1206 GBM patients from the discovery cohort. Additionally, a high m6A score was associated with poor prognoses. Significant TME features were found among the different m6A score groups, which demonstrated positive correlations with biological functions (i.e., EMT2) and immune checkpoints. CONCLUSIONS: m6A modification was important to characterize the tumorigenesis and TME infiltration in GBM. The m6Ascore provided GBM patients with valuable and accurate prognosis and prediction of clinical response to various treatment modalities, which could be useful to guide patient treatments.

Bioinformatic analyzes and validation of cystathionine gamma-lyase as a prognostic biomarker and related to immune infiltrates in hepatocellular carcinoma.

Background: The role of cystathionine γ-lyase (CTH) in the prognosis and immune invasion of hepatocellular carcinoma (HCC) remains poorly understood. Methods: In this study, the clinical data of patients with HCC were analyzed, and the expression level of CTH was compared between HCC and normal tissues using the R package and various databases. Results: We found that CTH expression was significantly decreased in HCC compared with normal tissues, and its expression was associated with various clinicopathological factors, including tumor stage, gender, tumor status, residual tumor, histologic stage, race, alpha-fetoprotein (AFP), albumin, drinking, and smoking. Our results suggest that CTH might be a protective factor for the survival of patients with HCC. Further functional analysis revealed that high CTH expression was enriched in the Reactome signaling by interleukins and the Reactome neutrophil degranulation. Moreover, CTH expression was closely correlated with a variety of immune cells, including a negative correlation with the CD56 (bright) NK cells and follicular helper T cell (TFH), while a positive correlation with Th17 cells and central memory T cell (Tcm). High expression of CTH in immune cells predicted a better prognosis of HCC. Our findings further indicated Pyridoxal phosphate, l-cysteine, Carboxymethylthio-3-(3-chlorophenyl)-1,2,4-oxadiazol, 2-[(3-Hydroxy-2-Methyl-5-Phosphonooxymethyl-Pyridin-4-Ylmethyl)-Imino]-5-phosphono-pent-3-enoic acid and L-2-amino-3-butynoic acid as potential target candidate medications for HCC treatment based on CTH. Conclusion: Our study suggests that CTH can serve as a biomarker to predict the prognosis and immune infiltration of HCC.

Tumor microenvironment-based signatures distinguish intratumoral heterogeneity, prognosis, and immunogenomic features of clear cell renal cell carcinoma.

Background: The tumor microenvironment (TME) performs a crucial function in the tumorigenesis and response to immunotherapies of clear cell renal cell carcinoma (ccRCC). However, a lack of recognized pre-clinical TME-based risk models poses a great challenge to investigating the risk factors correlated with prognosis and treatment responses for patients with ccRCC. Methods: Stromal and immune contexture were assessed to calculate the TMErisk score of a large sample of patients with ccRCC from public and real-world cohorts using machine-learning algorithms. Next, analyses for prognostic efficacy, correlations with clinicopathological features, functional enrichment, immune cell distributions, DNA variations, immune response, and heterogeneity were performed and validated. Results: Clinical hub genes, including INAFM2, SRPX, DPYSL3, VSIG4, APLNR, FHL5, A2M, SLFN11, ADAMTS4, IFITM1, NOD2, CCR4, HLA-DQB2, and PLAUR, were identified and incorporated to develop the TMErisk signature. Patients in the TMEhigh risk group (category) exhibited a considerably grim prognosis, and the TMErisk model was shown to independently function as a risk indicator for the overall survival (OS) of ccRCC patients. Expression levels of immune checkpoint genes were substantially increased in TMEhigh risk group, while those of the human leukocyte antigen (HLA) family genes were prominently decreased. In addition, tumors in the TMEhigh group showed significantly high infiltration levels of tumor-infiltrated lymphocytes, including M2 macrophages, CD8+ T cells, B cells, and CD4+ T cells. In heterogeneity analysis, more frequent somatic mutations, including pro-tumorigenic BAP1 and PBRM1, were observed in the TMEhigh group. Importantly, 19.3% of patients receiving immunotherapies in the TMEhigh group achieved complete or partial response compared with those with immune tolerance in the TMElow group, suggesting that TMErisk prominently differentiates prognosis and responses to immunotherapy for patients with ccRCC. Conclusions: We first established the TMErisk score of ccRCC using machine-learning algorithms based on a large-scale population. The TMErisk score can be utilized as an innovative independent prognosis predictive marker with high sensitivity and accuracy. Our discovery also predicted the efficacy of immunotherapy in ccRCC patients, indicating the intimate link between tumor immune microenvironment and intratumoral heterogeneity.

Immunogenomic Characteristics of Cell-Death-Associated Genes with Prognostic Implications in Bladder Cancer.

Bladder cancer is one of the most common genitourinary malignant cancers worldwide. Cell death processes, including apoptosis, ferroptosis, and necrosis, provide novel clinical and immunological insights promoting the management of precision medicine. Therefore, this study aimed to evaluate the transcriptomic profile of signatures in cell death pathways with significant prognostic implications in patients with bladder cancer from multiple independent cohorts (n = 1999). First, genes involved in apoptosis (n = 19), ferroptosis (n = 31), and necrosis (n = 6) were analyzed to evaluate the prognostic implications in bladder cancer. Significant genes were included to establish the cell-death index (CDI) of 36 genes that distinguished patients according to high and low risks. Survival analysis using the Kaplan-Meier curves clustered patients based on overall survival (18.8 vs. 96.7 months; hazard model [HR] = 3.12, P<00001). Cox proportional hazard model was significantly associated with a higher risk of mortality using 10 external independent cohorts in patients with CDIhigh (HR = 1.31, 95% CI: 1.04-1.62). To explore immune parameters associated with CDI, microenvironment cell-population-counter algorithms indicated increased intratumoral heterogeneity and macrophage/monocyte infiltration and CD8+ T cells in patients with CDIhigh group. Besides, the CDIhigh group showed an increased expression of the following immune checkpoints: CD276, PD-L1, CTLA-4, and T-cell exhaustion signatures. Cytokine expression analysis revealed the highest association of IL-9R, IL-17A, IL-17F, GDF7, and IFNW1 with the high-risk group. In addition, 42 patients with BCa receiving immunotherapies were enrolled from a real-world cohort, and expression patterns of three CDI hub genes (DRD5, SCL2A14, and IGF1) were detected using immunohistochemical staining. Patients with triple-negative staining of tumor tissues had significantly higher tumor-associated macrophage abundance, PD-L1 expression, predicted immunocompromised microenvironment, and prominently progressive progression (HR = 4.316, P = 0.0028). In conclusion, this study highlights the immunoevasive tumor microenvironment characterized by the higher tumor-associated macrophage infiltration with the presence of immune checkpoint and T-cell exhaustion genes in patients with BCa at CDIhigh risk who might suffer progression and be more suitable to benefit from immune checkpoint inhibitors or other immunotherapies.

Increased DNA Polymerase Epsilon Catalytic Subunit Expression Predicts Tumor Progression and Modulates Tumor Microenvironment of Hepatocellular Carcinoma.

Backgrounds: Liver hepatocellular carcinoma (HCC) is one of the most common cancers worldwide, and POLE, playing an important role in maintaining genetic stability, is closely connected with cancer prognosis. This study aimed to explore the significance role of POLE in HCC prognosis, clinical treatment and tumor immune microenvironment based on large-scale multiply cohorts. Methods: First, we found that the expression of POLE was prominently higher in tumor tissues than in normal tissues, and was closely related to clinical stage, grade and patient outcomes. Second, we found that patients with high POLE expression had significantly aggressive progression, indicating effective predictive role of POLE expression for Asian, male, low-risk HCC patients. Additionally, POLE mutation frequency was detected in several datasets with available genomic-wide data. Results: 130 HCC samples from real-world Renji cohort were included to demonstrate that elevated POLE expression was significantly connected to the invasive progression and poor prognosis. More importantly, the expression of POLE was closely related to the anti-tumoral activity of immune cells and immune checkpoints expression, suggesting a bright prospect of POLE as a predictive biomarker in immunotherapy. Conclusion: In conclusion, this study revealed that high expression of POLE significantly correlated to the malignant progression, poor prognosis and anti-tumoral activity of immune cells in HCC. Thus, POLE could function as a biomarker for the early diagnosis, prognosis, immune-excluded tumor microenvironment and response to immunotherapy of HCC.

Deciphering the role of miR-187-3p/LRFN1 axis in modulating progression, aerobic glycolysis and immune microenvironment of clear cell renal cell carcinoma.

Clear cell renal cell carcinoma (ccRCC) is one of the most common malignant genitourinary cancers with high recurrence risk worldwide. Recently, multi-omics data facilitate obtaining a molecular landscape of tumor development, and were implemented to affect pathogenesis, phenotype, and prognosis of ccRCC. In this study, after screening for differential expressed microRNAs based on multiply datasets, we tested expression levels and prognostic value of miR-187-3p in ccRCC samples, and transfected miR-187-3p mimics or negative controls into ccRCC cells. Up-regulation of miR-187-3p restrains proliferation, migration and promotes apoptosis ability in human ccRCC A498 and 786O cells. In addition, Luciferase reporter assay revealed that miR-187-3p directly targets LRFN1-3'-UTR and negatively modulates LRFN1 expression. LRFN1 rescues proliferation and invasion capacities after miR-187-3p mimic transfection in vitro and in subcutaneous xenograft models. We further performed deep-sequencing technology and bioinformatics analyses to evaluate the biological functions and potential clinical implications of LRFN1 expression in ccRCC. Interestingly, LRFN1 could serve as an independent and potential biomarker for prognosis in over 1000 patients with ccRCC from multiply independent cohorts. Besides, the up-regulated LRFN1 expression prominently promoted intra-tumoral heterogeneity and immune-infiltrating microenvironment, represented by elevated M2 macrophage infiltration, CD8+ T cells activity and PD-L1 expression. In conclusion, this study revealed the tumor-specific and immunological role of miR-187-3p/LRFN1 axis in the progression and reshaping of tumor immune microenvironment of ccRCC.

Transcriptome Profiles Reveal a 12-Signature Metabolic Prediction Model and a Novel Role of Myo-Inositol Oxygenase in the Progression of Prostate Cancer.

Prostate adenocarcinoma (PRAD) is an extremely common type of cancer in the urinary system. Here, we aimed to establish a metabolic signature to identify novel targets in a predictive model of PRAD patients. A total of 133 metabolic differentially expressed genes (MDEGs) were identified with significant prognostic value. Least absolute shrinkage and selection operator (LASSO) regression analysis was used to construct a 12-mRNA signature model, a metabolic prediction model (MPM), in 491 PRAD patients. The risk score of the MPM significantly predicted the progression of PRAD patients (p < 0.001, area under the curve (AUC) = 0.745). Furthermore, myo-inositol oxygenase (MIOX), the most prominently upregulated metabolic enzyme and hub gene in the protein-protein interaction network of the MPM, showed significant prognostic implications. Next, MIOX expression in normal prostate tissues was lower than in PRAD tissues, and high MIOX expression was significantly associated with disease progression (p = 0.005, HR = 2.274) in 81 PRAD patients undergoing first-line androgen receptor signaling inhibitor treatment from the Renji cohort. Additionally, MIOX was significantly involved in the abnormal immune infiltration of the tumor microenvironment and associated with the DNA damage repair process of PRAD. In conclusion, this study provides the first opportunity to comprehensively elucidate the landscape of prognostic MDEGs, establish novel prognostic modeling of MPM using large-scale PRAD transcriptomic data, and identify MIOX as a potential prognostic target in PRAD patients from multiple cohorts. These findings help manage risk assessment and provide valuable insights into treatment strategies for PRAD.

Prognostic value, DNA variation and immunologic features of a tertiary lymphoid structure-related chemokine signature in clear cell renal cell carcinoma.

BACKGROUND: The tumor microenvironment (TME) and tertiary lymphoid structures (TLS) affect the occurrence and development of cancers. How the immune contexture interacts with the phenotype of clear cell renal cell carcinoma (ccRCC) remains unclear. METHODS: We identified and evaluated TLS clusters in ccRCC using machine learning algorithms and the 12-chemokine gene signature for TLS. Analyses for functional enrichment, DNA variation, immune cell distribution, association with independent clinicopathological features and predictive value of CXCL13 in ccRCC were performed. RESULTS: We found a prominently enrichment of the 12-chemokine gene signature for TLS in patients with ccRCC compared with other types of renal cell carcinoma. We identified a prognostic value of CCL4, CCL5, CCL8, CCL19 and CXCL13 expression in ccRCC. DNA deletion of the TLS gene signature significantly predicted poor outcome in ccRCC compared with amplification and wild-type gene signature. We established TLS clusters (C1-4) and observed distinct differences in survival, stem cell-like characteristics, immune cell distribution, response to immunotherapies and VEGF-targeted therapies among the clusters. We found that elevated CXCL13 expression significantly predicted aggressive progression and poor prognosis in 232 patients with ccRCC in a real-world validation cohort. CONCLUSION: This study described a 12-chemokine gene signature for TLS in ccRCC and established TLS clusters that reflected different TME immune status and corresponded to prognosis of ccRCC. We confirmed the dense presence of TILs aggregation and TLS in ccRCC and demonstrated an oncogenic role of CXCL13 expression of ccRCC, which help develop immunotherapies and provide novel insights on the long-term management of ccRCC.

Protumorigenic Role of Elevated Levels of DNA Polymerase Epsilon Predicts an Immune-Suppressive Microenvironment in Clear Cell Renal Cell Carcinoma.

Increasing evidence indicates that DNA polymerase epsilon (POLE), which mediates DNA damage repair, is significantly associated with tumor prognosis. This study aimed to analyze POLE expression in tumor samples and its prognostic value for patients with clear cell renal cell carcinoma (ccRCC). We found significantly elevated POLE expression in ccRCC tissues compared with normal tissues of multiple independent cohorts. The POLE expression levels of 523 patients with ccRCC (The Cancer Genome Atlas RNA-seq data) and 179 patients with ccRCC with immunohistochemical data (Fudan University Shanghai Cancer Center) were analyzed to investigate the prognostic implications of POLE expression. Cox regression analyses were implemented to explore the effect of POLE expression on the prognosis of pan-cancer. These findings revealed that elevated POLE expression levels significantly correlated with shorter overall survival (p < 0.001, n = 701) of patients with ccRCC. These data indicate that POLE expression may serve as a prognostic biomarker for cancers. Although POLE mutations were not significantly associated with survival benefits conferred upon patients with ccRCC, a CD4+ T cell-regulated immune microenvironment was significantly activated. Moreover, we found that POLE expression in cancers significantly correlated with an immunosuppressive tumor microenvironment, higher intratumoral heterogeneity, and expression of immune checkpoint genes PDCD1, CTLA4, and CD86, possibly mediated via the JAK/STAT and Notch signaling pathways. In conclusion, the present study is the first to our knowledge to indicate that elevated POLE expression is significantly associated with poor survival and an immune-suppressive tumor microenvironment in ccRCC. These findings suggest that POLE can serve as a biomarker for guiding molecular diagnosis and facilitating the development of novel individual therapeutic strategies for patients with advanced ccRCC.

Prognostic Immunophenotyping Clusters of Clear Cell Renal Cell Carcinoma Defined by the Unique Tumor Immune Microenvironment.

Background: The tumor microenvironment affects the occurrence and development of cancers, including clear cell renal cell carcinoma (ccRCC). However, how the immune contexture interacts with the cancer phenotype remains unclear. Methods: We identified and evaluated immunophenotyping clusters in ccRCC using machine-learning algorithms. Analyses for functional enrichment, DNA variation, immune cell distribution, association with independent clinicopathological features, and predictive responses for immune checkpoint therapies were performed and validated. Results: Three immunophenotyping clusters with gradual levels of immune infiltration were identified. The intermediate and high immune infiltration clusters (Clusters B and C) were associated with a worse prognosis for ccRCC patients. Tumors in the immune-hot Clusters B and C showed pro-tumorigenic immune infiltration, and these patients showed significantly worse survival compared with patients in the immune-cold Cluster A in the training and testing cohorts (n = 422). In addition to distinct immune cell infiltrations of immunophenotyping, we detected significant differences in DNA variation among clusters, suggesting a high degree of genetic heterogeneity. Furthermore, expressions of multiple immune checkpoint molecules were significantly increased. Clusters B and C predicted favorable outcomes in 64 ccRCC patients receiving immune checkpoint therapies from the FUSCC cohort. In 360 ccRCC patients from the FUSCC validation cohort, Clusters B and C significantly predicted worse prognosis compared with Cluster A. After immunophenotyping of ccRCC was confirmed, significantly increased tertiary lymphatic structures, aggressive phenotype, elevated glycolysis and PD-L1 expression, higher abundance of CD8+ T cells, and TCRn cell infiltration were found in the immune-hot Clusters B and C. Conclusion: This study described immunophenotyping clusters that improved the prognostic accuracy of the immune contexture in the ccRCC microenvironment. Our discovery of the novel independent prognostic indicators in ccRCC highlights the relationship between tumor phenotype and immune microenvironment.

Integrative 5-Methylcytosine Modification Immunologically Reprograms Tumor Microenvironment Characterizations and Phenotypes of Clear Cell Renal Cell Carcinoma.

The tumor microenvironment (TME) affects the biologic malignancy of clear cell renal cell carcinoma (ccRCC). The influence of the 5-methylcytosine (m5C) epigenetic modification on the TME is unknown. We comprehensively assessed m5C modification patterns of 860 ccRCC samples (training, testing, and real-world validation cohorts) based on 17 m5C regulators and systematically integrated the modification patterns with TME cell-infiltrating characterizations. Our results identified distinct m5C modification clusters with gradual levels of immune cell infiltration. The distinct m5C modification patterns differ in clinicopathological features, genetic heterogeneity, patient prognosis, and treatment responses of ccRCC. An elevated m5C score, characterized by malignant biologic processes of tumor cells and suppression of immunity response, implies an immune-desert TME phenotype and is associated with dismal prognosis of ccRCC. Activation of exhausted T cells and effective immune infiltration were observed in the low m5C score cluster, reflecting a noninflamed and immune-excluded TME phenotype with favorable survival and better responses to immunotherapy. Together, these findings provide insights into the regulation mechanisms of DNA m5C methylation modification patterns on the tumor immune microenvironment. Comprehensive assessment of tumor m5C modification patterns may enhance our understanding of TME cell-infiltrating characterizations and help establish precision immunotherapy strategies for individual ccRCC patients.

CDC20 regulates the cell proliferation and radiosensitivity of P53 mutant HCC cells through the Bcl-2/Bax pathway.

Purpose: The incidence of hepatocellular carcinoma (HCC) is extremely high, and China accounts for approximately 50% of global liver cancer cases. Previous studies reported that CDC20 is involved in the occurrence and progression of a variety of malignant tumors. So, whether CDC20 will affect the development of HCC, we have conducted in-depth research on this. Methods: We selected Hep3B and HepG2 for cell culture, and performed siRNA transfection, lentiviral infection, western blot, MTS determination, cell cycle determination, apoptosis test, immunodeficiency test, clone survival test and subcutaneous parthenogenesis in nude mice. Results: Knockdown of CDC20 greatly enhanced the radiation efficacy on the growth retardation in HepG2, and protein level of CDC20 was decreased for the activation of P53 by radiation. Downregulation of CDC20 combined with radiation can inhibit proliferation, aggravate DNA damage, increase G2/M arrest, and promote apoptosis of HCC cells to a greater extent, and the relative survival fraction of HCC cells was gradually reduced with radiation dose increased in P53 mutated Hep3B cells. After knocking down CDC20 in HCC, Bcl-2 was down-regulated and Bax expression increased. Down-regulation of CDC20 can inhibit further invasion by promoting the radiosensitivity of HCC. Conclusion: In this study, we found that that CDC20 was highly expressed in HCC and participated in radio resistance of HCC cells with P53 mutation Bcl-2/Bax via signaling pathway. This study is the first to present evidence that CDC20 may play a role in improving the efficacy of radiotherapy in HCC.