Survival In ccRCC Research Articles

TRIM2 downregulation in clear cell renal cell carcinoma affects cell proliferation, migration, and invasion and predicts poor patients' survival.

Background and aimTripartite motif containing (TRIM) family protein has been involved in multiple pathogenesis of cancers. TRIM2 is a member of the family, and its role in clear cell renal cell carcinoma (ccRCC) remains to be unclarifid. Here, we showed the clinical value and biological role of TRIM2 in ccRCC.MethodsROC curves analyzed the clinicopathological parameters, Kaplan-Meier survival analysis determined the correlation of OS and DFS time, multivariate analysis demonstrated the prognostic indicator in overall survival and disease-free survival of ccRCC with TRIM2 expression in The Cancer Genome Atlas Kidney Clear Cell Carcinoma (TCGA-KIRC) database. Western blotting and immunohistochemistry were used to check the level of TRIM2 expression. Gain-of-function assay by exogenous overexpression of TRIM2 studied the biological role of TRIM2 in renal cell carcinoma cells.ResultsTRIM2 expression was associated with various clinicopathologicalfactors and lower TRIM2 expression was interrelated to a poor prognosis. The levels of TRIM2 expression were also scanty in ccRCC tissues and renal cancer cell lines than in normal control. The biological role of TRIM2 in ccRCC was identifid by bioinformatics analysis and functional analysis. Exogenous overexpression of TRIM2 with the gain-of-function assay in renal cell carcinoma cells showed that the cell proliferation, migration, and invasion were signifiantly suppressed.ConclusionThese results showed that TRIM2 acted as an antitumor gene and a specifi prognostic indicator for patients with ccRCC, which indicated that positive modulation of TRIM2 might be a novel treatment strategy for ccRCC.

Endogenous retroviral signatures predict immunotherapy response in clear cell renal cell carcinoma.

Human endogenous retroviruses (hERVs) are remnants of exogenous retroviruses that have integrated into the genome throughout evolution. We developed a computational workflow, hervQuant, which identified more than 3,000 transcriptionally active hERVs within The Cancer Genome Atlas (TCGA) pan-cancer RNA-Seq database. hERV expression was associated with clinical prognosis in several tumor types, most significantly clear cell renal cell carcinoma (ccRCC). We explored two mechanisms by which hERV expression may influence the tumor immune microenvironment in ccRCC: (i) RIG-I-like signaling and (ii) retroviral antigen activation of adaptive immunity. We demonstrated the ability of hERV signatures associated with these immune mechanisms to predict patient survival in ccRCC, independent of clinical staging and molecular subtyping. We identified potential tumor-specific hERV epitopes with evidence of translational activity through the use of a ccRCC ribosome profiling (Ribo-Seq) dataset, validated their ability to bind HLA in vitro, and identified the presence of MHC tetramer-positive T cells against predicted epitopes. hERV sequences identified through this screening approach were significantly more highly expressed in ccRCC tumors responsive to treatment with programmed death receptor 1 (PD-1) inhibition. hervQuant provides insights into the role of hERVs within the tumor immune microenvironment, as well as evidence that hERV expression could serve as a biomarker for patient prognosis and response to immunotherapy.

Loss of Nuclear BAP1 Expression Is Associated with High WHO/ISUP Grade in Clear Cell Renal Cell Carcinoma.

BackgroundBRCA1-associated protein 1 (BAP1) mutations are frequently reported in clear cell renal cell carcinoma (ccRCC); however, very few studies have evaluated the role of these mutations in other renal cell carcinoma (RCC) subtypes. Therefore, we analyzed BAP1 protein expression using immunohistochemistry in several RCC subtypes and assessed its relationship with clinicopathological characteristics of patients.MethodsBAP1 expression was immunohistochemically evaluated in tissue microarray blocks constructed from 371 samples of RCC collected from two medical institutions. BAP1 expression was evaluated based on the extent of nuclear staining in tumor cells, and no expression or expression in < 10% of tumor cells was defined as negative.ResultsLoss of BAP1 expression was observed in ccRCC (56/300, 18.7%), chromophobe RCC (6/26, 23.1%), and clear cell papillary RCC (1/4, 25%), while we failed to detect BAP1 expression loss in papillary RCC, acquired cystic disease-associated RCC, or collecting duct carcinoma. In ccRCC, loss of BAP1 expression was significantly associated with high World Health Organization (WHO)/International Society of Urological Pathology (ISUP) grade (p = .002); however, no significant correlation was observed between loss of BAP1 expression and survival in ccRCC. Loss of BAP1 expression showed no association with prognostic factors in chromophobe RCC.ConclusionsLoss of BAP1 nuclear expression was observed in both ccRCC and chromophobe RCC. In addition, BAP1 expression loss was associated with poor prognostic factors such as high WHO/ISUP grade in ccRCC.

Abstract 200: FOXD1 promotes stromal investment in clear cell renal cell carcinoma

Abstract Forkhead transcription factors (FOX) play a role in the development of many cancers, including clear cell renal cell carcinoma (ccRCC). FOXD1, essential for angiogenesis in the developing kidney, has not previously been implicated in ccRCC. Tumor microarrays (TMAs) containing 41 patient-derived ccRCC tumors and 26 healthy cortex samples were stained for FOXD1, with 65% of tumor samples staining positively. Due to the effect of FOXD1 on angiogenesis, staining for stromal markers PECAM, αSMA, PDGFRβ and NG2 was also performed. A direct correlation was found only between FOXD1 and PDGFRβ (p&amp;lt;4.5x10-6), suggesting interstitial fibroblast recruitment by FOXD1+ cancer cells. Increased expression of FOXD1 was found to correlate with increased stage (p=9.1x10-5) and reduced survival (FOXD1 low= 2830 days; FOXD1 high= 1913 days) from a Kaplan-Meier analysis of RNA-seq data from The Cancer Genome Atlas (TCGA). Potential FOXD1 target binding sites were determined using the TRANSFAC FOXD1 binding site matrix. SLIT2, a factor known to inhibit migration of pericytes, was uncovered from the analysis and was further found to be downregulated in response to FOXD1 overexpression in renal proximal tubule cells (RPTECs). To test the influence of SLIT2 on interstitialfibroblast migration, scratch assays on NRK-49F (rat kidney fibroblast) and Gli-1 (cardiac fibroblast) cells were performed and showed that SLIT2 reduced PDGFBB-induced cell migration (p&amp;lt;0.027). A multiplex proximity ligation assay for multiple signaling pathways showed that SLIT2 treatment reduced the STAT signaling response to PGDFBB. To model the influence of SLIT2 on ccRCC stromal invasion in vitro, we devised a novel 3D invasion assay. In summary, 786-O ccRCC cancer cells were seeded into a 200um thick silk scaffold and cultured for 3 days to allow deposition of ECM. Scaffolds were then placed on top of a monolayer of interstitial fibroblasts and invasion into the cancer cell-filled scaffold was measured using confocal microscopy. The results showed a significant reduction of cell invasion into the cancer-filled matrix with the addition of recombinant SLIT2 to media (5µm p=1.34x10-4; 50µm p=1.64x10-2). In conclusion, we show that FOXD1 expression has prognostic relevance to patient survival in ccRCC, and that this may be due to modulation of SLIT2 expression. Citation Format: Kyle H. Bond, Jennifer Fetting, Ashwani Gupta, Christine W. Duarte, Michele Karolak, Clare B. Congdon, Ivette Emery, Eoghainín Ó hAinmhire, Benjamin D. Humphreys, Leif Oxburgh. FOXD1 promotes stromal investment in clear cell renal cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 200.

126 - Loss of Epitranscriptomic Control of Seleno-cysteine Utilization Engages Senescence and Metabolic Reprogramming through mTOR Activation and Predicts Poor Survival in Clear Cell Renal Cell Cancer Patients

Over 14,000 Americans die each year from kidney cancer with clear-cell Renal Cell Carcinoma (ccRCC) being the most prevalent and malignant type of kidney cancer. Recent work has implicated disruptions in glutathione (GSH) metabolism as predictors of poor survival in ccRCC. Maintenance of the GSH redox cycle is reliant on the activities of selenocysteine-containing GSH metabolizing enzymes whose translation is controlled by the alkylation repair homolog 8 (Alkbh8) tRNA methyltransferase. In cells from Alkbh8-/- mice the absence of the tRNA methyltransferase Alkbh8 decreases the levels of many GSH metabolizing selenoproteins, promotes increased ROS and DNA damage levels, and confers enhanced sensitivity to oxidizing agents. Surprisingly the Alkbh8-/- mice reproduce, thrive normally but display phenotypic characteristics of premature ageing such as weight loss, reduced subcutaneous fat, alopecia (hair loss), kyphosis (curvature of the spine) when compared to their aged-matched WT control. Alkbh8-deficiency also triggers a senescence and metabolic gene signature that is predictive (Hazard Ratio 2.94, p

Characterization and Impact of TERT Promoter Region Mutations on Clinical Outcome in Renal Cell Carcinoma

BackgroundMutations in the promoter region of the TERT gene have been detected in a variety of cancers. These mutations can potentially lead to unlimited cell divisions and result in poor clinical prognosis. ObjectiveTo determine the role and relevance of TERT promoter region mutations in both clear cell (ccRCC) and non–clear cell (nccRCC) renal cell carcinoma using ultra-deep and whole-genome sequencing methods on primary tumor samples. Design, setting, and participantsDNA from 281 kidney tumors (147 ccRCC and 134 nccRCC) was sequenced between 2013 and 2015, and clinical outcomes for these patients from a single institution were retrospectively analyzed. Outcome measurements and statistical analysisDifferences in patient characteristics and mutational status were tested using Fisher's exact test for categorical variables and the Wilcoxon rank sum test for continuous variables. Survival times were estimated using the Kaplan-Meier method and differences were tested using the log-rank test. Results and limitationsTERT mutations occurred in 12.2% of ccRCC and 10.4% of nccRCC cases. In >80% of the cases, mutations were located at C228T and were found to co-occur only rarely with other relevant RCC driver genes. The median follow-up among survivors overall was 2.5 yr (range 0.1–18.3). TERT promoter mutations were significantly associated with cancer-specific survival in ccRCC (hazard ratio 2.68, 95% confidence interval 1.19–6.01; p=0.013). In nccRCC, TERT mutations were significantly associated with larger tumors and metastatic development. Assessment of further relevant clinical associations was precluded in the nccRCC group by the heterogeneous and small sample size. ConclusionsOur data suggests that TERT mutational status reflects a distinct pathogenesis with an aggressive disease course in RCC. Stratifying patients with this unique tumorigenesis that leads to poor clinical outcomes could be a putative target for novel therapeutics. Patient summaryWe show a previously unrecognized frequency of TERT promoter mutations in both clear cell and non–clear cell renal cell carcinoma. TERT promoter mutations were associated with some worse outcomes in patients with clear cell renal cell carcinoma.

IRF5 is associated with adverse postoperative prognosis of patients with non-metastatic clear cell renal cell carcinoma.

BackgroundIRF5 is one member of IRFs family, and is critical for host immunity and cell response. In the present study, we sought to search the clinical and prognostic value of IFR5 in patients with non-metastatic ccRCC.ResultsIRF5 proved to be an adverse independent prognostic factor for overall survival (p < 0.001) and recurrence free survival (p = 0.002). The newly built nomograms could give better prediction for overall survival and recurrence free survival in ccRCC patients.Materials and MethodsWe included 264 individuals who were diagnosed with non-metastatic clear cell renal cell carcinoma in the present study. Immunohistochemistry staining was performed on tissue microarrays to evaluate the IRF5 expression. χ2 test, Fisher's exact test, t test, Kaplan-Meier method and Cox proportional hazard model were applied to evaluate the prognostic value of IRF5. Two nomograms were constructed to predict clinical outcomes for ccRCC patients after surgery.ConclusionsIRF5 was an adverse independent prognostic factor for both overall survival and recurrence free survival in patients with non-metastatic ccRCC.

Intratumoral heterogeneity analysis reveals hidden associations between protein expression losses and patient survival in clear cell renal cell carcinoma.

Intratumoral heterogeneity (ITH) is a prominent feature of kidney cancer. It is not known whether it has utility in finding associations between protein expression and clinical parameters. We used ITH that is detected by immunohistochemistry (IHC) to aid the association analysis between the loss of SWI/SNF components and clinical parameters.160 ccRCC tumors (40 per tumor stage) were used to generate tissue microarray (TMA). Four foci from different regions of each tumor were selected. IHC was performed against PBRM1, ARID1A, SETD2, SMARCA4, and SMARCA2. Statistical analyses were performed to correlate biomarker losses with patho-clinical parameters. Categorical variables were compared between groups using Fisher's exact tests. Univariate and multivariable analyses were used to correlate biomarker changes and patient survivals. Multivariable analyses were performed by constructing decision trees using the classification and regression trees (CART) methodology. IHC detected widespread ITH in ccRCC tumors. The statistical analysis of the “Truncal loss” (root loss) found additional correlations between biomarker losses and tumor stages than the traditional “Loss in tumor (total)”. Losses of SMARCA4 or SMARCA2 significantly improved prognosis for overall survival (OS). Losses of PBRM1, ARID1A or SETD2 had the opposite effect. Thus “Truncal Loss” analysis revealed hidden links between protein losses and patient survival in ccRCC.

MiR-125b is associated with renal cell carcinoma cell migration, invasion and apoptosis.

MicroRNA (miR)-125b has been identified as deregulated in a number of types of cancer. Previous studies have detected the expression of miR-125b in clear cell renal cell carcinoma (ccRCC) tissues by in situ hybridization and revealed that miR-125b was upregulated in ccRCC tissues, and was associated with recurrence and survival of patients with ccRCC. However, the function of miR-125b in RCC remains unclear. Thus, the expression of miR-125b was detected with quantitative polymerase chain reaction (qPCR) in 24 paired RCC and adjacent normal tissues. The result of qPCR showed that miR-125b was upregulated in RCC tissues. Furthermore, the function of miR-125b in RCC (786-O and ACHN) cells was detected by transfecting miR-125 mimic or inhibitor to upregulate or downregulate miR-125b expression. Cell proliferation assays (MTT and Cell Counting Kit-8), cell mobility assays (cell scratch and Transwell assay) and a cell apoptotic assay (flow cytometry assay) were performed to assess the function of miR-125b on RCC cells. Results from the assays demonstrated that overexpression of miR-125b could promote cell migration and invasion, and reduce the cell apoptotic rate. It was also revealed that downregulation of miR-125b could reduce cell migration and invasion, and induce cell apoptosis. However, the results of the cell proliferation assay revealed that miR-125b had no significant effect on cell proliferation. Not only could miR-125b predict recurrence and survival of ccRCC; the present study revealed that miR-125b could regulate RCC cell migration, invasion and apoptosis. Additional studies are required to determine the mechanism of miR-125b in RCC cells and define the target genes of miR-125b in RCC.

Perinephric white adipose tissue inflammation in clear cell renal cell carcinoma (ccRCC).

507 Background: High body mass index (BMI) is an established risk factor for developing ccRCC but is associated with better survival in clinical studies. The obesity paradox may be influenced by metabolically healthy patients classified as obese, and metabolically unhealthy patients classified as normal weight. We evaluated white adipose tissue inflammation (WATi) in perinephric fat as a new marker of metabolic dysregulation and examined its association with clinicopathological characteristics. Methods: In July 2015, we established a prospective cohort study at Memorial Sloan Kettering Cancer Center to investigate the prognostic significance of perinephric WATi among patients undergoing nephrectomy. Perinephric WAT is collected during surgery and patients are followed for clinical outcomes. WATi is defined by the presence of dead/dying adipocytes surrounded by macrophages forming crown-like structures (CLS) and detected through immunohistochemistry. Clinicopathological data are abstracted from the electronic medical record. Wilcoxon rank-sum, Chi-square, or Fisher’s exact tests describe the relationship between CLS status and clinicopathological characteristics on the first 38 ccRCC patients. Results: The study cohort had a median age of 56 years (range 47-64 years) and was predominantly male (71%). CLS were detected in 50% of patients, did not differ by age or sex, and were present in all BMI levels; 59% of obese, 38.5% of overweight, and 33% of normal weight patients (p = 0.47). CLS was significantly associated with advanced disease characteristics including higher stage (p = 0.03) and local invasion (p = 0.02). Median tumor size was larger in patients who were CLS+ (3.5 cm, range 2.65-5.75) than CLS- (2.2 cm, range 1.55-3.00; p = 0.02). Conclusions: Perinephric WATi was found in ccRCC patients of all BMI levels and associated with factors related to poor prognosis. Patients with occult inflammation may be at higher mortality risk, regardless of their BMI. Recruitment of additional cases and analyses to examine how CLS influences ccRCC survival are on-going.

Overexpression of G6PD Represents a Potential Prognostic Factor in Clear Cell Renal Cell Carcinoma.

Glucose-6-phosphate dehydrogenase (G6PD) participates in glucose metabolism and it acts as the rate-limiting enzyme of the pentose phosphate pathway (PPP). Recently, G6PD dysregulation has been found in a variety of human cancers. Through analyzing published data in The Cancer Genome Atlas (TCGA), our pilot study indicated that G6PD mRNA expression was significantly higher in advanced Fuhrman grade in clear cell renal cell carcinoma (ccRCC). These clues promoted us to further evaluate the expression profile of G6PD and its prognostic impact in patients with ccRCC. In this study, G6PD expression levels were analyzed in 149 human ccRCC and normal tissues using immunohistochemistry. The results showed that compared with that in the normal renal samples, G6PD was found highly expressed in 51.0% of ccRCC (p<0.05). High expression of G6PD was significantly correlated to tumor extent, lymph node metastasis, Fuhrman grade, and TNM stage of ccRCC (all p<0.05). Moreover, positive G6PD expression was associated with poorer overall survival in ccRCC (p<0.001). In Cox regression analyses, high expression of G6PD also could be an independent prognostic factor for overall survival in ccRCC (p=0.007). This study suggests that overexpression of G6PD is associated with advanced disease status and therefore may become an important prognosticator for poor outcomes in ccRCC, as well as a potential therapeutic target for developing effective treatment modalities.

Identification and validation of soluble carrier family expression signature for predicting poor outcome of renal cell carcinoma.

The soluble carrier (SLC) family plays an important role in cell metabolism. The purpose of the current study was to screen SLCs as potential prognostic factors in clear cell renal cell carcinoma (ccRCC). A total of 509 patients with ccRCC from The Cancer Genome Atlas (TCGA) cohort were enrolled in this study. The expression profile of SLCs was obtained from the TCGA RNAseq database. Metadata of the TCGA cohort, including age, sex, TNM stage, tumor grade, American Joint Committee on Cancer stage, laterality, and overall survival, were collected. Univariate and multivariate Cox proportional hazards regression models were used to analyze the relative factors. Prognosis-associated genes were further validated in a Fudan University Shanghai Cancer Center (FUSCC) cohort consisting of 178 patients. Among a total of 364 SLC transporters, 61 were independent predictors of ccRCC patient overall survival. Among the 61 SLC transporters, 26 were significantly downregulated and 23 were significantly upregulated in tumor tissues compared with non-malignant kidney tissues. Analyses of two open source, RNA expression data sets on sunitinib response revealed that SLC10A2 was downregulated in tyrosine kinase inhibitor-resistant samples. We validated SLC10A2 expression in the FUSCC cohort and showed that SLC10A2 expression was an independent prognostic predictor of overall survival of ccRCC (hazard ratio=0.432, 95% CI: 0.204-0.915). Our results identified a number of associations of SLC gene expression with prognosis of ccRCC patients, indicating that these genes may represent possible oncogenes that could serve as therapeutic targets of ccRCC.

A four-gene signature predicts survival in clear-cell renal-cell carcinoma.

Clear-cell renal-cell carcinoma (ccRCC) is the most common pathological subtype of renal cell carcinoma (RCC), accounting for about 80% of RCC. In order to find potential prognostic biomarkers in ccRCC, we presented a four-gene signature to evaluate the prognosis of ccRCC. SurvExpress and immunohistochemical (IHC) staining of tissue microarrays were used to analyze the association between the four genes and the prognosis of ccRCC. Data from TCGA dataset revealed a prognostic prompt function of the four genes (PTEN, PIK3C2A, ITPA and BCL3). Further discovery suggested that the four-gene signature predicted survival better than any of the four genes alone. Moreover, IHC staining demonstrated a consistent result with TCGA, indicating that the signature was an independent prognostic factor of survival in ccRCC. Univariate and multivariate Cox proportional hazard regression analysis were conducted to verify the association of clinicopathological variables and the four genes’ expression levels with survival. The results further testified that the risk (four-gene signature) was an independent prognostic factors of both Overall Survival (OS) and Disease-free Survival (DFS) (P<0.05). In conclusion, the four-gene signature was correlated with the survival of ccRCC, and therefore, may help to provide significant clinical implications for predicting the prognosis of patients.

A Prospective Observational Study for Assessment and Outcome Association of Circulating Endothelial Cells in Clear Cell Renal Cell Carcinoma Patients Who Show Initial Benefit from First-line Treatment. The CIRCLES (CIRCuLating Endothelial cellS) Study (SOGUG-CEC-2011-01)

BackgroundMarkers able to predict the response to antiangiogenics in metastatic clear cell renal cell carcinoma (ccRCC) are not available. The development of new treatment options like immunotherapy are reaching the clinic; therefore, predictors of benefit from these different available treatments are increasingly needed. ObjectiveIn this study, we prospectively assessed the association of circulating endothelial cells (CECs) in peripheral blood with long-term benefit from first-line treatment in ccRCC. Design, setting, and participantsA prospective observational study was designed involving 13 institutions of the Spanish Oncology Genitourinary Group. Adult patients diagnosed with advanced ccRCC who had achieved response or disease stabilization after 3 mo on first-line therapy were eligible. Outcome measurements and statistical analysisCECs were isolated from peripheral blood, captured with ferrofluids coated with monoclonal antibodies directed against the CD146 antigen, and assessed centrally with an automated standardized system. CECs were defined as 4′,6-diamidino-2-phenylindole+, CD105+, and CD45–. Blood samples were systematically taken every 6 wk for 15 mo or until tumor progression, whichever occurred first. Clinical data were externally monitored at all centers. Results and limitationsFrom August 9, 2011, to January 17, 2013, 75 patients were enrolled in the study. Patients with baseline CECs above the median showed a significantly longer progression-free survival than those with low CECs (22.2 mo vs 12.2 mo) with a hazard ratio of 2.5 (95% confidence interval: 1.2–5.3, p=0.016). There was no difference between CEC levels at baseline and at tumor progression (medians of 50 CECs/4ml and 52 CECs/4ml, respectively). ConclusionsUnder antiangiogenic treatment, the detection of higher CEC levels is associated with clinical benefit in terms of progression-free survival in ccRCC. Patient summaryAntiangiogenics are the cornerstone of treatment in kidney cancer. Since they target endothelial rather than tumor cells, we studied the correlation between levels of circulating endothelial cells in peripheral blood and long-term benefit in patients on antiangiogenic therapy. Higher levels were associated with long-term benefit, suggesting that this determination could help to separate best responders from those who could require a more intensive approach.

Cooperative Effect of miR-141-3p and miR-145-5p in the Regulation of Targets in Clear Cell Renal Cell Carcinoma.

BackgroundDue to the poor prognosis for advanced renal cell carcinoma (RCC), there is an urgent need for new therapeutic targets and for prognostic markers to identify high risk tumors. MicroRNAs (miRNAs) are frequently dysregulated in tumors, play a crucial role during carcinogenesis and therefore might be promising new biomarkers. In previous studies, we identified miR-141-3p and miR-145-5p to be downregulated in clear cell RCC (ccRCC). Our objective was to investigate the functional association of these miRNAs, focusing on the cooperative regulation of new specific targets and their role in ccRCC progression.MethodsThe effect of miR-141-3p and miR-145-5p on cell migration was examined by overexpression in 786-O cells. New targets of both miRNAs were identified by miRWalk, validated in 786-O and ACHN cells and additionally characterized in ccRCC tissue on mRNA and protein level.ResultsIn functional analysis, a tumor suppressive effect of miR-141-3p and miR-145-5p by decreasing migration and invasion of RCC cells could be shown. Furthermore, co-overexpression of the miRNAs seemed to result in an increased inhibition of cell migration. Both miRNAs were recognized as post-transcriptional regulators of the targets EAPP, HS6ST2, LOX, TGFB2 and VRK2. Additionally, they showed a cooperative effect again as demonstrated by a significantly increased inhibition of HS6ST2 and LOX expression after simultaneous overexpression of both miRNAs. In ccRCC tissue, LOX mRNA expression was strongly increased compared to normal tissue, allowing also to distinguish between non-metastatic and already metastasized primary tumors. Finally, in subsequent tissue microarray analysis LOX protein expression showed a prognostic relevance for the overall survival of ccRCC patients.ConclusionThese results illustrate a jointly strengthening effect of the dysregulated miR-141-3p and miR-145-5p in various tumor associated processes. Focusing on the cooperative effect of miRNAs provides new opportunities for the development of therapeutic strategies and offers novel prognostic and diagnostic capabilities.

MP71-08 SELECT CONCURRENT CHROMOSOME 3P MUTATIONS PREDICT WORSE OVERALL AND RECURRENCE-FREE SURVIVAL IN CLEAR CELL RENAL CARCINOMA IN THE CANCER GENOME ATLAS

You have accessJournal of UrologyKidney Cancer: Basic Research & Pathophysiology I1 Apr 2016MP71-08 SELECT CONCURRENT CHROMOSOME 3P MUTATIONS PREDICT WORSE OVERALL AND RECURRENCE-FREE SURVIVAL IN CLEAR CELL RENAL CARCINOMA IN THE CANCER GENOME ATLAS Christopher Keith, John Petros, Michael Rossi, and Rebecca Arnold Christopher KeithChristopher Keith More articles by this author , John PetrosJohn Petros More articles by this author , Michael RossiMichael Rossi More articles by this author , and Rebecca ArnoldRebecca Arnold More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2016.02.1454AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES In clear cell renal cell carcinoma (ccRCC), mutations in BAP1 and, in some cohorts, PBRM1 are associated with worse survival, while VHL and SETD2 mutations do not independently predict poor prognosis. While studies have analyzed prognostic impact of independent mutations on these chromosome 3p (chr 3p) genes, the prognostic significance of co-existing mutations in these near-neighboring genes is unknown. METHODS The Cancer Genome Atlas (TCGA) ccRCC database was probed for mutations in VHL, BAP1, SETD2, and PBRM1. Kaplan-Meier survival log-rank and Cox multivariate analysis were performed. RESULTS Of the 418 patients with ccRCC in TCGA, at least one of the four chr 3p genes was mutated in 74% of cases. Co-existing somatic mutations in VHL and BAP1 (n=16), VHL and SETD2 (n=9), or PBRM1 and SETD2 and/or BAP1 (n=13) compared to all other cases was associated with worse recurrence-free (1.83 vs. 6.0 years; p=0.0004) and overall survival (2.42 vs. 7.13 years; p<0.0001). The survival difference remained significant after controlling for total mutational burden (p<0.0001), BAP1 mutations (p=0.014), and PBRM1 mutations (p<0.0001). On multivariate analysis, high-risk chromosome 3p status was associated with significantly decreased recurrence-free survival and overall survival. Interestingly, the deleterious effects of BAP1 in the ccRCC TCGA cohort are negated when the group with only coexisting VHL or PBRM1 mutations is excluded (p=0.83). When comparing the high-risk BAP1 group to all other cases with BAP1 alterations, there is a significant difference in survival (2.42 vs. 5.38 years; p=0.01). CONCLUSIONS Concurrent mutations in VHL and BAP1, VHL and SETD2, or PBRM with SETD2 and/or BAP1 are associated with significantly decreased recurrence-free and overall survival in ccRCC in the TCGA. In addition, the deleterious effect of BAP1 mutations in TCGA is dependent on concurrent chr 3p mutations. Such information may be useful in risk stratification of patients with ccRCC. © 2016FiguresReferencesRelatedDetails Volume 195Issue 4SApril 2016Page: e918 Advertisement Copyright & Permissions© 2016MetricsAuthor Information Christopher Keith More articles by this author John Petros More articles by this author Michael Rossi More articles by this author Rebecca Arnold More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Promoter Methylation of CDO1 Identifies Clear-Cell Renal Cell Cancer Patients with Poor Survival Outcome.

In this era of molecular diagnostics, prediction of clear-cell renal cell cancer (ccRCC) survival requires optimization, as current prognostic markers fail to determine individual patient outcome. Epigenetic events are promising molecular markers. Promoter CpG island methylation of cysteine dioxygenase type 1 (CDO1), which was identified as prognostic marker for breast cancer, is studied as a potential marker for ccRCC survival. We collected primary tissues of 365 ccRCC cases identified within the prospective Netherlands Cohort Study (NLCS). In this population-based series, CDO1 promoter methylation was observed in 124 of 324 (38.3%) patients with successful methylation-specific PCR analysis. Kaplan-Meier curves and Wilcoxon tests were used to evaluate 10-year ccRCC-specific survival. Cox regression analysis was used to obtain crude and multivariate HRs and 95% confidence intervals (CI). The relative prognostic value of multivariate models with and without CDO1 promoter methylation was compared using likelihood-ratio tests. Patients with CDO1 promoter methylation have a significantly poorer survival than those without (Wilcoxon P = 0.006). Differences in survival were independent of other prognostic factors, including age and sex (HR, 1.66; 95% CI, 1.12-2.45) and TNM stage, tumor size, and Fuhrman grade (HR, 1.89; 95% CI, 1.25-2.85). Multivariate models performed better with than without CDO1 promoter methylation status (likelihood-ratio P = 0.003). Survival curves were validated in an independent series of 280 ccRCC cases from The Cancer Genome Atlas (TCGA; Wilcoxon P < 0.001). CDO1 promoter methylation may not substitute common prognostic makers to predict ccRCC survival, but offers additional, relevant prognostic information, indicating that it might be a novel molecular marker to determine ccRCC prognosis.

ITGA2B and ITGA8 are predictive of prognosis in clear cell renal cell carcinoma patients.

Integrins play an important role in cancer growth and metastasis. This study aimed at determining the predictive ability of integrins and associated genes identified through molecular network in clear cell renal cell carcinoma. A total of 525 patients with ccRCC from The Cancer Genome Atlas (TCGA) cohorts were collected in this study. The expression profile of integrins and related genes were obtained from the TCGA RNAseq database. Clinicopathological characteristics, including age, gender, tumor size, tumor node metastasis (TNM), tumor grade, stage, laterality, and overall survival were collected. Cox proportional hazards regression model as well as Kaplan-Meier curve were used to assess the relative factors. Genes of integrin family that showed certain correlations with overall survival (OS) were further validated in the Fudan University Shanghai Cancer Center (FUSCC) cohort. In the TCGA cohort, after Cox proportional hazards analysis, ITGA2B (hazards ratio (HR) = 1.232, 95% CI 1.097 to 1.383) and ITGA8 (HR = 0.804, 95% CI 0.696 to 0.930) were shown predictive of ccRCC prognosis. Low ITGA8 expression was associated with poor prognosis for OS (log-rank test, p < 0.0001), while high level of ITGA2B expression was correlated with poor prognosis for OS (log-rank test, p < 0.0001). This finding was validated in FUSCC cohort (log-rank test, all p < 0.05). As a result, low ITGA8 expression was associated with poor prognosis for OS (log-rank test, p = 0.0053), while high level of ITGA2B expression was correlated with poor prognosis for OS (log-rank test, p < 0.0001). Plus, low ITGA8 expression was associated with poor prognosis for disease-free survival (DFS) in the TCGA cohort (log-rank test, p < 0.0001). In the gene cluster network analysis, GIT1 and SHC1 associated with ITGA2B and ITGA8 were identified as independent predictive factors of overall survival of ccRCC. ITGA2B, ITGA8, GIT1, and SHC1 were identified as independent prognostic factors of overall survival of ccRCC. This method may act as a tool to reveal more prognostic-associated genes in ccRCC.

Upregulation of long non-coding RNA MALAT1 correlates with tumor progression and poor prognosis in clear cell renal cell carcinoma.

Long noncoding RNAs (lncRNAs) have been investigated as a new class of regulators of cellular processes, such as cell growth, apoptosis, and carcinogenesis. LncRNA metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) has recently been identified to be involved in tumorigenesis of several cancers such as lung cancer, pancreatic cancer, and cervical cancer. However, the role of lncRNA MALAT1 in clear cell renal cell carcinoma (ccRCC) remains unclear. Expression levels of lncRNA MALAT1 in ccRCC tissues and renal cancer cell lines were evaluated by quantitative real-time PCR (qRT-PCR), and its association with overall survival of patients was analyzed by statistical analysis. Small interfering RNA (siRNA) was used to suppress MALAT1 expression in renal cancer cells. In vitro assays were conducted to further explore its role in tumor progression. The expression level of MALAT1 was higher in ccRCC tissues and renal cancer cells compared to adjacent non-tumor tissues and normal human proximal tubule epithelial cells HK-2. The ccRCC patients with higher MALAT1 expression had an advanced clinical features and a shorter overall survival time than those with lower MALAT1 expression. And multivariate analysis showed that the status of MALAT1 expression was an independent predictor of overall survival in ccRCC. Additionally, our data indicated that knockdown expression of MALAT1 decreased renal cancer cell proliferation, migration, and invasion. Our data suggested that lncRNA MALAT1 was a novel molecule involved in ccRCC progression, which provided a potential prognostic biomarker and therapeutic target.

Src pathway activation in RCC and the correlation with grade and survival and the development of a rational new target in RCC.

453 Background: Loss of the VHL tumor suppressor in clear cell renal cell carcinoma (ccRCC) leads to the accumulation of Hypoxia Inducible Factor (HIF) and aberrant transcription of downstream targets such as VEGF. Inhibition of VEGF is the underlying therapeutic principle of several approved targeted agents; however, other HIF transcriptional targets remain to be explored. Increased expression of the HIF transcriptional target CUB domain-containing protein 1 (CDCP1) is associated with poor survival in ccRCC. CDCP1 is a substrate and binding partner for Src-family tyrosine kinases (SFKs) and dictates substrate specificity. Crosstalk between the SFK signaling cascade and the HIF pathway may result in feed forward signal amplification. Upregulation of Src signaling increases HIF-dependent transcription by both VHL-dependent and -independent mechanisms. Reciprocally, HIF accumulation can activate Src through PDGFRα, focal adhesion kinase, and CDCP1. Methods: Data from The Cancer Genome Atlas (TCGA) was used to assess mRNA expression and protein phosphorylation as well as clinical parameters and outcome in patients with ccRCC. Correlations were analyzed via Mann-Whitney U-test. Survival data was obtained using the TCGA portal, and analyzed by logrank test. To investigate the therapeutic potential of Src inhibition, the IC50 of Dasatinib in primary tumors derived cell lines were determined by an AlamarBlue Assay. Results: Increased Src and CDCP1 mRNA expression and decreased phosphorylation on the Src inhibitory site Y527 correlated with increased Fuhrman grade. Decreased phosphorylation on the inhibitory site Y527 correlated with worse median overall survival (78.4 vs. 19.7 months, p&lt; 0.00001). Similarly, increased Src and CDCP1 mRNA expression correlated with worse overall survival (78.4 vs. 26.9 months, p &lt; 0.00001, 78.4 vs. 37.2 months, p = 0.005). In vitro inhibition of Src with Dasatinib induces growth inhibition of primary cell lines with sarcomatoid or rhabdoid features, histomorphologic criteria associated with higher grade disease and worse prognosis. Conclusions: The Src pathway might be biologically relevant for ccRCC, and Src inhibition may have therapeutic potential.