Biomarkers For Detection Of Bladder Cancer Research Articles

Properties of non-coding mutation hotspots as urinary biomarkers for bladder cancer detection

Mutations at specific hotspots in non-coding regions of ADGRG6, PLEKHS1, WDR74, TBC1D12 and LEPROTL1 frequently occur in bladder cancer (BC). These mutations could function as biomarkers for the non-invasive detection of BC but this remains largely unexplored. Massively-parallel sequencing of non-coding hotspots was applied to 884 urine cell pellet DNAs: 591 from haematuria clinic patients (165 BCs, 426 non-BCs) and 293 from non-muscle invasive BC surveillance patients (29 with recurrence). Urine samples from 142 non-BC haematuria clinic patients were used to optimise variant calling. Non-coding mutations are readily detectable in the urine of BC patients and undetectable, or present at much lower frequencies, in the absence of BC. The mutations can be used to detect incident BC with 66% sensitivity (95% CI 58–75) at 92% specificity (95% CI 88–95) and recurrent disease with 55% sensitivity (95% CI 36–74) at 85% specificity (95% CI 80–89%) using a 2% variant allele frequency threshold. In the NMIBC surveillance setting, the detection of non-coding mutations in urine in the absence of clinically detectable disease was associated with an increased relative risk of future recurrence (RR = 4.62 (95% CI 3.75–5.48)). As urinary biomarkers, non-coding hotspot mutations behave similarly to driver mutations in BC-associated genes and could be included in biomarker panels for BC detection.

Discovery of Volatile Biomarkers for Bladder Cancer Detection and Staging through Urine Metabolomics.

Timely diagnosis is crucial to improve the long-term survival of bladder cancer (BC) patients. The discovery of new BC biomarkers based in urine analysis is very attractive because this biofluid is in direct contact with the inner bladder layer, in which most of the neoplasms develop, and is non-invasively collected. Hence, this work aimed to unveil alterations in the urinary volatile profile of patients diagnosed with BC compared with cancer-free individuals, as well as differences among patients diagnosed at different tumor stages, to identify candidate biomarkers for non-invasive BC diagnosis and staging. Urine analysis was performed by headspace solid-phase microextraction coupled with gas chromatography-mass spectrometry (HS-SPME-GC-MS). The results unveiled that BC patients have a distinct urinary volatile profile characterized by higher levels of several alkanes and aromatic compounds, and lower levels of aldehydes, ketones and monoterpenes. Seventeen significantly altered volatiles were used to evaluate the performance for overall BC detection, disclosing 70% sensitivity, 89% specificity and 80% accuracy. Moreover, distinct urinary volatile profiles were found among patients diagnosed at different tumor stages (Ta/Tis, T1 and ≥T2). This work identified distinct urinary volatile signatures of BC patients with potential for non-invasive detection and staging of bladder cancer.

Urinary MicroRNA96 as Biomarkers for Bladder Cancer Detection

Bladder malignancy is viewed as the fifth most basic disease with expanded disorder and demise. As of late, incessant examinations showed that microRNAs are arising as indicative biomarkers for bladder malignancy. Explicit miRNA profiles have been distinguished for a few examples from patients with bladder malignancy. MicroRNAs are noncoding RNA particles of around 23 nucleotides that assume significant parts in different strides during the movement of bladder malignant growth. Here, we audit the declaration of miRNAs and their organic capacities, guideline, and clinical ramifications in bladder malignant growth. Either downregulation or upregulation of miRNAs happens in bladder malignant growth through epigenetic changes or imperfections of the biogenesis mechanical assembly. Liberation of miRNAs is engaged with cell cycle capture, apoptosis, multiplication, metastasis, drug obstruction, and different capacities in bladder malignancy. Various miRNAs, have been related with tumor type, stage, or patient endurance, and miRNAs may be created as demonstrative or prognostic markers. A superior comprehension of the jobs of miRNAs in bladder malignant growth will reveal insight into the sub-atomic systems of bladder disease.

A Panel of Urinary Long Non-coding RNAs Differentiate Bladder Cancer from Urocystitis.

Liquid biopsy is becoming a promising method for non-invasive cancer detection. In several proof-of-concept studies, long non-coding RNAs (lncRNAs) were found to be potential biomarkers for bladder cancer detection. The objective of this study was to discover a panel of cell-free, urinary lncRNAs as liquid biopsy biomarkers to non-invasively differentiate bladder cancer from chronic urocystitis. To this end, we collected urine samples from both bladder cancer patients and urocystitis patients. These samples were divided into discovery group and validation group. In the discovery group, the expression levels of 16 cell-free urinary lncRNAs were measured by qPCR to discover candidate biomarkers. The diagnostic performance of the candidate lncRNA biomarkers was then evaluated, which led to a panel of lncRNA biomarkers for bladder cancer detection. The performance of this panel of biomarkers was further evaluated in the validation group to see if these lncRNA biomarkers could discriminate the bladder cancer patients from urocystitis patients. We found that all of the 16 lncRNAs evaluated in this study demonstrated significant difference (p<0.05) of expression between bladder cancer patients and urocystitis patients. Nine lncRNAs provided decent diagnostic performance with area under the receiver operating characteristic (ROC) curve (AUC) reaching 0.70 or higher. We then selected the top four lncRNAs, namely UCA1-201, HOTAIR, HYMA1 and MALAT1, to form a panel of urinary biomarkers. Using this panel, bladder cancer patients could be discriminated from urocystitis patients, with sensitivity and specificity reaching 95.7% and 94.3%, respectively. Finally, we confirmed the applicability of the four-lncRNA panel in an independent validation study that included 60 bladder cancer patients and 60 urocystitis patients. Our study paves the way for further studies aimed at large-scale clinical tests of developing lncRNA biomarkers in urine for bladder cancer diagnostics.

Diagnostic value of urinary survivin as a biomarker for bladder cancer: a systematic review and meta-analysis of published studies.

This study is a meta-analysis and aims to determine the value of urinary survivin for detecting bladder cancer (BC) on the basis of preceding statistical performance and to compare their diagnostic value. Considering that the urinary survivin data were from both RNA and protein levels, the key words "bladder cancer" AND "survivin" and "bladder cancer" AND "survivin RNA" were used; and PubMed, Web of Science, and Cochrane Library were systematically searched to identify relevant articles. The methodological quality of each study was assessed by QUADAS-2. Data were analyzed by STATA 12.0 and Meta-disc v.1.4 software package. A random-effects model was used and subgroup analysis was carried out to identify possible sources of heterogeneity. Nine articles for survivin protein test with 789 patients and 684 controls, and 12 articles for survivin RNA test with 880 patients and 922 controls were identified. The results showed that the pooled sensitivity was 0.79 (95% CI 0.73, 0.84), specificity was 0.87 (95% CI 0.79, 0.92) of the survivin protein test for bladder cancer, and the sensitivity and specificity was 0.84 (95% CI 0.79, 0.88) and 0.94 (95% CI 0.89, 0.97) of the survivin RNA test. The AUC of the two approaches was 0.89 (95% CI 0.86, 0.91) and 0.94 (95% CI 0.92, 0.96), respectively. The survivin protein and survivin RNA both had great potential as biomarkers for BC detection, and survivin RNA showed higher accuracy than survivin protein on BC diagnosis.

MicroRNAs as Biomarkers for the Diagnostics of Bladder Cancer: a Meta-Analysis.

Bladder cancer (BCa) is the fifth most common cancer with significant morbidity and mortality. Recently, numerous studies demonstrated that microRNAs (miRNAs) are emerging as diagnostic biomarkers for BCa. However, the findings in these studies are inconsistent. To systematically assess the potential diagnostic value of miRNAs for BCa, a meta-analysis was performed in this study. Relevant literature was researched in PubMed, Embase, Cochrane Library, Chinese National Knowledge Infrastructure (CNKI), and WanFang databases up to September 1, 2014. The pooled sensitivity, specificity, positive likelihood ratio (PLR), negative LR (NLR), diagnostic odds ratio (DOR), and area under the SROC curve (AUC) value were analyzed by the random-effects model, whose parameters reflected the overall diagnostic performance of miRNAs. Thirty studies from 10 individual publications, including 1019 BCa patients and 690 controls, were included in this meta-analysis. The pooled sensitivity, specificity, PLR, NLR, DOR, and AUC were 0.80 (95% CI: 0.78 - 0.81), 0.74 (95% CI: 0.72 - 0.76), 3.22 (95% CI: 2.68 - 3.87), 0.26 (95% CI: 0.21 - 0.32), 15.20 (95% CI: 10.25 - 22.53) and 0.85, respectively, indicating a moderate diagnostic accuracy for BCa. Moreover, our subgroup analyses showed that analysis of multiple miRNAs (AUC, sensitivity, and specificity of 0.913, 0.86, and 0.80, respectively) yielded a higher diagnostic accuracy than of single miRNAs (AUC, sensitivity, and specificity of 0.84, 0.78, and 0.73, respectively) in BCa diagnosis. In addition, as biomarkers, miRNAs are more suitable for the diagnosis of non-muscle-invasive BCa (NMIBCa) with AUC of 0.84, sensitivity of 0.74, and specificity of 0.77 than muscle-invasive BCa (MIBCa) with AUC of 0.79, sensitivity of 0.73, and specificity of 0.73. The present meta-analysis suggests that miRNAs are potential novel biomarkers for detection of BCa. However, further validation studies are still needed to confirm our findings.

Investigation of CCL18 and A1AT as potential urinary biomarkers for bladder cancer detection.

BackgroundIn this study, we further investigated the association of two biomarkers, CCL18 and A1AT, with bladder cancer (BCa) and evaluated the influence of potentially confounding factors in an experimental model.MethodsIn a cohort of 308 subjects (102 with BCa), urinary concentrations of CCL18 and A1AT were assessed by enzyme-linked immunosorbent assay (ELISA). In an experimental model, benign or cancerous cells, in addition to blood, were added to urines from healthy controls and analyzed by ELISA. Lastly, immunohistochemical staining for CCL18 and A1AT in human bladder tumors was performed.ResultsMedian urinary protein concentrations of CCL18 (52.84 pg/ml vs. 11.13 pg/ml, p < 0.0001) and A1AT (606.4 ng/ml vs. 120.0 ng/ml, p < 0.0001) were significantly elevated in BCa subjects compared to controls. Furthermore, the addition of whole blood to pooled normal urine resulted in a significant increase in both CCL18 and A1AT. IHC staining of bladder tumors revealed CCL18 immunoreactivity in inflammatory cells only, and there was no significant increase in these immunoreactive cells within benign and cancerous tissue and no association with BCa grade nor stage was noted. A1AT immunoreactivity was observed in the cytoplasm of epithelia cells and intensity of immunostaining increased with tumor grade, but not tumor stage.ConclusionsFurther development of A1AT as a diagnostic biomarker for BCa is warranted.

A multi-analyte assay for the noninvasive detection of bladder cancer.

306 Background: Accurate urinary assays for bladder cancer (BCa) detection would benefit both patients and healthcare systems. Through genomic and proteomic profiling of urine components, we have previously identified a panel of biomarkers that can outperform current urine-based biomarkers for the non-invasive detection of BCa. Herein, we report the diagnostic utility of various multivariate combinations of these biomarkers. Methods: We performed a case-controlled validation study in which voided urines from 550 patients (220 tumor bearing subjects) were analyzed. The urinary concentrations of 14 biomarkers (IL-8, MMP-9,MMP-10, SDC1,CCL18, PAI-1, CD44, VEGF, ANG, CA9,A1AT, OPN, PTX3, and APOE) were assessed by enzyme-linked immunosorbent assay (ELISA). Diagnostic performance of each biomarker and multivariate models were compared using receiver operating characteristic curves and the chi-square test. Results: An 3-biomarker model (CA9, PAI-1 and IL-8) achieved the most accurate BCa diagnosis (sensitivity 90%, specificity 88%). These data show that a multivariate urine-based assay can markedly improve the accuracy of non-invasive BCa detection. Conclusions: Further validation studies are under way to investigate the clinical utility of this panel of biomarkers for BCa diagnosis and disease monitoring.

Upregulation of Tissue and Urinary Nicotinamide N-Methyltransferase in Bladder Cancer: Potential for the Development of a Urine-Based Diagnostic Test

Carcinoma of the bladder is one of the most common urologic malignancies occurring worldwide. Diagnosis and monitoring of bladder urothelial carcinoma (UC) are based on cystoscopy and urinary cytology. However, these diagnostic methods still have some limitations, mainly related to invasive nature and lack of sensitivity. New reliable and non-invasive biomarkers for bladder cancer detection are therefore required. To explore the involvement of enzymes of drug metabolism in bladder cancer, in the present study, we analyzed the gene expression profiles of tumor and normal looking tissues obtained from the same patient by cDNA macroarray. The enzyme nicotinamide N-methyltransferase (NNMT) was identified as a highly expressed gene in bladder cancer. RT-PCR, Real-Time PCR, Western blot analysis, and catalytic activity assay, performed on a large cohort of patients with bladder UC, confirmed NNMT upregulation. NNMT mRNA and protein levels were also determined in urine specimens obtained from patients with bladder UC and healthy subjects. We found that NNMT expression levels were significantly higher in patients with bladder tumor compared to controls that showed very low or undetectable amounts of NNMT transcript and protein. Our results indicate that a marked NNMT increase is a peculiar feature of bladder UC and suggest the potential suitability of urine NNMT expression levels determination for early and non-invasive diagnosis of bladder cancer.

Urinary levels of Hepatocarcinoma-intestine-pancreas/Pancreatitis-associated protein as a diagnostic biomarker in patients with bladder cancer

BackgroundTo assess the possibility of hepatocarcinoma-intestine-pancreas/pancreatitis-associated protein (HIP/PAP) as a biological marker for detecting Bladder cancer (BCa), we examined the expression of HIP/PAP in both BCa specimens and BCa cell lines and measured HIP/PAP levels in urine from patients with BCa.MethodsHIP/PAP expression in BCa samples was evaluated by western blot analysis, and urinary levels of HIP/PAP in patients with BCa were measured by enzyme-linked immunosorbent assay. Urine samples were collected from 10 healthy volunteers and 109 with benign urological disorders as controls, and from 101 patients who were diagnosed with BCa.ResultsHIP/PAP was highly expressed in BCa samples as compared with control bladder. Urinary HIP/PAP concentrations were significantly higher in BCa patients than in controls (median value; 3.184 pg/mL vs. 55.200 pg/mL, P <0.0001, by Mann–Whitney U test). Urinary HIP/PAP levels in BCa patients correlated positively with pathological T stages and progression-risk groups among non-muscle invasive BCa (P = 0.0008, by Kruskal-Wallis test). Regarding the recurrence-risk classifications of non-muscle invasive BCa, the urinary levels of HIP/PAP were significantly higher in the intermediate than in the low risk group (P = 0.0002, by Mann–Whitney U test). Based on a cut-off of 8.5 pg/mL, the ability of urinary HIP/PAP levels to detect BCa had a sensitivity of 80.2%, specificity of 78.2%, positive predictive value (PPV) of 75.7%, and negative predictive value (NPV) of 82.3%.ConclusionsHIP/PAP was abundantly expressed in BCa, and the urinary levels of HIP/PAP could be a novel and potent biomarker for detection of BCa, and also for predicting the risks of recurrence- and progression-risk of non-muscle invasive BCa. A large scale study will be needed to establish the usefulness of this biomarker.

Bladder Cancer Diagnosis and Identification of Clinically Significant Disease by Combined Urinary Detection of Mcm5 and Nuclear Matrix Protein 22

BackgroundUrinary biomarkers for bladder cancer detection are constrained by inadequate sensitivity or specificity. Here we evaluate the diagnostic accuracy of Mcm5, a novel cell cycle biomarker of aberrant growth, alone and in combination with NMP22.Methods1677 consecutive patients under investigation for urinary tract malignancy were recruited to a prospective blinded observational study. All patients underwent ultrasound, intravenous urography, cystoscopy, urine culture and cytologic analysis. An immunofluorometric assay was used to measure Mcm5 levels in urine cell sediments. NMP22 urinary levels were determined with the FDA-approved NMP22® Test Kit.ResultsGenito-urinary tract cancers were identified in 210/1564 (13%) patients with an Mcm5 result and in 195/1396 (14%) patients with an NMP22 result. At the assay cut-point where sensitivity and specificity were equal, the Mcm5 test detected primary and recurrent bladder cancers with 69% sensitivity (95% confidence interval = 62–75%) and 93% negative predictive value (95% CI = 92–95%). The area under the receiver operating characteristic curve for Mcm5 was 0.75 (95% CI = 0.71–0.79) and 0.72 (95% CI = 0.67–0.77) for NMP22. Importantly, Mcm5 combined with NMP22 identified 95% (79/83; 95% CI = 88–99%) of potentially life threatening diagnoses (i.e. grade 3 or carcinoma in situ or stage ≥pT1) with high specificity (72%, 95% CI = 69–74%).ConclusionsThe Mcm5 immunoassay is a non-invasive test for identifying patients with urothelial cancers with similar accuracy to the FDA-approved NMP22 ELISA Test Kit. The combination of Mcm5 plus NMP22 improves the detection of UCC and identifies 95% of clinically significant disease. Trials of a commercially developed Mcm5 assay suitable for an end-user laboratory alongside NMP22 are required to assess their potential clinical utility in improving diagnostic and surveillance care pathways.

Vascular Endothelial Growth Factor, Carbonic Anhydrase 9, and Angiogenin as Urinary Biomarkers for Bladder Cancer Detection

To investigate whether elevated urinary levels of vascular endothelial growth factor (VEGF), carbonic anhydrase 9 (CA9), and angiogenin are associated with bladder cancer (BCa). This was a case-control study in which voided urine samples from 127 patients (63 control subjects and 64 patients with BCa) were analyzed. The urinary concentrations of VEGF, CA9, angiogenin, and bladder tumor antigen (BTA) were assessed using enzyme-linked immunosorbent assays. We used the area under the curve of receiver operating characteristic curves to determine the ability of VEGF, CA9, and angiogenin to detect BCa in voided urine samples. Data were also compared with the findings from a commercial enzyme-linked immunosorbent assay-based BCa detection assay (BTA-Trak). The sensitivity, specificity, and positive and negative predictive values were calculated. The urinary concentrations of VEGF, CA9, angiogenin, and BTA were significantly elevated in those with BCa. VEGF was the most accurate urinary biomarker (area under the curve 0.886, 95% confidence interval 0.8301-0.9418). Furthermore, multivariate regression analysis highlighted VEGF (odds ratio 5.90, 95% confidence interval 2.60-13.40, P < .0001) as an independent variable. The sensitivity and specificity for VEGF (83% sensitivity and 87% specificity) outperformed those for BTA (80% sensitivity and 84% specificity). VEGF could be a valuable addition to voided urine sample analysis for the detection of BCa. Larger, prospective studies are needed to determine the clinical utility of urinary VEGF and angiogenin as biomarkers in the noninvasive evaluation of patients with BCa.

Abstract 4160: Glycoprotein biomarkers for bladder cancer detection identified using LC-MS/MS and label free quantification

Abstract PURPOSE: Cancers of the urinary bladder are the fifth most commonly diagnosed malignancy in the US. Early clinical diagnosis of bladder cancer remains a major challenge and the development of noninvasive methods for detection and surveillance is desirable for both patients and health care providers. METHODS: In order to identify urinary proteins with potential clinical utility we enriched and profiled the glycoprotein component of urine samples using a dual-lectin affinity chromatography and LC-MS/MS platform. RESULTS: From a primary sample set obtained from 56 cancer patients and 47 healthy individuals, a total of 265 distinct glycoproteins were identified with high confidence, and changes in glycoprotein abundance between groups were quantified by a label-free spectral counting method. Validation of candidate biomarker disease association was performed on second set of 70 independent samples using an antibody-lectin sandwich microarray and ELISA. Increased levels of urinary alpha-1-antitrypsin (A1AT) glycoprotein were identified and confirmed as being indicative of the presence of bladder cancer (p value &amp;lt; 0.0001). CONCLUSION: The described strategy can be used to discover biomarkers for disease status in other body fluids. Application of the strategy in this study has provided novel candidate biomarkers for the non-invasive molecular detection of bladder cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4160. doi:10.1158/1538-7445.AM2011-4160

Genotypic and phenotypic biomarker profiles for individual risk assessment and cancer detection (lessons from bladder cancer risk assessment in symptomatic patients and workers exposed to benzidine).

There is a need for improved methods for detecting individuals at risk for cancer to target subsets of patients for more intensive individual screening and targeted cancer therapy and chemoprevention. One approach for accomplishing this objective is to detect premalignant molecular fingerprints in an organ at risk for cancer or to define biomarkers reflective of treatment selection and response. Bladder cancer is an excellent model for testing this approach; however, comprehending the strategy for biomarker selection and analysis is more complicated than is generally appreciated. The objective of this article is to provide a succinct overview of our experience with the selection of biomarkers for bladder cancer detection, first in symptomatic patients and then in high-risk cohorts of workers at risk for bladder cancer. Biomarker selection depends on multiple parameters, each of which must be optimized to enhance the utility of a biomarker for clinical application. Many markers that initially show promise fail in the clinical arena for a variety of reasons. Important parameters include when a biomarker is expressed in carcinogenesis (i.e., early vs. late), the sample type, and the method of analysis. These all contribute to the sensitivity, specificity, and ultimate clinical utility of a biomarker. New technologies/ support the notion that all diseases start in the cell, and Seymore West indicated the cell, under appropriate conditions, can function as a microcuvette for biophysical cytochemical analysis. Spectroscopy provides an accurate and sensitive method for quantitative single-cell proteomics. Improved and more stable fluorescence probes will enhance the utility of cellular chemistry, as will a rationale approach for biomarker selection based on the concepts of field cancerization, complemented by improved quantitative analysis of protein markers at the single-cell level. Our laboratory has developed a platform for single-cell proteomic analysis that can be applied to multiple basic science and clinical problems. Single-cell proteomics also facilitates the study of genetic instability and epigenetic signaling (stromal-epithelial interactions) in relation to cancer therapy and diagnosis. Because most cancers arise through multiple signaling pathways and are heterogeneous, the identification of appropriate biomarker profiles provides a number of strategic advantages over a single biomarker. Complex networks of signaling pathways lead to increased cell proliferation, decreased cell adhesion, cellular differentiation, genetic instability, and other functions associated with the malignant phenotype. The purpose of this presentation is to illustrate the fundamental concepts for selection and profile analysis of high-level phenotypic biomarkers developed for bladder cancer risk assessment, screening, and early bladder cancer detection.