Urine Cytology Research Articles

Evaluating artificial intelligence-enhanced digital urine cytology for bladder cancer diagnosis.

This study evaluated the diagnostic effectiveness of the AIxURO platform, an artificial intelligence-based tool, to support urine cytology for bladder cancer management, which typically requires experienced cytopathologists and substantial diagnosis time. One cytopathologist and two cytotechnologists reviewed 116 urine cytology slides and corresponding whole-slide images (WSIs) from urology patients. They used three diagnostic modalities: microscopy, WSI review, and AIxURO, per The Paris System for Reporting Urinary Cytology (TPS) criteria. Performance metrics, including TPS-guided and binary diagnosis, inter- and intraobserver agreement, and screening time, were compared across all methods and reviewers. AIxURO improved diagnostic accuracy by increasing sensitivity (from 25.0%-30.6% to 63.9%), positive predictive value (PPV; from 21.6%-24.3% to 31.1%), and negative predictive value (NPV; from 91.3%-91.6% to 95.3%) for atypical urothelial cell (AUC) cases. For suspicious for high-grade urothelial carcinoma (SHGUC) cases, it improved sensitivity (from 15.2%-27.3% to 33.3%), PPV (from 31.3%-47.4% to 61.1%), and NPV (from 91.6%-92.7% to 93.3%). Binary diagnoses exhibited an improvement in sensitivity (from 77.8%-82.2% to 90.0%) and NPV (from 91.7%-93.4% to 95.8%). Interobserver agreement across all methods showed moderate consistency (κ=0.57-0.61), with the cytopathologist demonstrating higher intraobserver agreement than the two cytotechnologists across the methods (κ=0.75-0.88). AIxURO significantly reduced screening time by 52.3%-83.2% from microscopy and 43.6%-86.7% from WSI review across all reviewers. Screening-positive (AUC+) cases required more time than negative cases across all methods and reviewers. AIxURO demonstrates the potential to improve both sensitivity and efficiency in bladder cancer diagnostics via urine cytology. Its integration into the cytopathological screening workflow could markedly decrease screening times, which would improve overall diagnostic processes.

Cytomorphological characteristics of low-grade papillary urothelial carcinoma in voided urine samples: Distinction from benign and high-grade papillary urothelial carcinoma.

Given its frequent recurrence and the potential for high-grade transformation, accurate diagnosis of low-grade papillary urothelial carcinoma (LGPUC) in urine cytology is clinically important. We attempted to identify cytomorphologic features in urine samples, which could be helpful for the identification of LGPUC. We conducted a retrospective review of voided urine specimens collected from patients with histopathologic diagnoses of LGPUC. Their cytomorphological features were compared with those from patients with benign conditions and high-grade papillary urothelial carcinoma (HGPUC). A total of 115 voided urine specimens were evaluated, including 30 benign, 41 LGPUC, and 44 HGPUC cases. In LGPUC, 18 cases (44%) were diagnosed as atypical, a proportion significantly higher than that observed in benign cases (4 cases, 13%), while the remaining 23 cases (56%) were diagnosed as negative. LGPUC urine samples tended to have higher cellularity than benign cases, but the difference was not statistically significant. Three cytological features, namely nuclear enlargement, higher nuclear-to-cytoplasmic (N/C) ratio, and presence of small cell clusters, were statistically more prevalent in LGPUC compared to benign cases, although the changes were relatively subtle. In contrast, cytomorphological distinction between LGPUC and HGPUC was evident, as high cellularity, nuclear enlargement, hyperchromasia, high N/C ratio, irregular nuclear membrane, and apoptosis were significantly more prevalent in HGPUC cases. Several cytomorphologic features in voided urine samples were more prevalent in cases with LGPUC, albeit not observed in all instances. Since these alterations were relatively subtle, meticulous attention to these cytomorphologic details is crucial to suggest the possibility of LGPUC.

Cytomorphologic comparison of upper urinary tract urothelial carcinomas and renal cell carcinomas on urine cytology.

Compared to urothelial carcinomas (UCs), the cytomorphology of renal cell carcinomas (RCCs) is underdescribed. This study aims to investigate whether UCs and RCCS of the upper urinary tract can be differentiated cytologically, and to identify distinguishing cytomorphological features. Consecutive urine cytology specimens with atypical/C3, suspicious/C4 or malignant/C5 diagnoses matched with a nephrectomy or ureterectomy specimen with UC or RCC over a 15-year period were reviewed for cellularity, architecture, background composition and cytomorphologic features. Totally 132 specimens were retrieved, comprising 24 RCCs and 108 UCs. Clear cell RCC (CCRCC) (n = 18) was the most common RCC. Urine cytology specimens from UC showed a trend of higher cellularity (p = 0.071) against RCC and was significant in subgroup analysis with CCRCC (p < .001). Epithelial structures in sheets, tubules, and papillae were exclusive in specimens of UC (p < .05). For background features, squamous cells were more common for RCC (p = .006) including CCRCC (p = .003), whereas polymorphs (p = .011) and necrotic material (p = .010) were associated with UC. Average nuclear size was larger and nuclear size variation (p < .001) and nuclear-cytoplasmic ratio (p = .001) were greater in UC (p = .001) than RCC. Comparing RCC to high-grade UCs only, nuclear-cytoplasmic ratio maintained statistical significance (p = .006) while average nuclear size showed a trend (p = .063). A clean background free of tumor necrosis and polymorphs, and the lack of complex tumor fragments favors RCC. UCs also display larger nuclear size, higher nuclear size variation and nuclear-cytoplasmic ratio. These cytomorphological features with corroboration of clinical/radiological findings, can aid in raising a diagnosis of RCC.

New edition of the Paris classification 2022: What is new?

As an internationally accepted diagnostic system, the Paris classification has achieved aglobal breakthrough in the standardization of diagnoses in urine cytology. Based on experience over the past few years since its first publication, the new edition of the Paris classification refines the diagnostic criteria and discusses diagnostic pitfalls. While the detection of high-grade urothelial carcinoma remains the main focus, other aspects of urine cytology, including cytology of the upper urinary tract and the associated challenges, have also been addressed. Low-grade urothelial neoplasia is no longer listed as aseparate category but is now included in the category "negative for high-grade urothelial carcinoma" (NGHUC). Essentially, the Paris classification provides an important basis for estimating the risk of malignancy and further clinical management.

Utility of Image Morphometry in the Atypical Urothelial Cells and High-Grade Urothelial Carcinoma Categories of the Paris System for Reporting Urinary Cytology.

Urinary cytology (UrCy) is highly sensitive to diagnosing high-grade urothelial carcinoma (HGUC) but cannot predict muscularis propria invasion. Further, the atypical urothelial cell category (AUC) may have variable outcomes. Image morphometry (IM) may be a valuable adjunct technique in this setting. Hence, we evaluated IM in the AUC and HGUC categories to improve the diagnostic performance. The following six nuclear parameters were evaluated by IM on 3150 cells: nucleo-cytoplasmic (N:C) ratio, nuclear area, diameter, perimeter, standard deviation of the nuclear area (SDNA; pleomorphism) and integrated density (ID; nuclear chromasia), using the ImageJ software, in three cohorts based on the histopathology outcome: 20 cases of AUC - benign non-neoplastic outcome (AUC-B); 22 cases of HGUC Muscle invasive (HGUC-MI) and 21 cases of HGUC non-muscle invasive (HGUC-MF). A retrospective analysis of urine cytology. The patient's ages ranged from 36 to 85 years, with a mean age of 60.6. The male-to-female ratio was 5.4:1. A total of 20 cases of AUC-B and 43 cases of HGUC were selected for IM analysis. HGUC cases had higher nuclear parameters than AUC-B, and HGUC-MI had higher SDNA, ID, diameter, and area than HGUC-MF. SDNA and ID predict muscularis propria invasion in HGUC. Image morphometry successfully differentiates HGUC cases from benign non-neoplastic ones and might help to identify muscularis propria invasion in HGUC using a combination of nuclear parameters.

Impact of implementing the first edition of the Paris system for reporting: A systematic review and meta-analysis.

Urine cytology is a noninvasive, widely used diagnostic tool for screening and surveillance of genitourinary tract neoplasms. However, the absence of unified terminology and clear objective morphological criteria limits the clinical benefit of urine cytology. The Paris System for Reporting Urine Cytology (TPS) was developed with the goal of standardizing reporting and improving urine cytology performance in detecting high-grade malignancy (HGM). We aimed to evaluate potential effects of TPS on improving urine cytology diagnostic performance and clinical utility by conducting a systematic review and meta-analysis. We searched six electronic databases to identify cross-sectional and cohort studies written in English assessing the accuracy of urine cytology in detecting genitourinary tract malignancies of patients under surveillance or with clinical suspicion of malignancy from January 2004 to December 2022. We extracted relevant data from eligible studies to calculate relative distribution of cytology diagnostic categories; ratio of atypical to HGM cytology diagnosis; and risk of HGM (ROHGM) and HGM likelihood ratio (HGM-LR) associated with cytology diagnostic categories. We used a generalized linear mixed model with logit transformation to combine proportions and multilevel mixed-effect logistic regression to pool diagnostic accuracy measurements. We performed meta-regression to evaluate any significant difference between TPS and non-TPS cohorts. We included 64 studies for 99,796 combined total cytology samples, across 31 TPS and 49 non-TPS cohorts. Pooled relative distribution [95% confidence interval (CI)] of negative for high-grade urothelial carcinoma (NHGUC)/negative for malignancy (NM); atypical urothelial cells (AUC); suspicious for high-grade urothelial carcinoma (SHGUC)/suspicious for malignancy (SM); low-grade urothelial neoplasm (LGUN); and HGM categories among satisfactory cytology cases were 83.8% (80.3%-86.9%), 8.0% (6.0%-10.6%), 2.2% (1.4%-3.3%), 0.01% (0.0%-0.1%), and 4.2% (3.2%-5.5%) in TPS versus 80.8% (76.8-2.7%), 11.3% (8.6%-14.7%), 1.8% (1.2%-2.7%), 0.01% (0.0%-0.1%), and 3.3% (2.5%-4.3%) in non-TPS cohorts. Adopting TPS classification resulted in a significant increase in the frequency of NHGUC and a reduction in AUC cytology diagnoses, respectively. The AUC/HGM ratio in TPS cohort was 2.0, which showed a statistically significant difference from the atypical/HGM ratio of 4.1 in non-TPS cohort (p-value: 0.01). Moreover, the summary rate (95% CI) of LGUN called AUC on cytology significantly decreased to 20.8% (14.9%-28.3%) in the TPS compared with 34.1% (26.4%-42.8%) in non-TPS cohorts. The pooled ROHGM (95% CI) was 20.4% (6.2%-50.0%) in nondiagnostic (NDX), 15.5% (9.6%-24.2%) in NHGUC, 40.2% (30.9%-50.2%) in AUC, 80.8% (72.9%-86.8%) in SHGUC, 15.1% (5.7%-34.3%) in LGUN, and 91.4% (87.3%-94.3%) in HGM categories in TPS studies. NHGUC, AUC, SHGUC, and HGM categories were associated with HGM-LR (95% CI) of 0.2 (0.1-0.3), 0.9 (0.6-1.3), 6.9 (2.4-19.9), and 16.8 (8.3-33.8). Our results suggest that TPS 1.0 has reduced the relative frequency of AUC diagnosis, AUC/HGM ratio, and the frequency of LGUNs diagnosed as AUC on cytology. Adopting this classification has improved the clinical utility of SHGUC and HGM cytology diagnoses in ruling in high-grade lesions. However, an NHGUC diagnosis does not reliably rule out the presence of a high-grade lesion.

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

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. 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. 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. 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.

Machine learning assisted microfluidics dual fluorescence flow cytometry for detecting bladder tumor cells based on morphological characteristic parameters

BackgroundBladder cancer (BC) is the most common malignant tumor and has become a major public health problem, leading the causes of death worldwide. The detection of BC cells is of great significance for clinical diagnosis and disease treatment. Urinary cytology based liquid biopsy remains high specificity for early diagnosis of BC, however, it still requires microscopy examination which heavily relies on manual operations. It is imperative to investigate the potential of automated and indiscriminate cell differentiation technology to enhance the sensitivity and efficiency of urine cytology. ResultsHere, we developed a machine learning algorithm empowered dual-fluorescence flow cytometry platform (μ-FCM) for urinary cytology analysis. A phenotype characteristic parameter (CP) which correlated with the size of the cell and nucleus was defined to achieve the differentiation of the BC cells and uroepithelial cells with high throughput and high accuracy. Based on CP analysis, SV-HUC-1 cells were almost differentiated from EJ cells and effectively reduced the overlap with 5637 cells. To further differentiate SV-HUC-1 cells and 5637 cells, support vector machine (SVM) machine learning algorithm was optimized to assist data analysis with the highest accuracies of 84.7 % for cell differentiation including the specificity of 91.0 % and the sensitivity of 75.0 %. Furthermore, the false positive rate (FPR) compensation enabled the detection rates of rare BC cells predicted by the well-trained SVM model were close to the true proportions with the recognition error in 0.4 % for the tumor cells. SignificanceAs a proof of concept, the developed μ-FCM system successfully demonstrates the capacity to identify the distribution of exfoliated cells in real urine samples. This system underscores the significance of integrating AI with microfluidics to perform high-throughput phenotyping of exfoliated cells, offering a pathway toward scalable, efficient, and automatic microfluidic systems in the fields of both biosensing and in vitro diagnosis of BC.

Violation of onco-surgical principles is associated with survival outcomes in upper tract urothelial carcinomas after radical nephroureterectomy.

Disease recurrence, particularly intravesical recurrence (IVR) after radical nephroureterectomy (RNU) for upper tract urothelial carcinoma (UTUC), is common. We investigated whether violations of onco-surgical principles before or during RNU, collectively referred to as surgical violation (SV), were associated with survival outcomes. Material and methods: Data from a consecutive series of patients who underwent RNU for UTUC 2001-2012 at Skåne University Hospital Lund/Malmö were collected. Preoperative insertion of a nephrostomy tube, opening the urinary tract during surgery or refraining from excising the distal ureter were considered as SVs. Survival outcomes in patients with and without SV (IVR-free [IVRFS], disease-specific [DSS] and overall survival [OS]) were assessed using multivariate Cox regression analyses (adjusted for tumour stage group, prior or concomitant bladder cancer, comorbidity and preoperative urinary cytology). Of 150 patients, 47 (31%) were subjected to at least one SV. Overall, SV was not associated with IVRFS (HR 0.81, 95% CI 0.4-1.6) but with worse DSS (HR 1.9, 95% CI 1.03-3.7) and OS (HR 1.9, 95% CI 1.2-3) in multivariable analysis. Additional analyses with a broader definition of SV including also preoperative instrumentation of the upper urinary tract (ureteroscopy and/or double J stenting) showed similar outcomes for DSS (HR 2.1, 95% CI 1.1-4.3). Worse survival outcomes, despite no difference in IVR, for patients that were subjected to the violation of sound onco-surgical principles before or during RNU for UTUC strengthen the notion that adhering to such principles is a cornerstone in upper tract urothelial cancer surgery.

Predicting the Lymphovascular Invasion in Carcinoma Bladder at Transurethral Resection.

Bladder cancer is among the most common malignant neoplasms in the world. Transurethral resection of bladder tumor (TURBT) is consideredthe standard procedure for diagnosis, staging, and risk classification of bladder tumors. Lymphovascular invasion (LVI) is considered a poor prognostic factor. Its assessment of TURBT is very important for risk stratification and decision-making for further treatment. The purpose of our clinical study is to attempt to predict/assess the correlation between LVI and various preoperative (age, gender, history of smoking, hematuria, urine cytology, and hydronephrosis/hydroureteronephrosis), intraoperative (tumor number, size, and appearance - sessile/ pedunculated) and histopathological (tumor histology, grading, and muscle invasion) factors. In this prospective study, 75 patients with bladder tumors underwent TURBT (standard monopolar TURBT with 1.5% glycine as irrigation solution) in the Department of Urology at Sri Venkateswara Institute of Medical Sciences (SVIMS), Tirupati between October 2021 and March 2023. Histopathological examination (HPE) reports were looked for the presence or absence of LVI. Accordingly, patients were divided into two groups, i.e., those with LVI and those without LVI. Various preoperative and intraoperative variables were analyzed for each subject in both groups. Statistically significant variables occurring in those patients with LVI compared to those without LVI were considered predictors of LVI in bladder tumors. Results: Sixteen patients out of 75 (21.33%) had LVI on their histopathology examination. The mean age was 68.19 years in the group with LVI and 64.14 years in the group without LVI. A total of 60 men (80%) and 15 women (20%) were included in our study. Thirteen men (21.7%) and three women (20%) were found to have LVI. We observed a significant association between the appearance of the tumor and LVI. Fifty-four subjects in our study had sessile tumors. Fifteen out of them (27.8%) hadLVI, while only one out of 21 patients (4.8%) with pedunculated tumors had LVI (p-value=0.028). 30% of subjects who had high-grade tumors on HPE also had LVI. On the contrary, only one of 25 patients (4%) with low-grade tumors had LVI (p-value=0.010). Our study also showed a significant association between muscle invasion and LVI. Thirty-four (45.3%) and 41 (54.7%) patients had muscle-invasive and non-muscle-invasive tumors, respectively. While 12 (35.3%) patients with muscle-invasive tumors had LVI, only four (9.8%) patients with non-muscle-invasive tumors showed LVI (p-value=0.007). We observed that LVI of bladder tumors at first TURBT is significantly associated with tumor grade, tumor appearance, and depth of invasion of the tumor. Though statistically not significant, we further observed that LVI was more commonly found in smokers, patients with hematuria, and larger tumor sizes. We conclude that these factors can be used as reliable predictors of LVI of bladder tumors at their first TURBT.

Computer-assisted urine cytology: Faster, cheaper, better?

Recent advancements in computer-assisted diagnosis (CAD) have catalysed significant progress in pathology, particularly in the realm of urine cytopathology. This review synthesizes the latest developments and challenges in CAD for diagnosing urothelial carcinomas, addressing the limitations of traditional urinary cytology. Through a literature review, we identify and analyse CAD models and algorithms developed for urine cytopathology, highlighting their methodologies and performance metrics. We discuss the potential of CAD to improve diagnostic accuracy, efficiency and patient outcomes, emphasizing its role in streamlining workflow and reducing errors. Furthermore, CAD tools have shown potential in exploring pathological conditions, uncovering novel biomarkers and prognostic/predictive features previously unknown or unseen. Finally, we examine the practical issues surrounding the integration of CAD into clinical practice, including regulatory approval, validation and training for pathologists. Despite the promising results, challenges remain, necessitating further research and validation efforts. Overall, CAD presents a transformative opportunity to revolutionize diagnostic practices in urine cytopathology, paving the way for enhanced patient care and outcomes.

From Detection to Cure - Emerging Roles for Urinary Tumor DNA (utDNA) in Bladder Cancer.

This review sought to define the emerging roles of urinary tumor DNA (utDNA) for diagnosis, monitoring, and treatment of bladder cancer. Building from early landmark studies the focus is on recent studies, highlighting how utDNA could aid personalized care. Recent research underscores the potential for utDNA to be the premiere biomarker in bladder cancer due to the constant interface between urine and tumor. Many studies find utDNA to be more informative than other biomarkers in bladder cancer, especially in early stages of disease. Points of emphasis include superior sensitivity over traditional urine cytology, broad genomic and epigenetic insights, and the potential for non-invasive, real-time analysis of tumor biology. utDNA shows promise for improving all phases of bladder cancer care, paving the way for personalized treatment strategies. Building from current research, future comprehensive clinical trials will validate utDNA's clinical utility, potentially revolutionizing bladder cancer management.

An assessment of chromosomal alterations detected by FISH in urothelial carcinoma and its correlation to liquid-based urine cytology and histopathology

BackgroundThe present study aimed to evaluate the diagnostic usefulness, sensitivity, and specificity of urinary liquid-based cytology (LBC) versus multitarget, multicolor UroVysion fluorescence in situ hybridization (uFISH) assay for the diagnosis of urinary bladder cancer in light of the histological diagnosis. MethodsFifty patients and negative controls each were selected for the study. A 10 ml voided urine specimen was processed for urine cytology to prepare LBC smears. For urinary FISH analysis, 50 ml of voided urine was processed using multi-target FISH-containing probes for the centromeres of chromosomes 3, 7, and 17 and the 9p21 locus. Histopathological examination of transurethral resection of bladder tumor (TURBT) chips was performed on diseased cases only. ResultsAll 50 diseased cases exhibited 62%, 80%, 76%, and 68% of uFISH sensitivity, uFISH specificity, positive predictive value (PPV) and negative predictive value (NPV) respectively. Among negative controls, results of 40%, 84%, 71%, and 58% for LBC sensitivity, LBC specificity, PPV, and NPV were found, respectively. Among the diseased cases uFISH positivity of 20%, 45%, 83.3%, 82.4% and 100% for Ta, T1, T2a, T2b and T3 respectively. Among the diseased cases LBC positivity was 0%, 20%, 33.3%, 70% and 100% for Ta, T1, T2a, T2b and T3 respectively. ConclusionIncreased sensitivity and specificity of uFISH than LBC urine to detect urothelial carcinoma (UC) reflect the better potentiality of the former to detect UCs. The limitations of uFISH analysis include sophisticated equipment, higher cost, and experienced cytopathologists/technicians. It is proposed to conduct more such prospective trials with recent biomarkers so as to reduce the frequency of cystoscopy in such patients.

Sequential Intravesical Chemotherapy for Treatment Naïve, High-Risk Non-Muscle Invasive Bladder Cancer: Oncologic Outcomes, Tolerance, and Comparison to Contemporary Controls

ObjectivesTo assess if receiving sequential intravesical chemotherapy (Gemcitabine-Docetaxel, Gem-Doce) therapy was associated with similar oncologic efficacy to bacillus Calmette-Guérin (BCG) in patients with treatment-naïve, high-risk non-muscle invasive bladder cancer (HR-NMIBC). MethodsSingle-center, retrospective cohort study of 80 patients with HR-NMIBC initiating first-line Gem-Doce or BCG between August 2020 and August 2023. Surveillance conducted with cystoscopy, urine cytology, and cross-sectional imaging. The primary oncologic outcome was high-grade bladder tumor recurrence during surveillance. Kaplan-Meier method applied to determine 12- and 24-month recurrence free survival (RFS) after initiation of therapy. Tolerance of each intravesical therapies was assessed. Results53/80 (66%) received Gem-Doce and 27/80 (34%) received BCG with overall 18-month median follow up. There were 10 recurrences after Gem-Doce and seven after BCG. The RFS at 12- and 24-months for Gem-Doce (12-months: 87%, 24-months: 75%) was not significantly different than BCG (12-months: 85%, 24-months: 81%). Lastly, Gem-Doce had significantly fewer patients with AEs compared to BCG (40% vs 74%). Limitations include retrospective design, small cohort size, and intermediate oncologic follow-up. ConclusionsOur data suggest that sequential intravesical Gem-Doce is an oncologically efficacious and, potentially better tolerated, alternative to BCG for treatment-naïve HR-NMIBC.

Nuclear long diameter as a new criterion to distinguish high-grade urothelial carcinoma cells from benign reactive cells.

Recently, the nuclear area has attracted attention as a morphological parameter to differentiate high-grade urothelial carcinoma (HGUC) cells from benign reactive cells. The nuclear long diameter (NLD) strongly correlates with the nuclear area and is easy to subjectively estimate. Therefore, this study examined the usefulness of the NLD-to-neutrophil diameter ratio for detecting HGUC cells in urine cytology. This study included 29, 26 and 18 patients with HGUC, glomerular disease and urolithiasis respectively. An image analysis system was used to measure the NLD of HGUC and benign reactive cells (reactive renal tubular cells and reactive urothelial cells) and the neutrophil diameter that appeared in the voided urine in these cases. The NLD index was calculated using the NLD-to-neutrophil diameter ratio. We subsequently compared HGUC and benign reactive cells with respect to NLD and NLD indices. In addition, the HGUC cell group and benign reactive cell group were compared by selecting the five cells with the largest NLD and NLD index on each slide. The NLD and NLD indices of HGUC cells were significantly higher than those of benign reactive cells in all cells and in the five cells with the largest NLD and NLD indices. The cut-off value of the NLD index for detecting HGUC cells was 1.25 in all cells and 1.80 in the five cells with the largest NLD index. The NLD index is a useful parameter that can be introduced into routine microscopic examinations to differentiate HGUC cells from benign reactive cells.

The value of urinary exosomal microRNA-21 in the early diagnosis and prognosis of bladder cancer.

Bladder cancer (BC) poses high morbidity and mortality, with urinary exosomal microRNA (miR)-21 showing potential value in its diagnosis and prognosis, and we probed its specific role. We prospectively selected 116 BC patients and 116 healthy volunteers as the BC and control groups, respectively. BC urinary exosomal miR-146a-5p, miR-93-5p, miR-663b, miR-21, and miR-4454 relative expression levels were assessed. The correlations between clinical indexes and urinary exosomal miR-21, prognostic value of miR-21, and diagnostic value of the five candidate miRNAs, urine cytology, and miRNA joint diagnostic panel for BC and urinary exosomal miR-21, miR-4454, and urine cytology for Ta-T1 and T2-T4 stage BC were analyzed. Urinary exosomal miR-146a-5p, miR-93-5p, miR-663b, miR-21, and miR-4454 were highly expressed in BC patients. miR-146a-5p, miR-93-5p, miR-663b, miR-21, miR-4454, miRNA combined diagnostic panel, and urine cytology had certain diagnostic value for BC, with miR-21, miR-4454, and miRNA co-diagnostic panel showing the highest diagnostic value. Collectively, urinary exosomal miR-21 was closely related to Tumor-Node-Metastasis staging and grading in BC patients. Urinary exosomal miR-21 had high diagnostic value for BC and Ta-T1 and T2-T4 stage BC, and had high predictive value for BC poor prognosis, providing an effective indicator for the occurrence, development, and prognostic assessment of BC.

P53 and Vimentin Double Immunostaining to Differentiate between High-Grade Urothelial Carcinoma Cells and Benign Atypical Cells

Introduction: Urine cytology is an indispensable test for detecting high-grade urothelial carcinoma (HGUC); however, the distinction between HGUC cells and morphologically similar benign atypical cells poses clinical challenges. In this study, we performed double immunostaining for p53 and vimentin to establish a diagnostic method to accurately distinguish HGUC cells from benign atypical cells. Methods: This study included 41 cases of HGUC, 11 of urolithiasis, and 22 of glomerular disease diagnosed histopathologically or clinically. After preparing urine cytology specimens from voided urine samples, p53 immunostaining was performed, and the p53-positive intensity and p53 positivity rate were calculated. Subsequently, vimentin immunostaining was performed on the same specimens to calculate the rate of vimentin positivity. Results: The HGUC cell group had a mean p53-positive intensity of 2.40, a mean p53 positivity rate of 73.2%, and a mean vimentin positivity rate of 5.1%. In contrast, the mean p53-positive intensity, p53 positivity rate, and vimentin positivity rate were 1.63, 36.7%, and 66.2%, respectively, in the benign atypical cell group. There were significant differences between the two groups for each parameter. Moreover, two multiple logistic regression models combining the results of these three parameters exhibited higher sensitivity and specificity than solely assessing the p53-positive intensity, positivity rate, and vimentin positivity rate. Conclusion: Since double immunostaining with p53 and vimentin distinguishes HGUC cells from benign atypical cells, it could be to improve the diagnostic accuracy of urine cytology.

Routine Surveillance of Upper Urinary Tract Imaging for Diagnosing Upper Urinary Tract Urothelial Cancer Recurrence in Patients with Nonmuscle Invasive Bladder Cancer.

Although routine surveillance imaging to examine upper urinary tract urothelial cancer recurrence during follow-up of nonmuscle invasive bladder cancer is recommended, its necessity remains invalidated. A single-institute long-term follow-up cohort study to evaluate the clinical impact of routine surveillance imaging and identify risk factors for upper urinary tract urothelial cancer recurrence after nonmuscle invasive bladder cancer treatment was conducted. A retrospective chart review of 864 patients with primary nonmuscle invasive bladder cancer who underwent initial transurethral resection of bladder tumor between 1980 and 2020 was conducted. The opportunities to diagnose its recurrence were examined. Moreover, oncological outcomes included upper urinary tract urothelial cancer recurrence-free survival and overall survival. Of 864 patients, 19 (2.2%) experienced upper urinary tract urothelial cancer recurrence. Among 19 patients, recurrence was detected through routine imaging in 12 (63.2%), cystoscopy in 2 (10.5%), urine cytology in 2 (10.5%), and presence of gross hematuria in 1 (5.3%). All patients had high- or highest-risk NMIBC at diagnosis of primary nonmuscle invasive bladder cancer. On multivariate Fine-Gray proportional regression analyses, a tumor size of ≥30 mm and carcinoma in situ were independently associated with short upper urinary tract urothelial cancer recurrence-free survival (P=0.040 and 0.0089, respectively). Most patients experiencing upper urinary tract urothelial cancer recurrence were diagnosed by routine surveillance imaging, suggesting its clinical importance, especially for patients with nonmuscle invasive bladder cancer accompanied by a tumor size of ≥30 mm and carcinoma in situ.

Abstract B018: KDM6A and MTAP expression loss enable identification of a large fraction of low grade non-invasive urothelial neoplasia of the urinary bladder

Abstract The identification of low-grade urothelial neoplasia in cytologic samples or in biopsies with dysplastic lesions poses a particular problem for pathologists in routine diagnostics. Improved diagnosis of low-grade urothelial neoplasia could help to reduce the number of cystoscopic follow-up examinations. KDM6A, also known as UTX (ubiquitously transcribed X chromosome tetratricopeptide repeat protein) is an epigenetic regulator which is frequently mutated in urothelial carcinoma. Because KDM6A is localized on chromosome Xp11.3, and because of the male predominance in bladder cancer, many KDM6A mutations should result in a complete loss of KDM6A protein expression and become detectable by immunohistochemistry (IHC). Similarly, S-methyl-5′-thioadenosine phosphorylase (MTAP), a critical enzyme for DNA and RNA synthesis, is co-deleted with the CDKN2A tumor suppressor located on chromosome 9p21 in about 25% of bladder cancers, resulting in reduced or lost protein expression. To study the prevalence and diagnostic potential of immunohistochemical loss of KDM6A and MTAP expression, 928 tumors were analyzed by IHC in a tissue microarray format. The cohort included 628 non-invasive papillary tumors (pTa) and 300 tumors from patients who underwent radical cystectomy for muscle-invasive disease (pT2-4). A complete KDM6A expression loss occurred in 35% of 342 pTa G2 low-grade, 23% of 150 pTa G2 high-grade, 17.8% of 90 pTa G3 tumors, and 20.1% of 243 interpretable muscle invasive (pT2-4) cancers (p&amp;lt;0.0001). Within pTa tumors, there was a strong correlation between KDM6A loss and low tumor grade (p=0.0003), whereas no significant difference was found between pT3 G3 and muscle invasive cancers (p=0.8828). Complete absence of MTAP expression occurred in 11% of 297 pTa G2 low-grade, 29.6% of 125 pTa G2 high-grade, 18.4% of 76 pTa G3 tumors, and 26.3% of 217 muscle invasive (pT2-4) cancers (p&amp;lt;0.0001). A combined analysis of 702 cancers with interpretable data for KDM6A and MTAP showed that 6.6% of tumors had expression loss of both KDM6A and MTAP, 20% had KDM6A loss only, and 13% had MTAP loss only. The rate of expression loss of KDM6A and/or MTAP was markedly higher in pTa G2 low (41.2%) and in pTa G2 high (40.3%) than in pTa G3 tumors (29%). In summary, our data demonstrate that about 40% of low-grade non-invasive urothelial neoplasias – which are usually not detected by urine cytology – had a completed expression loss of either KDM6A or MTAP. In patients with these alterations, follow-up could be potentially facilitated by immunohistochemistry. Citation Format: Florian Viehweger, Neele Heckmann, Natalia Gorbokon, Henning Plage, Sebastian Hofbauer, Sarah Weinberger, Florian Roßner, Simon Schallenberg, Sefer Elezkurtaj, Maximilian Lennartz, Tim Mandelkow, Elena Bady, Margit Fisch, Thorsten Ecke, Stefan Koch, Nico Adamini, Ronald Simon, Guido Sauter, Tobias Klatte, Thorsten Schlomm, David Horst, Henrik Zecha, Martina Kluth, Sarah Minner. KDM6A and MTAP expression loss enable identification of a large fraction of low grade non-invasive urothelial neoplasia of the urinary bladder [abstract]. In: Proceedings of the AACR Special Conference on Bladder Cancer: Transforming the Field; 2024 May 17-20; Charlotte, NC. Philadelphia (PA): AACR; Clin Cancer Res 2024;30(10_Suppl):Abstract nr B018.

Abstract IA020: AI-powered tool for rapid &amp; reliable bladder cancer screening and surveillance: multicenter validation efforts

Abstract Bladder cancer is the most recurrent form of cancer, requiring frequent screening and surveillance, which contributes to its status as the costliest per capita. While urine cytology serves as a crucial adjunct to invasive cystoscopy, it is high volume, labor-intensive, and relatively inconsistent. Despite the implementation of The Paris System, which introduced updated quantitative guidelines, implementation of digital tools for urine cytology still trails the technological advancements seen in semi-automated Pap smear screenings. This lag can be attributed to the diverse cytomorphology of urine specimens and variation in specimen preparation and screening. Our team has developed AutoParis-X (APX), a state-of-the-art technology leveraging deep learning algorithms to extract objective (e.g., nucleus-to-cytoplasm ratio) and subjective (e.g., hyperchromasia) features from whole slide images (WSI). APX transforms these features into the Atypia Burden Score (ABS), offering a quantitative assessment of malignancy and recurrence risks on a scale from 0 to 1. Initially validated with 1300 samples (ThinPrep; Leica Aperio GT450) at Dartmouth Hitchcock, achieving an AUROC of 0.9, APX has evolved into APX-WEB, a user-friendly, web-based tool for cytopathologists. Notable challenges persist, including: 1) validating across nationally representative cohorts with varied specimen preparations and imaging, 2) addressing the intricacies of z-stacking for imaging non-monolayer preparations, and 3) integration of genomics to heighten the sensitivity of imaging assays. This study aimed to establish a performance baseline for APX's adaptability to these challenges without algorithmic adjustments. APX was applied across three distinct cohorts: 1) a preliminary national validation on 500 ThinPrep WSI in collaboration with Hologic; 2) assessment on non-monolayer specimen preparations (e.g., SurePath), using z-stacking, conducted with Johns Hopkins (n=100 WSI); and 3) a pilot study at Dartmouth (n=19) for integrating APX's imaging features with whole exomic sequencing (WES). Preliminary findings indicate: 1) Promising performance on a held-out Hologic national cohort (sensitivity=0.87, specificity=0.73; original APX study: sensitivity=0.88, specificity=0.83). 2) In SurePath z-stack WSI, the number of detected urothelial cells per cluster decreased with distance from the focus plane (rho =-0.13, p&amp;lt;0.001). 3) Of eight cases deemed negative or atypical by cytology, three exhibited TERT mutations, suggesting subclinical recurrence, in contrast to the consistent findings in all nine suspicious or positive cases. This highlights the potential of integrating genomics with image-based classifiers to enhance diagnostic precision. The findings underscore the need for future multicenter work aimed at refining machine learning models to achieve enhanced predictive performance across various operational parameters and settings, facilitating their implementation. Citation Format: Joshua Levy, Keluo Yao, Jonathan Marotti, Darcy Kerr, Edward Gutmann, Sam Harvey, Yoseph Sayegh, Chris VandenBussche, Michael Quick, Peter Costa, Lauren Wainman, Donald Green, Parth Shah, Xiaoying Liu, Louis Vaickus. AI-powered tool for rapid &amp; reliable bladder cancer screening and surveillance: multicenter validation efforts [abstract]. In: Proceedings of the AACR Special Conference on Bladder Cancer: Transforming the Field; 2024 May 17-20; Charlotte, NC. Philadelphia (PA): AACR; Clin Cancer Res 2024;30(10_Suppl):Abstract nr IA020.