CTCF-activated FUCA1 functions as a tumor suppressor by promoting autophagy flux and serum α-L-fucosidase serves as a potential biomarker for prognosis in ccRCC.

Notably, clear cell renal cell carcinoma (ccRCC) is characterized by a distinct metabolic tumor phenotype that involves the reprogramming of multiple metabolic pathways. Although there is increasing evidence linking FUCA1 to malignancies, its specific role and downstream signaling pathways in ccRCC remain poorly understood. Here we found that FUCA1 expression was significantly downregulated in ccRCC tissues, which also predicts poor prognosis of ccRCCpatients. Moreover, enhancing FUCA1 expression resulted in reduced invasion and migration of ccRCC cells, further indicating its protective role. CHIP-qPCR and luciferase assays showed that CTCF was an upstream transcription factor of FUCA1 and could reverse the effects caused by FUCA1 inactivation. The change in FUCA1 led to changes in the results of various autophagy-related proteins and the mRFP-GFP-LC3 dual fluorescence system, indicating that it may play a role in the fusion stage of autophagy. Protein-protein interaction analysis revealed that FUCA2 exhibited the closest interaction with FUCA1 and strongly predicted the prognosis of ccRCC patients. Additionally, serum AFU encoded by FUCA2 could serve as a valuable predictor for survival in ccRCC patients. FUCA1 suppresses invasion and migration of ccRCC cells, with its activity being modulated by CTCF. FUCA1 regulates the autophagy process in ccRCC cells by influencing the fusion between autophagosomes and lysosomes. FUCA2 shares similarities with FUCA1, and elevated serum AFU levels along with increased expression of FUCA2 are indicative of a favorable prognosis in ccRCC.

High peripheral neutrophil and monocyte count distinguishes renal cell carcinoma from renal angiomyolipoma and predicts poor prognosis of renal cell carcinoma.

Background: The presence of peripheral inflammatory cells has been linked to the prognosis of cancer. This study aims to investigate the distinct roles of absolute neutrophil count (ANC) and absolute monocyte count (AMC) in differentiating renal cell carcinoma (RCC) from renal angiomyolipoma (RAML), as well as their prognostic significance in RCC. Methods: We conducted a comprehensive analysis of peripheral immune cell data, clinicopathological data, and tumor characteristics in patients diagnosed with RCC or RAML from January 2015 to December 2021. Receiver operating characteristic (ROC) curves, as well as univariate and multivariate analyses, were employed to assess the diagnostic utility of AMC and ANC in differentiating between RCC and RAML. Kaplan-Meier curve analysis was used to study the survival of RCC patients with different AMC and ANC. The prognostic value of AMC and ANC in RCC was investigated using COX univariate and multivariate analysis. The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases were used for bioinformatic correlation analysis. Results: A total of 1120 eligible patients were included in the study. The mean preoperative AMC and ANC in patients with RCC were found to be significantly higher compared to those in patients with RAML (P = 0.001 and P < 0.001, respectively). High preoperative AMC and ANC significantly correlated with smoking history, tumor length, gross hematuria, and high T Stage, N stage, and pathological grade. In multivariate analyses, an ANC> 3.205 *10^9/L was identified to be independently associated with the presence of RCC (HR = 1.618, P = 0.008). High AMC and ANC were significantly associated with reduced OS and PFS (P < 0.05), and ANC may be an independent prognostic factor. Public database analysis showed that signature genes of tumor-associated macrophages (TAMs) and tumor-associated neutrophils (TANs) were highly expressed in ccRCC. Conclusions: Elevated preoperative ANC and AMC can distinguish RCC from RAML and predict poor prognosis in patients with RCC. Furthermore, the signature genes of TAMs and TANs exhibit high expression levels in clear cell RCC.

Identification and validation of m6A-associated ferroptosis genes in renal clear cell carcinoma.

Urinary cancer is synonymous with clear cell renal cell carcinoma (ccRCC). Unfortunately, existing treatments for this illness are ineffective and unpromising. Finding novel ccRCC biomarkers is crucial to creating successful treatments. The Cancer Genome Atlas provided clear cell renal cell carcinoma transcriptome data. Functional enrichment analysis was performed on ccRCC and control samples' differentially expressed N6-methyladenosine RNA methylation and ferroptosis-related genes (DEMFRGs). Machine learning was used to find and model ccRCC patients' predicted genes. A nomogram was created for clear cell renal cell carcinoma patients. Prognostic genes were enriched. We examined patients' immune profiles by risk score. Our prognostic genes predicted ccRCC treatment drugs. We found 37 DEMFRGs by comparing 1913 differentially expressed ccRCC genes to 202 m6A RNA methylation FRGs. Functional enrichment analysis showed that hypoxia-induced cell death and metabolism pathways were the most differentially expressed methylation functional regulating genes. Five prognostic genes were found by machine learning: TRIB3, CHAC1, NNMT, EGFR, and SLC1A4. An advanced renal cell carcinoma nomogram with age and risk score accurately predicted the outcome. These five prognostic genes were linked to various cancers. Immunological cell number and checkpoint expression differed between high- and low-risk groups. The risk model successfully predicted immunotherapy outcome, showing high-risk individuals had poor results. NIACIN, TAE-684, ROCILETINIB, and others treat ccRCC. We found ccRCC prognostic genes that work. This discovery may lead to new ccRCC treatments.

Laparoscopic pyelotomy combined with ultrasonic lithotripsy via a nephroscope for the treatment of complex renal stones.

The purpose of the study was to introduce a novel surgical approach of combining laparoscopic pyelotomy with ultrasonic lithotripsy via a nephroscope for the treatment of complex renal stones. Between May 2021 and April 2023, 32 patients underwent laparoscopic pyelotomy combined with ultrasonic lithotripsy via a nephroscope and their perioperative variables were retrospectively collected and outcomes were assessed. Dissection and incision of the anterior renal pelvis wall was performed via a laparoscope. A 19.5 F nephroscope was introduced into the renal pelvis through a laparoscopic trocar from the incision. Stones were fragmented and sucked out using a 3.3 mm ultrasonic probe placed through the nephroscope. All operations were completed successfully and the stone-free rate at 3 days after operation was 87.5% (28/32). Four (12.5%, 4/32) patients with staghorn stones had a small residual stone in the lower calyx after operation and did not require reintervention. No patient required perioperative transfusion and four (12.5%, 4/32) patients with struvite stones developed postoperative fever, which was successfully treated with intravenous antibiotics. The mean follow-up time was 14.0 ± 7.2 months, with no patient developing long-term complications. This approach offers a safe and effective treatment option for complex renal stones, as the method exhibits a high clearance rate with few complications.

PBRM1 mutation and WDR72 expression as potential combinatorial biomarker for predicting the response to Nivolumab in patients with ccRCC: a tumor marker prognostic study.

PURPOSE: Immune checkpoint therapy (ICT) provides a new idea for the treatment of advanced clear cell renal cell carcinoma (ccRCC), which can bring significant benefits to patients. However, the clinical application of ICT is limited because of the lack of predictive biomarkers to select potential responders. This study aims to propose a new biomarker to predict the response to Nivolumab in patients with ccRCC. MATERIALS AND METHODS: The genes that significantly improve the prognosis of ccRCC were retrieved from The Cancer Genome Atlas (TCGA) database. The genomic and clinical data were from patients that had been registered in prospective clinical trials (CheckMate 009, CheckMate 010 and CheckMate 025). TCGA, Gene Expression Omnibus (GEO), and The Human Protein Atlas database were used to analyze the gene and protein expression of WD repeat-containing protein 72 (WDR72) in ccRCC. Gene Ontology (GO) & The Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Set Enrichment Analysis (GSEA) were performed to dig relevant mechanisms of WDR72. Single sample gene set enrichment analysis (ssGSEA) was conducted to evaluate the role of WDR72 in immune infiltration. Cell proliferation assay, FAO and ATP quantification were used to explore and verify the molecular mechanisms. The expression of WDR72, FOXP3, CD8, and CPT1A was examined by IHC in 20 advanced ccRCC tissue samples at the Urology Department of our hospital. The MethSurv was used to identify PBRM1 and WDR72 gene methylation and its effect on prognosis of ccRCC. RESULTS: WDR72 is the most significant gene for improving overall survival (OS) in ccRCC. In all three checkmates, OS and progression free survival (PFS) were found to be significantly higher in WDR72 high expression group than that in WDR72 low expression group (P=0.040 and P=0.012, respectively), and similar conclusions could be drawn from the PBRM1-mutation (MUT) compared with the PBRM1-wildtype (WT) (P=0.007 and P=0.006, respectively). What's more, high expression of WDR72 plus PBRM1-MUT as a combinatorial biomarker showed improved OS (HR=0.388, P=0.0026) and PFS (HR=0.39, P=0.0066) compared to low expression of WDR72 plus PBRM1-WT. Functional enrichment analysis showed that WDR72 was closely positively related to fatty acid degradation and fatty acid beta oxidation pathway in ccRCC. In vitro experiments showed that high expression of WDR72 can promote fatty acids oxidation and inhibit the proliferation of ccRCC cells. Immune analysis revealed that WDR72 high expression was associated with decreased infiltration of Treg cells and low ssGSEA score of check-point. IHC results showed that WDR72 was negatively correlated with FOXP3 expression (r=-0.506, P=0.023) and positively correlated with CPT1A expression (r=0.529, P=0.017). CONCLUSIONS: The present study indicated that high expression of WDR72 may indicate a good prognosis of patients treated with Nivolumab and WDR72 expression combined with PBRM1 mutation could be more persuasive to predict the response for ICT in ccRCC patients.

Dyskerin and telomerase RNA component are sex-differentially associated with outcomes and Sunitinib response in patients with clear cell renal cell carcinoma.

BACKGROUND: Clear cell renal cell carcinoma (ccRCC) displays sex-biased incidence, outcomes, molecular alterations and treatment efficacy; however, clinical managements are largely identical in male and female patients. Moreover, many biomarkers have been identified as predictors for ccRCC outcomes and response to therapeutic drugs, such as multitargeted tyrosine-kinase receptor (TKR) inhibitors, but little is known about their sex-specificity. Dyskerin (DKC1), encoded by the DKC1 gene within Xq28, is a telomerase co-factor stabilizing telomerase RNA component (TERC) and overexpressed in various cancers. Here, we determined whether DKC1 and/or TERC affect ccRCC sex-differentially. METHODS: DKC1 and TERC expression in primary ccRCC tumors was assessed using RNA sequencing and qPCR. DKC1 association with molecular alterations and overall or progression-free survival (OS or PFS) was analyzed in the TCGA cohort of ccRCC. The IMmotion 151 and 150 ccRCC cohorts were analyzed to evaluate impacts of DKC1 and TERC on Sunitinib response and PFS. RESULTS: DKC1 and TERC expression was significantly upregulated in ccRCC tumors. High DKC1 expression predicts shorter PFS independently in female but not male patients. Tumors in the female DKC1-high group exhibited more frequent alterations in PIK3CA, MYC and TP53 genes. Analyses of the IMmotion 151 ccRCC cohort treated with the TKR inhibitor Sunitinib showed that female patients in the DKC1-high group was significantly associated with lower response rates (P = 0.021) accompanied by markedly shortened PFS (6.1 vs 14.2 months, P = 0.004). DKC1 and TERC expression correlated positively with each other, and higher TERC expression predicted poor Sunitinib response (P = 0.031) and shorter PFS (P = 0.004), too. However, DKC1 rather than TERC acted as an independent predictor (P < 0.001, HR = 2.0, 95% CI 1.480-2.704). In male patients, DKC1 expression was associated with neither Sunitinib response (P = 0.131) nor PFS (P = 0.184), while higher TERC levels did not predict response rates. Similar results were obtained from the analysis of the Sunitinib-treated IMmotion 150 ccRCC patients. CONCLUSIONS: DKC1 serves as an independent female-specific predictor for survival and Sunitinib efficacy in ccRCC, which contribute to better understanding of the sex-biased ccRCC pathogenesis and improve personalized interventions of ccRCC.

Many types of cancer including clear cell renal cell carcinoma (ccRCC) are known to display sex-biased survival, genomic alterations and treatment efficacy; however, clinical managements are largely identical in male and female ccRCC patients. Many molecules have been identified as predictors for ccRCC survival and response to therapeutic drugs, such as multitargeted tyrosine-kinase receptor inhibitor Sunitinib, but little is known about their sex-specificity. Dyskerin (DKC1), encoded by the DKC1 gene on X chromosome, is a telomerase co-factor stabilizing telomerase RNA component (TERC), whereas telomerase plays key roles in cancer development and progression. In this study, we observed increased DKC1 expression in ccRCC tumors. High DKC1 expression predicts shorter disease progression-free survival (PFS) in female but not male patients. Oncogene activation and tumor suppressor inactivation are more frequent in the female DKC1-high tumors. By analyzing two cohorts of ccRCC patients treated with Sunitinib, we showed that female patients in the DKC1-high group was significantly associated with lower response rates accompanied by markedly shortened PFS. DKC1 and TERC expression correlated positively with each other, and higher TERC expression predicted poor Sunitinib response and shorter PFS, too. However, DKC1 rather than TERC acted as an independent predictor. In male patients, DKC1 expression was associated with neither Sunitinib response nor PFS. Thus, DKC1 serves as a female-specific predictor for survival and Sunitinib response in ccRCC. Our findings are expected to improve personalized management of ccRCC.

Absolute monocyte count has a diagnostic role in distinguishing tumor marker-negative TGCT from benign testicular tumor via CCL2 regulation.

Clinically, for testicular tumor patients with negative tumor markers, how to distinguish the malignant from the benign is a difficult problem. This study aimed to assess the clinical significance of the absolute monocyte count (AMC) in differential diagnosis of testicular germ cell tumor with stage S0 (TGCTS0) and benign testicular tumor. In this retrospective single-center study, a total of 90 patients newly diagnosed with benign testicular tumor or TGCTS0 were reviewed. All patients received surgical intervention as the primary treatment method. AMC and other clinicopathological parameters were analyzed. Receiver operating characteristic (ROC) curves were constructed to assess the diagnostic power of investigated parameters, and to determine the optimal cutoff values. Kaplan-Meier curve analysis was used to study the survival of patients with TGCTS0. qRT-PCR and immunohistochemistry (IHC) were performed to examine the expression of C-C motif chemokine ligand 2 (CCL2) mRNA and protein respectively. Differential gene expression and functional enrichment analysis were performed using Gene Expression Omnibus and the Cancer Genome Atlas databases. The mean preoperative AMC in patients with TGCTS0 was significantly higher than that in patients with benign testicular tumor (P = .020). AMC > 0.485*10^9/L was identified to be associated with the presence of TGCTS0 (hazard ratio [HR] = 3.074, P = .026), and patients with higher AMC level had worse progression free survival (PFS) (P = .047). Furthermore, AMC combined with lactate dehydrogenase (LDH) achieved a better diagnostic efficacy for TGCTS0 (area under curve [AUC] = 0.695). Tumor-associated macrophages (TAMs) signature gene CCL2 was highly expressed in TGCT compared with normal testicular tissue. Functional enrichment analysis showed that CCL2 is closely involved in the Extracellular Matrix Organization pathway and positively correlated with the expression of various matrix metalloproteinases (MMPs). Elevated AMC may serve as a predictor of higher risk of TGCTS0, and CCL2 mediated TAMs infiltration and MMPs secretion is essential for the tumorigenesis of TGCT.

The cuproptosis-associated 11 gene signature as a predictor for outcomes and response to Bacillus Calmette-Guerin and immune checkpoint inhibitor therapies in bladder carcinoma.

Bladder cancer (BC) or carcinoma (BLCA) is predominantly derived from urothelium and includes non-muscle invasive BC (NMIBC) and muscle invasive BC (MIBC). Bacillus Calmette-Guerin (BCG) has long been applied for NMIBC to effectively reduce disease recurrence or progression, whereas immune checkpoint inhibitors (ICIs) were recently introduced to treat advanced BLCA with good efficacy. For BCG and ICI applications, reliable biomarkers are required to stratify potential responders for better personalized interventions, and ideally, they can replace or reduce invasive examinations such as cystoscopy in monitoring treatment efficacy. Here we developed the cuproptosis-associated 11 gene signature (CuAGS-11) model to accurately predict survival and response to BCG and ICI regimens in BLCA patients. In both discovery and validation cohorts where BLCA patients were divided into high- and low-risk groups based on a median CuAGS-11 score as the cutoff, the high-risk group was associated with significantly shortened overall survival (OS) and progression-free survival (PFS) independently. The survival predictive accuracy was comparable between CuAGS-11 and stage, and their combination-based nomograms showed high consistence between predicted and observed OS/PFS. The analysis of 3 BLCA cohorts treated with BCG unveiled lower response rates and higher frequencies of recurrence or progression coupled with shorter survival in CuAGS-11 high-risk groups. In contrast, almost none of patients underwent progression in low-risk groups. In IMvigor210 cohort of 298 BLCA patients treated with ICI Atezolizumab, complete/partial remissions were 3-fold higher accompanied by significantly longer OS in the CuAGS-11 low- than high-risk groups (P = 7.018E-06). Very similar results were obtained from the validation cohort (P = 8.65E-05). Further analyses of Tumor Immune Dysfunction and Exclusion (TIDE) scores revealed that CuAGS-11 high-risk groups displayed robustly higher T cell exclusion scores in both discovery (P = 1.96E-05) and validation (P = 0.008) cohorts. Collectively, the CuAGS-11 score model is a useful predictor for OS/PFS and BCG/ICI efficacy in BLCA patients. For BCG-treated patients, reduced invasive examinations are suggested for monitoring the CuAGS-11 low-risk patients. The present findings thus provide a framework to improve BLCA patient stratification for personalized interventions and to reduce invasive monitoring inspections.

The chromosomal instability 25 gene signature is identified in clear cell renal cell carcinoma and serves as a predictor for survival and Sunitinib response.

Background: Chromosomal instability (CIN) is a cancer hallmark and it is difficult to directly measure its phenotype, while a CIN25 gene signature was established to do so in several cancer types. However, it is currently unclear whether there exists this signature in clear cell renal cell carcinoma (ccRCC), and if so, which biological and clinical implications it has. Methods: Transcriptomic profiling was performed on 10 ccRCC tumors and matched renal non-tumorous tissues (NTs) for CIN25 signature analyses. TCGA and E-MBAT1980 ccRCC cohorts were analyzed for the presence of CIN25 signature, CIN25 score-based ccRCC classification, and association with molecular alterations and overall or progression-free survival (OS or PFS). IMmotion150 and 151 cohorts of ccRCC patients treated with Sunitinib were analyzed for the CIN25 impact on Sunitinib response and survival. Results: The transcriptomic analysis of 10 patient samples showed robustly upregulated expression of the CIN25 signature genes in ccRCC tumors, which were further confirmed in TCGA and E-MBAT1980 ccRCC cohorts. Based on their expression heterogeneity, ccRCC tumors were categorized into CIN25-C1 (low) and C2 (high) subtypes. The CIN25-C2 subtype was associated with significantly shorter patient OS and PFS, and characterized by increased telomerase activity, proliferation, stemness and EMT. The CIN25 signature reflects not only a CIN phenotype, but also levels of the whole genomic instability including mutation burden, microsatellite instability and homologous recombination deficiency (HRD). Importantly, the CIN25 score was significantly associated with Sunitinib response and survival. In IMmotion151 cohort, patients in the CIN25-C1 group exhibited 2-fold higher remission rate than those in the CIN25-C2 group (P = 0.0004) and median PFS in these two groups was 11.2 and 5.6 months, respectively (P = 7.78E-08). Similar results were obtained from the IMmotion150 cohort analysis. Higher EZH2 expression and poor angiogenesis, well characterized factors leading to Sunitinib resistance, were enriched in the CIN25-C2 tumors. Conclusion: The CIN25 signature identified in ccRCC serves as a biomarker for CIN and other genome instability phenotypes and predicts patient outcomes and response to Sunitinib treatment. A PCR quantification is enough for the CIN25-based ccRCC classification, which holds great promises in clinical routine application.

Widespread genomic/molecular alterations of DNA helicases and their clinical/therapeutic implications across human cancer.

DNA helicases are essential to genomic stability by regulating DNA metabolisms and their loss-of-function mutations lead to genomic instability and predisposition to cancer. Paradoxically, overexpression of DNA helicases is observed in several cancers. Here we analyzed genomic and molecular alterations in 12 important DNA helicases in TCGA pan-cancers to provide an overview of their aberrations. Significant expression heterogeneity of 12 DNA helicases was observed. We calculated DNA helicase score (DHS) based on their expression, and categorized tumors into high, low and intermediate subtypes. High DHS subtypes were robustly associated with stemness, proliferation, hyperactivated oncogenic signaling, longer telomeres, total mutation burden, copy number alterations (CNAs) and shorter survival. Importantly, tumors with high DHSs exhibited stronger expression of alternative end-join (alt-EJ) factors, indicative of sensitivity to chemo- and radio-therapies. High DHSs were also associated with homologous recombination deficiency (HRD), BRCA1/2 mutations and sensitivity to PARP inhibitors. Moreover, several drugs are identified to inhibit DNA helicases, with the Auror A kinase inhibitor Danusertib as the strongest candidate that was confirmed experimentally. The aberrant expression of DNA helicases was associated with CNAs, DNA methylation and m6A regulators. Our findings thus reveal widespread dysregulation of DNA helicases and their broad connection with featured oncogenic aberrations across human cancers. The close association of DHS with the alt-EJ pathway and HRD, and identification of Danusertib as a putative DNA helicase inhibitor have translational significance. Taken together, these findings will contribute to DNA helicase-based cancer therapy.

Genomic, epigenomic, and transcriptomic signatures for telomerase complex components: a pan-cancer analysis.

Telomerase activation is required for malignant transformation. Recent advances in high-throughput technologies have enabled the generation of complex datasets, thus providing alternative approaches to exploring telomerase biology more comprehensively, which has proven to be challenging due to the need for laborious assays required to test for telomerase activity. To solve these issues, several groups have analyzed TCGA pan-cancer tumor datasets by investigating telomerase reverse transcriptase (TERT), the catalytic subunit for telomerase activity, or its surrogates. However, telomerase is a multiunit complex containing not only TERT, but also numerus cofactors required for telomerase function. Here we determined genomic and molecular alterations of 10 well-characterized telomerase components in the TCGA and CCLE datasets. We calculated a telomerase score (TS) based on their expression profiles and clustered tumors into low, high, and intermediate subtypes. To validate the in silico analysis result, we used immunoblotting and telomerase assays. High TS subtypes were significantly associated with stemness, proliferation, epithelial to mesenchymal transition, hyperactivation of oncogenic signaling pathways, shorter patient survival, and infiltration of dysfunctional T-cells or poor response to immunotherapy. Copy number alterations in 10 telomerase components were widespread and associated with the level of their expression. Surprisingly, primary tumors and cancer cell lines frequently displayed a homozygous deletion of the TCAB1 gene, encoding a telomerase protein essential for telomerase trafficking, assembling, and function, as previously reported. However, tumors or cells carrying a TCAB1 deletion still exhibited telomerase activity comparable to or even higher than their wildtype counterparts. Collectively, applying telomerase component-based TS in complex datasets provided a robust tool for telomerase analyses. Our findings also reveal a tight connection between telomerase and other oncogenic signaling pathways; TCAB1 may acts as a dispensable telomerase component. Moreover, TS may serve as a useful biomarker to predict patient outcomes and response to immunotherapy.

The cuproptosis-associated 13 gene signature as a robust predictor for outcome and response to immune- and targeted-therapies in clear cell renal cell carcinoma.

Cuproptosis, the newly identified form of regulatory cell death (RCD), results from mitochondrial proteotoxic stress mediated by copper and FDX1. Little is known about significances of cuproptosis in oncogenesis. Here we determined clinical implications of cuproptosis in clear cell renal cell carcinoma (ccRCC). Based on the correlation and survival analyses of cuproptosis-correlated genes in TCGA ccRCC cohort, we constructed a cuproptosis-associated 13 gene signature (CuAGS-13) score system. In both TCGA training and two validation cohorts, when patients were categorized into high- and low-risk groups according to a median score as the cutoff, the CuAGS-13 high-risk group was significantly associated with shorter overall survival (OS) and/or progression-free survival (PFS) independently (P<0.001 for all). The CuAGS-13 score assessment could also predict recurrence and recurrence-free survival of patients at stage I - III with a high accuracy, which outperformed the ccAccB/ClearCode34 model, a well-established molecular predictor for ccRCC prognosis. Moreover, patients treated with immune checkpoint inhibitors (ICIs) acquired complete/partial remissions up to 3-time higher coupled with significantly longer PFS in the CuAGS-13 low- than high-risk groups in both training and validation cohorts of ccRCCs (7.2 - 14.1 vs. 2.1 - 3.0 months, P<0.001). The combination of ICI with anti-angiogenic agent Bevacizumab doubled remission rates in CuAGS-13 high-risk patients while did not improve the efficacy in the low-risk group. Further analyses showed a positive correlation between CuAGS-13 and TIDE scores. We also observed that the CuAGS-13 score assessment accurately predicted patient response to Sunitinib, and higher remission rates in the low-risk group led to longer PFS (Low- vs. high-risk, 13.9 vs. 5.8 months, P = 5.0e-12). Taken together, the CuAGS-13 score assessment serves as a robust predictor for survival, recurrence, and response to ICIs, ICI plus anti-angiogenic drugs and Sunitinib in ccRCC patients, which significantly improves patient stratifications for precision medicine of ccRCC.

GABPA-activated TGFBR2 transcription inhibits aggressiveness but is epigenetically erased by oncometabolites in renal cell carcinoma.

BACKGROUND: The ETS transcription factor GABPA has long been thought of as an oncogenic factor and recently suggested as a target for cancer therapy due to its critical effect on telomerase activation, but the role of GABPA in clear cell renal cell carcinoma (ccRCC) is unclear. In addition, ccRCC is characterized by metabolic reprograming with aberrant accumulation of L-2-hydroxyglurate (L-2HG), an oncometabolite that has been shown to promote ccRCC development and progression by inducing DNA methylation, however, its downstream effectors remain poorly defined. METHODS: siRNAs and expression vectors were used to manipulate the expression of GABPA and other factors and to determine cellular/molecular and phenotypic alterations. RNA sequencing and ChIP assays were performed to identify GABPA target genes. A human ccRCC xenograft model in mice was used to evaluate the effect of GABPA overexpression on in vivo tumorigenesis and metastasis. ccRCC cells were incubated with L-2-HG to analyze GABPA expression and methylation. We carried out immunohistochemistry on patient specimens and TCGA dataset analyses to assess the effect of GABPA on ccRCC survival. RESULTS: GABPA depletion, although inhibiting telomerase expression, robustly enhanced proliferation, invasion and stemness of ccRCC cells, whereas GABPA overexpression exhibited opposite effects, strongly inhibiting in vivo metastasis and carcinogenesis. TGFBR2 was identified as the GABPA target gene through which GABPA governed the TGFß signaling to dictate ccRCC phenotypes. GABPA and TGFBR2 phenocopies each other in ccRCC cells. Higher GABPA or TGFBR2 expression predicted longer survival in patients with ccRCC. Incubation of ccRCC cells with L-2-HG mimics GABPA-knockdown-mediated phenotypic alterations. L-2-HG silenced the expression of GABPA in ccRCC cells by increasing its methylation. CONCLUSIONS: GABPA acts as a tumor suppressor by stimulating TGFBR2 expression and TGFß signaling, while L-2-HG epigenetically inhibits GABPA expression, disrupting the GABPA-TGFß loop to drive ccRCC aggressiveness. These results exemplify how oncometabolites erase tumor suppressive function for cancer development/progression. Restoring GABPA expression using DNA methylation inhibitors or other approaches, rather than targeting it, may be a novel strategy for ccRCC therapy.

SETD8 stabilized by USP17 epigenetically activates SREBP1 pathway to drive lipogenesis and oncogenesis of ccRCC.

Recently, epigenetic modifications, including DNA methylation, histone modification and noncoding RNA (ncRNA)-associated gene silencing, have received increasing attention from the scientific community. Many studies have demonstrated that epigenetic regulation can render dynamic alterations in the transcriptional potential of a cell, which then affects the cell's biological function. The initiation and development of clear cell renal cell carcinoma (ccRCC), the most common subtype of renal cell cancer (RCC), is also closely related to genomic alterations by epigenetic modification. For ccRCC, lipid accumulation is one of the most typical characteristics. In other words, dysregulation of lipid uptake and synthesis occurs in ccRCC, which inversely promotes cancer proliferation and progression. However, the link among epigenetic alterations, lipid biosynthesis and renal cancer progression remains unclear. SETD8 is a histone methyltransferase and plays pivotal roles in cell cycle regulation and oncogenesis of various cancers, but its role in RCC is not well understood. In this study, we discovered that SETD8 was significantly overexpressed in RCC tumors, which was positively related to lipid storage and correlated with advanced tumor grade and stage and poor patient prognosis. Depletion of SETD8 by siRNAs or inhibitor UNC0379 diminished fatty acid (FA) de novo synthesis, cell proliferation and metastasis in ccRCC cells. Mechanistically, SETD8, which was posttranslationally stabilized by USP17, could transcriptionally modulate sterol regulatory element-binding protein 1 (SREBP1), a key transcription factor in fatty acid biosynthesis and lipogenesis, by monomethylating the 20th lysine of the H4 histone, elevating lipid biosynthesis and accumulation in RCC and further promoting cancer progression and metastasis. Taken together, the USP17/SETD8/SREBP1 signaling pathway plays a pivotal role in promoting RCC progression. SETD8 might be a novel biomarker and potential therapeutic target for treating RCC.

Berberine enhances gemcitabine­induced cytotoxicity in bladder cancer by downregulating Rad51 expression through inactivating the PI3K/Akt pathway.

Although gemcitabine (GEM) has been used to treat bladder cancer (BC) for a number of years, severe adverse events or drug resistance frequently develops. A series of drugs have been proved to sensitize patients to GEM and reduce the side effects. The aim of the present study was to evaluate the potential effects of berberine (BER) on GEM­induced cytotoxicity in BC and to explore the possible underlying mechanisms. T24 and 5637 human BC cell lines were treated with GEM and/or BER before cell proliferation, apoptosis and migration were studied. Oncomine databases and Gene Expression Profiling Interactive Analysis (GEPIA) were used to retrieve RAD51 recombinase (Rad51) mRNA expression. Overexpression plasmid or specific Rad51 small interfering RNA were used to examine the role of Rad51 in drug­treated BC cells. BC model mice were administered with GEM and/or BER before changes in tumor volume, size and Ki67 expression were assessed. BER enhanced GEM­induced cytotoxicity, apoptosis and inhibition of migration, whilst attenuating the GEM­induced upregulation of phosphorylated Akt and Rad51 expression. According to Oncomine and GEPIA analyses, Rad51 was found to be significantly upregulated in BC tissues compared with that in normal tissues, where there was a weak positive correlation between Rad51 and Akt1 expression. Knockdown of Rad51 enhanced GEM­induced cytotoxicity, whilst overexpression of Rad51 reversed the suppressed cell viability induced by BER and GEM. Inactivation of the PI3K/Akt pathway by LY294002 or BER enhanced GEM­induced cytotoxicity and downregulated Rad51 expression, whilst overexpression of constitutively active Akt restored Rad51 expression and cell viability that was previously decreased by BER and GEM. BER additively inhibited tumor growth and Ki67 expression when combined with GEM in vivo. These results suggest that BER can enhance GEM­induced cytotoxicity in BC by downregulating Rad51 expression through inactivating the PI3K/Akt pathway, which may represent a novel therapeutic target for BC treatment.

Clinical, Pathological, and Prognostic Analysis of Urachal Carcinoma.

OBJECTIVE: The aim of this study was to improve understanding the clinical, pathologic, and prognostic features of urachal carcinoma (UrC), a retrospectively descriptive study was done in 2 clinical centers. METHODS: After excluding the 2 missed patients, the clinical and pathological data of 59 patients with UrC, who were diagnosed or treated at 2 clinical centers between 1986 and 2019, was retrospectively analyzed. SPSS 22.0 (IBM) and GraphPad Prism 8.0.1 were used for statistics and data visualization. Survival data were analyzed by the Kaplan-Meier method and Log-rank tests. Cox proportional hazards regression were performed for find risk factors on predicting the prognosis. RESULTS: Of all 59 patients, 47 were male and 12 were female. The median age at diagnosis was 51.6 years (range: 22-84 years). Gross hematuria was the most common symptom (79.66%). The majority of urachal neoplasms were adenocarcinomas (94.92%). Forty-two patients (72.41%) underwent extended partial cystectomy with en bloc resection of the entire urachus. The mean follow-up was 52 months (3-277 months). Median overall survival was 52.8 months (4-93 months). The 3-year cancer-specific survival (CSS) rate and 5-year CSS rate were 69.1% and 61.2%. There was no significant difference among localized T stage, tumor histologic grade and surgical procedures in determining prognosis by survival analyze. While patients with high-risk TNM stage (local abdominal metastasis, lymph node metastasis, or distant metastasis) (p = 0.003) and positive surgical margin (p < 0.001) had significantly worse prognosis. CONCLUSIONS: The results indicate that high-risk TNM stage and positive surgical margin are risk predictors of prognosis. Localized T stage, histologic grade, and surgical procedure cause no significant effect on patient prognosis. The extended partial cystectomy is the recommended surgical approach for patients with UrC. Active multimodal treatments may improve the survival of patients with recurrent and metastatic disease.

FTO promotes tumour proliferation in bladder cancer via the FTO/miR-576/CDK6 axis in an m6A-dependent manner.

The aberrant expression of fat mass and obesity-associated protein (FTO) has been confirmed to be associated with a variety of cancers and participates in the regulation of multiple biological behaviours. FTO plays an oncogenic role in bladder cancer, but few studies have focused on how FTO promotes bladder cancer progression by regulating miRNA synthesis. Here, we confirmed that FTO expression was significantly increased in bladder cancer and was associated with a poor prognosis. FTO overexpression promoted bladder cancer cell proliferation, whereas FTO knockdown inhibited bladder cancer cell proliferation. We also demonstrated that FTO promoted bladder cancer cell proliferation via the FTO/miR-576/CDK6 pathways. Taken together, our work revealed that FTO plays a critical role in bladder cancer and could be a potential diagnostic or prognostic biomarker for this disease.

Multi-Omics Analysis of the Expression and Prognosis for FKBP Gene Family in Renal Cancer.

BACKGROUND: The FK506-binding protein (FKBP) is a family of intracellular receptors that can bind specifically to the immunosuppressant FK506 and rapamycin. Although FKBPs play crucial roles in biological processes and carcinogenesis, their prognostic value and molecular mechanism in clear cell renal cell carcinoma (ccRCC) remain unclear. METHODS: Using pan-cancer data from The Cancer Genome Atlas (TCGA) and public databases, we analyzed the expression and correlation of FKBPs in 33 tumor types. Survival and Cox regression analyses were employed to explore the prognostic value of FKBPs. The relationship with tumor microenvironment and stemness indices was taken into account to evaluate the function of FKBPs. We constructed a risk score model to predict the prognosis of patients with ccRCC. The receiver operating characteristic (ROC) curve was performed to further test the prognostic ability of our model. Nomogram, joint effects analysis, and clinical relevance were performed to assist the clinician. Gene set enrichment analysis (GSEA) and cell line experiments were performed to investigate the function and molecular mechanisms of FKBPs in patients with ccRCC. Paired clinical specimens and multi-omics analysis were used to further validate and explore the factors affecting gene expression in ccRCC patients. RESULTS: The expression levels of FKBP10 and FKBP11 were higher in ccRCC tissues than in normal tissues. The alteration in expression may be because of the degree of DNA methylation. Increased expression levels of FKBP10 and FKBP11 were associated with worse overall survival (OS). More importantly, GSEA revealed that FKBP10 is mainly involved in cell metabolism and autophagy, whereas FKBP11 is mainly associated with immune-related biological processes and autophagy. Cell Counting Kit 8 (CCK-8) and Transwell assays revealed that knockdown of FKBP10 and FKBP11 inhibits proliferation, migration, and invasion of the ccRCC cell line. CONCLUSION: FKBP10 and FKBP11 play important roles in ccRCC phenotypes and are potential prognostic markers as well as new therapeutic targets for patients with ccRCC.

Regulatory region mutations of TERT, PLEKHS1 and GPR126 genes as urinary biomarkers in upper tract urothelial carcinomas.

Background: The hotspot regulatory region mutations of the TERT, PLEKHS1 and GPR126 genes have been shown to occur frequently in urothelial bladder carcinoma (UBC). However, it is currently unclear whether these mutations are all present in upper tract urothelial carcinomas (UTUC) including renal pelvic carcinoma (RPC) and ureter carcinoma (UC), although TERT promoter mutations were previously observed in these malignancies. Methods: The hotspot mutations of TERT and PLEKHS1 promoters and GPR126 intron 6 (enhancer) in tumors derived from 164 patients with UTUC were determined using Sanger sequencing, and the obtained results were further compared with the mutation frequency in 106 UBCs. The mutations were also assessed in urine from patients with UTUC and UBC. Results: The mutation frequencies in UTUC tumors were 28%, 5.8% and 11% for TERT and PLEKHS1 promoters and GPR126 intron 6, respectively, which were lower than those (44.3%, 26.4%, and 31.4%, respectively) in UBCs. The total frequencies for the presence of any of these mutations were 50.8% and 34.4% for RPCs and UCs, respectively. All these mutated DNA sequences were detectable in urine from both UTUC and UBC patients and disappeared rapidly in most patients after surgery. Conclusions: This proof-of-concept study demonstrates that the hotspot mutations in the TERT, PLEKHS1 and GPR126 non-coding regions are present in UTUCs, and that urinary assays of these mutated sequences serve as potential biomarkers for UTUC diagnostics and disease monitoring.