Real-world performance of Uromonitor® in urothelial bladder cancer detection: a multicentric trial.

OBJECTIVES: To compare Uromonitor® (U-Monitor Lda, Porto, Portugal), a multitarget DNA assay that detects mutated proto-oncogenes (telomerase reverse transcriptase [TERT], fibroblast growth factor receptor 3 [FGFR-3], Kirsten rat sarcoma viral oncogene homologue [KRAS]), with urine cytology in the urine-based diagnosis of urothelial carcinoma of the bladder (UCB) within a multicentre real-world setting. PATIENTS AND METHODS: This multicentre, prospective, double-blind study was conducted across four German urological centres from 2019 to 2024. We evaluated the diagnostic performance of Uromonitor compared to urine cytology in a cohort of patients with UCB and in healthy controls within a real-world setting. Sensitivity, specificity, positive-predictive value (PPV), negative-predictive value (NPV), and accuracy of the tests were measured, in addition to multivariate analyses to assess the ability of individual proto-oncogene mutations in detecting UCB. The biometric sample size was designed to achieve a 10% difference in sensitivity. RESULTS: Patients with UCB comprised 63.7% (339/532) of the study group. Uromonitor showed a sensitivity, specificity, PPV, NPV, accuracy, and an area-under-the-curve of 49.3%, 93.3%, 92.8%, 51.1%, 65.2%, and 0.713%, respectively. These metrics did not demonstrate statistical superiority over urine cytology in terms of sensitivity (44.6%; P = 0.316). Moreover, the comparison of additional test parameters, as well as the comparison within various sensitivity analyses, yielded no significant disparity between the two urinary tests. Multivariate logistic regression underscored the significant predictive value of a positive Uromonitor for detecting UCB (odds ratio [OR] 9.03; P < 0.001). Furthermore, mutations in TERT and FGFR-3 were independently associated with high odds of UCB detection (OR 13.30 and 7.04, respectively), while KRAS mutations did not exhibit predictive capability. CONCLUSION: Despite its innovative approach, Uromonitor fell short of confirming the superior results anticipated from previous studies in this real-world setting. The search for an optimal urine-based biomarker for detecting and monitoring UCB remains ongoing. Results from this study highlight the complexity of developing non-invasive diagnostic tools and emphasise the importance of continued research efforts to refine these technologies.

PD-L1 expression in tumor and inflammatory cells is associated with favorable tumor features and favorable prognosis in muscle-invasive urothelial carcinoma of the bladder not treated by immune checkpoint inhibitors.

BACKGROUND: A high level of PD-L1 expression is the most relevant predictive parameter for response to immune checkpoint inhibitor (CPI) therapy in urinary bladder cancer. Existing data on the relationship between PD-L1 expression and the natural course of disease are controversial and sparse. METHODS: To expand our understanding of the relationship between PD-L1 expression and parameters of cancer aggressiveness, PD-L1 was analyzed on tissue microarrays containing 2710 urothelial bladder carcinomas including 512 patients with follow-up data who underwent radical cystectomy and follow-up therapies in the pre-immune checkpoint inhibitor therapy era. RESULTS: Tumor cell positivity in ≥10% of cells were seen in 513 (20%) and an immune cell positivity occurred in 872 (34%) of 2566 interpretable cancers. PD-L1 positivity in tumor cells increased from pTaG2 low grade (0.9% positive) to pTaG3 high grade (4.1%; p = 0.0255) and was even higher in muscle-invasive (pT2-4) carcinomas (29.3%; p < 0.0001). However, within pT2-4 carcinomas, PD-L1 positivity was linked to low pT stage (p = 0.0028), pN0 (p < 0.0001), L0 status (p = 0.0005), and a better prognosis within 512 patients with cystectomy who never received CPIs (p = 0.0073 for tumor cells and p = 0.0086 for inflammatory cells). PD-L1 staining in inflammatory cells was significantly linked to PD-L1 staining in tumor cells (p < 0.0001) and both were linked to a positive p53 immunostaining (p < 0.0001). CONCLUSION: It cannot be fully excluded that the strong statistical link between PD-L1 status and favorable histological tumor features as well as better prognosis could influence the outcome of studies evaluating CPIs in muscle-invasive urothelial carcinoma.

Nerve-sparing radical prostatectomy using the neurovascular structure-adjacent frozen-section examination (NeuroSAFE): results after 20 years of experience.

OBJECTIVES: To evaluate the long-term oncological outcomes and functional results of the neurovascular structure-adjacent frozen-section examination (NeuroSAFE) during nerve-sparing (NS) radical prostatectomy (RP). MATERIALS AND METHODS: A 10-yr survival analysis on 11069 RPs performed with or without the NeuroSAFE, between January 2002 to June 2011 was carried out. In the NeuroSAFE cohort, the neurovascular structure-adjacent prostatic margins are removed and stained for cryo-sectioning during RP. In case of a PSM, partial or full removal of the neurovascular bundle was performed. The impact of NeuroSAFE on biochemical recurrence-free survival (BFS), salvage radiation therapy-free survival, metastasis-free survival, and prostate cancer-specific survival at 10 years was analyzed. 1-year (1-yr) erectile function (EF), 1-yr, and 2-yr continence rates were assessed in propensity score-based matched cohorts. RESULTS: Median follow-up was 121 (IQR: 73, 156) months. No differences in BFS between NeuroSAFE and non-NeuroSAFE were recorded (10-yr BFS: NeuroSAFE vs non-Neurosafe, pT2: 81% vs 84%, p = 0.06; pT3a: 58% vs. 63%, p = 0.6; ≥pT3b: 22% vs. 27%, p = 0.99). No differences were found between the two groups in terms of sRFS (pT2: p = 0.1; pT3a: p = 0.4; ≥pT3b: p = 0.4) (Fig. 1B, Table 2), and MTS (pT2: p = 0.3; pT3a: p = 0.6; ≥pT3b: p = 0.9). The NeuroSAFE-navigated patients reported a better 1-yr EF than non-NeuroSAFE (68% vs. 58%, p = 0.02) and no differences in 1-yr and 2-yr continence rates (92.4% vs. 91.8%, and 93.4% vs. 93%, respectively). The main limitation is the retrospective study design. CONCLUSIONS: While the NeuroSAFE approach did not show significant improvements in long-term oncologic or continence outcomes, it did provide an opportunity for a higher proportion of patients to improve postoperative functional results, possibly through increased nerve-sparing procedures.

CEA (CEACAM5) expression is common in muscle-invasive urothelial carcinoma of the bladder but unrelated to the disease course.

Objectives: Carcinoembryonic antigen (CEA) is a cell surface glycoprotein that represents a promising therapeutic target. Serum measurement of shedded CEA can be utilized for monitoring of cancer patients. Material and Methods: To evaluate the potential clinical significance of CEA expression in urothelial bladder neoplasms, CEA was analysed by immunohistochemistry in more than 2500 urothelial bladder carcinomas in a tissue microarray format. Results: CEA staining was largely absent in normal urothelial cells but was observed in 30.4% of urothelial bladder carcinomas including 406 (16.7%) with weak, 140 (5.8%) with moderate, and 192 (7.9%) with strong staining. CEA positivity occurred in 10.9% of 411 pTaG2 low-grade, 32.0% of 178 pTaG2 high-grade, and 43.0% of 93 pTaG3 tumours (p < 0.0001). In 1335 pT2-4 carcinomas, CEA positivity (34.1%) was lower than in pTaG3 tumours. Within pT2-4 carcinomas, CEA staining was unrelated to pT, pN, grade, L-status, V-status, overall survival, recurrence free survival, and cancer specific survival (p > 0.25). Conclusion: CEA increases markedly with grade progression in pTa tumours, and expression occurs in a significant fraction of pT2-4 urothelial bladder carcinomas. The high rate of CEA positivity in pT2-4 carcinomas offers the opportunity of using CEA serum measurement for monitoring the clinical course of these cancers. Moreover, CEA positive urothelial carcinomas are candidates for a treatment by targeted anti-CEA drugs.

Prognostic Impact and Spatial Interplay of Immune Cells in Urothelial Cancer.

BACKGROUND AND OBJECTIVE: Quantity and the spatial relationship of specific immune cell types can provide prognostic information in bladder cancer. The objective of the study was to characterize the spatial interplay and prognostic role of different immune cell subpopulations in bladder cancer. METHODS: A total of 2463 urothelial bladder carcinomas were immunostained with 21 antibodies using BLEACH&STAIN multiplex fluorescence immunohistochemistry in a tissue microarray format and analyzed using a framework of neuronal networks for an image analysis. Spatial immune parameters were compared with histopathological parameters and overall survival data. KEY FINDINGS AND LIMITATIONS: The identification of > 300 different immune cell subpopulations and the characterization of their spatial relationship resulted in numerous spatial interaction patterns. Thirty-nine immune parameters showed prognostic significance in univariate analyses, of which 16 were independent from pT, pN, and histological grade in muscle-invasive bladder cancer. Among all these parameters, the strongest association with prolonged overall survival was identified for intraepithelial CD8+ cytotoxic T cells (time-dependent area under receiver operating characteristic curve [AUC]: 0.70), while stromal CD8+ T cells were less relevant (AUC: 0.65). A favorable prognosis of inflamed cancers with high levels of "exhaustion markers" suggests that TIM3, PD-L1, PD-1, and CTLA-4 on immune cells do not hinder antitumoral immune response in tumors rich of tumor infiltrating immune cells. CONCLUSIONS AND CLINICAL IMPLICATIONS: The density of intraepithelial CD8+ T cells was the strongest prognostic feature in muscle-invasive bladder cancer. Given that tumor cell killing by CD8+ cytotoxic T lymphocytes through direct cell-to-cell-contacts represents the "terminal end route" of antitumor immunity, the quantity of "tumor cell adjacent CD8+ T cells" may constitute a surrogate for the efficiency of cancer recognition by the immune system that can be measured straightaway in routine pathology as the CD8 labeling index. PATIENT SUMMARY: Quantification of intraepithelial CD8+ T cells, the strongest prognosticfeature identified in muscle-invasive bladder cancer, can easily be assessed by brightfield immunohistochemistry and is therefore "ready to use" for routine pathology.

Loss of TROP2 and epithelial cell adhesion molecule expression is linked to grade progression in pTa but unrelated to disease outcome in pT2-4 urothelial bladder carcinomas.

Introduction: Trophoblast cell surface antigen 2 (TROP2; EpCAM2) is a transmembrane glycoprotein which is closely related to EpCAM (EpCAM; EpCAM1). Both proteins share partial overlapping functions in epithelial development and EpCAM expression but have not been comparatively analyzed together in bladder carcinomas. TROP2 constitutes the target for the antibody-drug conjugate Sacituzumab govitecan (SG; TrodelvyTM) which has been approved for treatment of metastatic urothelial carcinoma by the United States Food and Drug administration (FDA) irrespective of its TROP2 expression status. Methods: To evaluate the potential clinical significance of subtle differences in TROP2 and EpCAM expression in urothelial bladder cancer, both proteins were analyzed by multiplex fluorescence immunohistochemistry in combination with a deep-learning based algorithm for automated cell detection on more than 2,700 urothelial bladder carcinomas in a tissue microarray (TMA) format. Results: The staining pattern of TROP2 and EpCAM were highly similar. For both proteins, the staining intensity gradually decreased from pTa G2 low grade (TROP2: 68.8±36.1; EpCAM: 21.5±11.7) to pTa G2 high grade (64.6±38.0; 19.3±12.2) and pTa G3 (52.1±38.7; 16.0±13.0, p<0.001 each). In pT2-4 carcinomas, the average TROP2 and EpCAM staining intensity was intermediate (61.8±40.9; 18.3±12.3). For both proteins, this was significantly lower than in pTa G2 low grade (p<0.001 each) but also higher than in pTa G3 tumors (p=0.022 for TROP2, p=0.071 for EpCAM). Within pT2-4 carcinomas, the TROP2 and EpCAM staining level was unrelated to pT, grade, UICC-category, and overall or tumor-specific patient survival. The ratio TROP2/EpCAM was unrelated to malignant phenotype and patient prognosis. Conclusion: Our data show that TROP2 and EpCAM expression is common and highly interrelated in urothelial neoplasms. Despite of a progressive loss of TROP2/EpCAM during tumor cell dedifferentiation in pTa tumors, the lack of associations with clinicopathological parameters in pT2-4 cancer argues against a major cancer driving role of both proteins for the progression of urothelial neoplasms.