Improving detection of carcinoma in situ in bladder cancer: urinary cytology vs the Xpert® BC Monitor.

OBJECTIVE: To investigate and compare the performance of urinary cytology and the Xpert BC Monitor test in the detection of bladder cancer in various clinically significant patient cohorts, including patients with carcinoma in situ (CIS), in a prospective multicentre setting, aiming to identify potential applications in clinical practice. PATIENTS AND METHODS: A total of 756 patients scheduled for transurethral resection of bladder tumour (TURBT) were prospectively screened between July 2018 and December 2020 at six German University Centres. Central urinary cytology and Xpert BC Monitor tests were performed prior to TURBT. The diagnostic performance of urinary cytology and the Xpert BC Monitor was evaluated according to sensitivity (SN), specificity (SC), negative predictive value (NPV) and positive predictive value (PPV). Statistical comparison of urinary cytology and the Xpert BC Monitor was conducted using the McNemar test. RESULTS: Of 756 screened patients, 733 (568 male [78%]; median [interquartile range] age 72 [62-79] years) were included. Bladder cancer was present in 482 patients (65.8%) with 258 (53.5%) high-grade tumours. Overall SN, SC, NPV and PPV were 39%, 93%, 44% and 92% for urinary cytology, and 75%, 69%, 59% and 82% for the Xpert BC Monitor. In patients with CIS (concomitant or solitary), SN, SC, NPV and PPV were 59%, 93%, 87% and 50% for urinary cytology, and 90%, 69%, 95% and 50% for the Xpert BC Monitor. The Xpert BC Monitor missed four tumours (NPV = 98%) in patients with solitary CIS, while potentially avoiding 63.3% of TURBTs in inconclusive or negative cystoscopy and a negative Xpert result. CONCLUSION: Positive urinary cytology may indicate bladder cancer and should be taken seriously. The Xpert BC Monitor may represent a useful diagnostic tool for correctly identifying patients with solitary CIS and unsuspicious or inconclusive cystoscopy.

Preoperative Function Assessment of Ex Vivo Kidneys with Supervised Machine Learning Based on Blood and Urine Markers Measured during Normothermic Machine Perfusion.

Establishing an objective quality assessment of an organ prior to transplantation can help prevent unnecessary discard of the organ and reduce the probability of functional failure. In this regard, normothermic machine perfusion (NMP) offers new possibilities for organ evaluation. However, to date, few studies have addressed the identification of markers and analytical tools to determine graft quality. In this study, function and injury markers were measured in blood and urine during NMP of 26 porcine kidneys and correlated with ex vivo inulin clearance behavior. Significant differentiation of kidneys according to their function could be achieved by oxygen consumption, oxygen delivery, renal blood flow, arterial pressure, intrarenal resistance, kidney temperature, relative urea concentration, and urine production. In addition, classifications were accomplished with supervised learning methods and histological analysis to predict renal function ex vivo. Classificators (support vector machines, k-nearest-neighbor, logistic regression and naive bayes) based on relevant markers in urine and blood achieved 75% and 83% accuracy in the validation and test set, respectively. A correlation between histological damage and function could not be detected. The measurement of blood and urine markers provides information of preoperative renal quality, which can used in future to establish an objective quality assessment.

Urinary Markers for Bladder Cancer Diagnosis and Monitoring.

Hematuria is a typical symptom of bladder cancer which enables early detection of bladder cancer. However, reliable diagnostic tools for bladder cancer using urine samples or other non-invasive methods are lacking. Tremendous attempts have been tried and revealed fancy works to convey definitive diagnostic power using urine samples. In this paper, we reviewed urinary markers for bladder cancer and compared their efficacies.

[Rational follow-up of non-muscle invasive bladder cancer]. / Rationale Nachsorge des nicht-muskelinvasiven Harnblasenkarzinoms.

BACKGROUND: Follow-up for non-muscle invasive bladder cancer (NMIBC) is a challenge for urologists that has not been finally resolved. The intensity of follow-up is based on the recurrence and progression behavior of the tumor as well as the patient's individual situation. MATERIALS AND METHODS: The following article focuses on the current data situation, the valid German S3 guideline and the available instruments for the detection of relapses and progression, taking into account tumor stages and degree of malignancy. RESULTS: Urethrocystoscopy, imaging and urine cytology are generally recommended, but the recommendations appear to be too extensive in the case of so-called intermediate risk profiles. Depending on the situation, urine markers could optimize follow-up, although results from prospective randomized studies are still pending. CONCLUSIONS: The current follow-up of NMIBC is invasive, carries the risk of side effects and increases costs. In the absence of scientific evidence, recommendations for follow-up for NMIBC are naturally based on expert opinion. In the opinion of the authors, overdiagnosis is currently taking place particularly in patients with an intermediate risk profile. The first prospective, marker-based studies are ongoing and will be helpful in the near future to improve the data situation relevant to urological practice.

Detection of Novel Urine Markers Using Immune Complexome Analysis in Bladder Cancer Patients: A Preliminary Study.

BACKGROUND/AIM: Little is known on urine biomarkers that are associated with malignant behavior in patients with bladder cancer (BC). Our aim was to identify BC-related factors in urine samples using our original method "immune complexome analysis", based on detecting the immune complex (IC). PATIENTS AND METHODS: Immune complexome analysis was performed using urine samples from 97 BC patients, including 67 with non-muscle invasive BC (NMIBC). RESULTS: Eight IC-antigens were recognized as candidates for BC-related factors from 20,165 proteins. IC-serum albumin, -fibrinogen γ chain, -hemoglobin subunit α, -hemoglobin subunit ß, -ceruloplasmin, and fibrinogen ß chain were significantly associated with either pathological features and/or outcome. IC-ceruloplasmin was most widely associated with pathological features in all BC patients and lamina propria invasion and urinary tract recurrence in NMIBC. CONCLUSION: Based on detection of IC-antigens it was demonstrated that six IC-antigens, especially IC-ceruloplasmin, are potential urine biomarkers in BC.

New synthetic cannabinoids carrying a cyclobutyl methyl side chain: Human Phase I metabolism and data on human cannabinoid receptor 1 binding and activation of Cumyl-CBMICA and Cumyl-CBMINACA.

Synthetic cannabinoids (SCs) represent a large group of new psychoactive substances (NPS), sustaining a high prevalence on the drug market since their first detection in 2008. Cumyl-CBMICA and Cumyl-CBMINACA, the first representatives of a new subclass of SCs characterized by a cyclobutyl methyl (CBM) moiety, were identified in July 2019 and February 2020. This work aimed at evaluating basic pharmacological characteristics and human Phase I metabolism of these compounds. Human Phase I metabolites were tentatively identified by liquid chromatography-quadrupole time-of-flight mass spectrometry (LC-QToF-MS) of urine samples and confirmed by a pooled human liver microsome (pHLM) assay. The basic pharmacological evaluation was performed by applying a competitive ligand binding assay and a functional activation assay (GTPγS) using cell membranes carrying the human cannabinoid receptor 1 (hCB1 ). Investigation of the human Phase I metabolism resulted in the identification of specific urinary markers built by monohydroxylation or dihydroxylation. Although Cumyl-CBMICA was primarily hydroxylated at the indole ring, hydroxylation of Cumyl-CBMINACA mainly occurred at the CBM moiety. Both substances acted as agonists at the hCB1 receptor, although substantial differences could be observed. Cumyl-CBMINACA showed higher binding affinity (Ki = 1.32 vs. 29.3 nM), potency (EC50 = 55.4 vs. 497 nM), and efficacy (Emax = 207% vs. 168%) than its indole counterpart Cumyl-CBMICA. This study confirms that substitution of an indole by an indazole core tends to increase in vitro potency, which is potentially reflected by higher in vivo potency. The emergence and disappearance of SCs distributed via online shops carrying a CBM moiety once more demonstrate the "cat-and-mouse" game between manufacturers and legislation.

Impact of Histopathological Prostate Inflammation on Urine-Based Prostate Cancer Prediction Using the Prostate Cancer Gene 3 Score.

INTRODUCTION: The Prostate Cancer gene 3 (PCA3) urine test has gained importance in the diagnostic workup of prostate cancer (PC). Limited evidence suggests that PCA3 is not altered in the presence of inflammation. OBJECTIVE: To assess the impact of histological inflammation on PCA3. METHODS: PCA3 was evaluated in patients prior to prostate biopsy (n = 193) and to radical prostatectomy (n = 197). In patients without PC, inflammation was assessed and quantified by individual scores integrating grade and extent. Uni- and multivariate analyses were performed to assess the impact of inflammation grade on PCA3. RESULTS: The PCA3 scores prior to prostatectomy were lower (median 45) than those before positive biopsy (57; p = 0.008). Of 101 negative biopsies, 78% showed inflammation. The median PCA3 scores in the groups with no inflammation and with maximum grade 1 (n = 22), 2 (n = 38), and 3 (n = 19) inflammation were 45, 38, 27, and 25 (p = 0.016). The multivariate models revealed a decrease in PCA3 proportional to the grade and extent of inflammation (p < 0.04 each). CONCLUSIONS: The present data imply that the PCA3 score decreases in the presence of inflammation, which is relevant, for instance, to testing after a recently performed biopsy. In general, inflammation should be regarded as a factor putatively influencing PCA3 and other available and upcoming PC tests.

A novel diagnostic tool for the detection of bladder cancer: Measurement of urinary high mobility group box-1.

OBJECTIVE: We aimed to investigate the diagnostic value of urinary High Mobility Group Box-1 (HMGB1) level as a noninvasive tool that can be potentially used for diagnosis and during follow-up in patients with bladder cancer patients. METHOD: The study was conducted in a total of 121 participants including 61 patients diagnosed with primary bladder cancer, 30 patients with an acute urinary tract infection and 30 healthy controls. Age, gender and urinary HMGB1 levels of the study groups were evaluated. The association of clinical features (tumor diameter, number of foci, pathological grade, muscle invasion) with urinary HMGB1 levels was investigated in patients with bladder cancer. RESULTS: All 3 groups showed a normal age and gender distribution with no significant difference among them (P = 0.775 and P = 0.967, respectively). A significant difference was detected in urinary HMGB1 levels among the 3 groups (P < 0.001). When urinary HMGB1 levels were compared between patients with high grade vs. low grade tumors, the mean HMGB1 level was 44.39 pg/ml (12.1-505.2) in patients with low grade tumors and 280 pg/ml (18.7-2685.3) in patients with high grade tumors (P < 0.001). Patients with a greater number of tumor foci had higher HMGB1 levels in comparison to patients with a single tumor focus (P = 0.008). Urinary HMGB1 levels were higher in patients with a tumor diameter of ≥3 cm than in patients with a tumor diameter less than 3 cm (P = 0.001). Patients with muscle-invasive bladder cancer exhibited higher urinary HMGB1 levels compared to patients with non-muscle-invasive bladder cancer (P = 0.033). The cut-off values derived from the ROC analysis were 63.30 pg/ml for distinguishing bladder cancer from urinary tract infection, 30.94 pg/ml for urinary tract infection versus control group and 38.70 pg/ml for bladder cancer vs. control group, respectively. Sensitivity was 59% and specificity was found 77%. CONCLUSION: In future controlled studies involving larger patient groups, urinary HMGB1 levels can be used for diagnostic and screening purposes in bladder cancer patients.

Impact of variant microscopic interpretation of the uCyt+ immunocytological urine test for the detection of bladder cancer.

BACKGROUND: Urinary marker tests for bladder cancer (BC) detection and surveillance represent a desirable approach to diagnosis and follow-up. The SCIMEDX uCyt+ assay detects antigens expressed by BC cells (mucin glycoprotein and carcinoembryonic antigen) using green and red fluorescence and is interpreted according to specific manufacturer's recommendations. In the present study, we evaluated divergent approaches of numeric and morphological analysis of uCyt+ to generate a rationale for alternative test interpretation strategies. METHODS: A total of 444 patients with hematuria and without history of BC underwent uCyt+ analysis, cystoscopy and histological examination of tissue biopsies. Beside positive cells according to the manufacturer's definition (definitely positive cells, DPC), (i) cells showing borderline character (borderline cells, BLC), and (ii) cells with staining present below defined border (subliminal cells, SLC) were included into the analytical algorithm. Different cut-off levels for cell counts (>0, ≥3, ≥5) were evaluated separately with regard to their diagnostic accuracy. Moreover, the influence of clinical factors on test results were evaluated. RESULTS: Adding BLC at a cut-off of ≥3 cells resulted in Area Under the Curve (AUC) of 0.70 for green and 0.77 for red fluorescence, respectively. Adding SLC led to reduced AUC (0.62 and 0.73, respectively). Male gender was significantly associated with false positive results in the "best AUC" groups (P = .0101). No further correlations to clinical influencing factors were observed. CONCLUSIONS: Adding microscopic BLC as test-positive and adjusting cut-off level for the interpretation of uCyt+ may improve assay performance independent of clinical factors.

Rapid Assessment of Microbiota Changes in Individuals with Autism Spectrum Disorder Using Bacteria-derived Membrane Vesicles in Urine.

Individuals with autism spectrum disorder (ASD) have altered gut microbiota, which appears to regulate ASD symptoms via gut microbiota-brain interactions. Rapid assessment of gut microbiota profiles in ASD individuals in varying physiological contexts is important to understanding the role of the microbiota in regulating ASD symptoms. Microbiomes secrete extracellular membrane vesicles (EVs) to communicate with host cells and secreted EVs are widely distributed throughout the body including the blood and urine. In the present study, we investigated whether bacteria-derived EVs in urine are useful for the metagenome analysis of microbiota in ASD individuals. To address this, bacterial DNA was isolated from bacteria-derived EVs in the urine of ASD individuals. Subsequent metagenome analysis indicated markedly altered microbiota profiles at the levels of the phylum, class, order, family, and genus in ASD individuals relative to control subjects. Microbiota identified from urine EVs included gut microbiota reported in previous studies and their up- and down-regulation in ASD individuals were partially consistent with microbiota profiles previously assessed from ASD fecal samples. However, overall microbiota profiles identified in the present study represented a distinctive microbiota landscape for ASD. Particularly, the occupancy of g_Pseudomonas, g_Sphingomonas, g_Agrobacterium, g_Achromobacter, and g_Roseateles decreased in ASD, whereas g_Streptococcus, g_Akkermansia, g_Rhodococcus, and g_Halomonas increased. These results demonstrate distinctively altered gut microbiota profiles in ASD, and validate the utilization of urine EVs for the rapid assessment of microbiota in ASD.

Comparison of different concepts for interpretation of chromosomal aberrations in urothelial cells detected by fluorescence in situ hybridization.

PURPOSE: Urine fluorescence in situ hybridization (FISH) has become a broadly used marker for noninvasive detection of bladder cancer (BC). However, it has been discussed whether the interpretation algorithm proposed by the manufacturer could be improved. Aim of the present study was to compare alternative evaluation strategies of FISH for detection of BC. METHODS: We included 1048 patients suspicious for BC, who underwent urine FISH examination before cystoscopy (diagnostic cohort). Herefrom, we selected 122 patients (prognostic cohort) with a history of non-muscle-invasive BC who were cystoscopically tumor free and received FISH analysis ahead of a follow-up period of 24 months. FISH results were interpreted by the algorithms of UroVysion™, Bubendorf et al. and Zellweger et al. RESULTS: In the diagnostic cohort, 228 patients (21.8%) had BC at time of evaluation; in the prognostic cohort 39 patients (32.0%) experienced tumor recurrence. Alterations in chromosome 3, 7 and 17 correlated with the presence of BC. Relative loss of 9p21 was associated with BC and higher risk for progression. The evaluation strategy proposed by Zellweger et al. showed highest accuracy of all FISH assessments. Performance of evaluation strategies differed in voided urine samples and samples obtained after mechanical manipulation. CONCLUSIONS: The performance of FISH in BC diagnosis strongly depends on the interpretation criteria. Alternative evaluation methods partly show superior diagnostic performance compared to the manufacturer's algorithm. The introduction of specific cutoffs for tetraploid cells improves specificity. Further modifications of the interpretation algorithm of the Urovysion® FISH assay have the potential to positively affect the value of this test in diagnosis and surveillance of BC.

Urine TMPRSS2: ERG Fusion Transcript as a Biomarker for Prostate Cancer: Literature Review.

Prostate cancer (PCa) is one of the most common male malignancies. Serum prostate-specific antigen (PSA) is one of the most valuable biomarkers in tumor biology and remains the standard marker in detecting and monitoring PCa. However, the high number of serum PSA false positive and false negative results make the identification of novel biomarkers extremely welcome to improve our diagnostic accuracy in detecting PCa and distinguishing the aggressive from the indolent ones. In this study, we analyzed the current role of urinary gene fusion transcripts involving v-ets erythroblastosis virus E26 oncogene homolog, commonly known as ERG, and the androgen-regulated gene transmembrane protease, serine 2 (TMPRSS2), as a biomarker for PCa. Used as a single marker, urinary TMPRSS2:ERG has low sensitivity but high specificity. However, its combination with the other urinary marker PCa antigen 3 (PCA3) has been reported to provide high specificity and sensitivity. Finally, a commercially available assay combining serum PSA with urinary PCA3 and TMPRSS2:ERG provides a 90% specificity and 80% sensitivity in diagnosing PCa. Urinary TMPRSS2:ERG also seems to be indicative of PCa aggressiveness upon biopsy. Should these findings be confirmed in larger studies, urinary TMPRSS2:ERG might become a valuable test not only for diagnosing PCa but also for distinguishing the aggressive tumors from the indolent ones.

Interaction network analysis of differentially expressed genes and screening of cancer marker in the urine of patients with invasive bladder cancer.

OBJECTIVE: To detect the expression profile of bladder cancer and to delineate the interaction network of these genes in invasive bladder cancer. METHODS: A total of 126 differentially expressed genes were identified, and input into STRING online database to delineate interaction network. The network data were screened with central nodes. The expression of genes with the most evident change in the exfoliated cells of urine was detected. RNA markers with over-expression in stage Ta tumor and/or T1 to T4 tumors but low expression in blood or inflammatory cells were characterized. RESULTS: On the basis of assay of 21,639 whole-genome oligonucleotide microarray, a total of 126 differentially expressed genes were identified, of which 69 had up-regulated expression and 57 had down-regulated expression. STRING screening showed there were interactions among 103 genes in the bladder cancer which formed a complex network. A total of 23 central nodes were screened with Cytoscape and are involved in multiple signaling pathways related to tumorigenesis. The test specificity was 80% for the 30 control patients with urinary tract infections. The combination of BLCA-4 and HOXA13 could distinguish between low and high grade tumors, with specificity and sensitivity of 80%. CONCLUSION: The interaction network of differentially expressed genes, especially the central nodes of this network, can provide evidence for the early diagnosis and molecular targeted therapy of invasive bladder cancer, and combined detection of IGF-1, hTERT, BLCA-4 and HOXA13 genes is helpful to evaluate BTCC at different stages.