Current perspectives of urine-based tests for screening and post-treatment monitoring of urothelial carcinoma of the bladder

Introduction

Urothelial carcinoma (UC) originates from cells lining the upper (renal pelvis and ureter) and lower (bladder and urethra) urinary tracts. The Global Cancer Statistics 2020 report states that bladder cancer (BCa) is the seventh most common cancer among men worldwide and the thirteenth most common cancer in Japan (1,2). BCa is one of the most financially burdensome malignancies globally, spanning the diagnostic to terminal stages. Early detection, diagnosis, and intervention are crucial for reducing BCa mortality. In most patients with BCa, microscopic hematuria is the initial presentation.

Because cystoscopy is invasive and costly, both physicians and patients often hesitate to pursue adequate evaluation, which increases the risk of missed opportunities for early BCa detection and diagnosis. The American Urological Association/Society of Urodynamics, Female Pelvic Medicine & Urogenital Reconstruction (AUA/SUFU) microhematuria guidelines recommend evaluating patients with hematuria using a risk-stratification system (3). This system, based on extensive literature and systematic reviews, categorizes patients as follows:

• Low-risk (estimated UC risk <1%): women <50 years, men <40 years; never smoker or <10 pack years; 3–10 red blood cells (RBC)/high-power field (hpf) on a single urinalysis; no UC risk factors.
• Intermediate-risk (estimated UC risk 1–2%): women aged 50–59 years, men aged 40–59 years; 10–30 pack years; 11–25 RBC/hpf on a single urinalysis; low-risk patients without prior evaluation and 3–10 RBC/hpf on repeat urinalysis; additional UC risk factors.
• High-risk (estimated UC risk ≥10%): women or men aged ≥60 years; >30 pack years; >25 RBC/hpf on a single urinalysis; history of gross hematuria.

Patients classified as low risk may opt for surveillance instead of immediate cystoscopy, as UC is detected in approximately 0.8% of low-risk patients with microhematuria (4). This approach reduces the burden of potentially unnecessary cystoscopies. Urine is an ideal biological fluid for noninvasive UC diagnosis. However, conventional urine cytology cannot replace cystoscopy due to its low sensitivity and negative predictive value (NPV). A urine-based biomarker with high diagnostic accuracy, particularly high sensitivity and NPV, could help reduce unnecessary cystoscopies by facilitating the identification of individuals with cancer.

Results of the STRATA trial

As reported by Lotan et al. in a recent issue of The Journal of Urology, the STRATA trial was a prospective randomized controlled study comparing CxBladder Triage (CxbT; Pacific Edge Diagnostics, Dunedin, New Zealand) with cystoscopy in patients with asymptomatic microhematuria (5). CxbT measures the mRNA expression of five biomarkers (MDK, CDK1, IGFBP5, HOXA13, and CXCR2) in voided urine samples using quantitative reverse transcription polymerase chain reaction (PCR) (6). CXCR2 is a “negative” marker due to its high expression in neutrophils and inflammatory conditions, while the other four markers are overexpressed in BCa cells. Combined with clinical parameters such as sex, age, smoking history, and visible hematuria, the CxbT-specific algorithm generates a score ranging from 1 to 10. Scores <4 indicate a low probability of UC, with a sensitivity of 95.1% and an NPV of 98.5% (6). A prior study showed that 40% of patients were accurately triaged with a low probability of UC (6). Lotan et al. hypothesized that CxbT could triage patients requiring evaluation by identifying those at high risk of BCa while reducing evaluations in low-risk patients with negative CxbT results (5).

Patients included in the STRATA trial were classified as having lower risk (3–29 RBCs/hpf and smoking <10 pack-years) or not lower risk (>29 RBCs/hpf and/or smoking >10 pack-years). Low-risk patients were randomized into the CxbT marker or standard of care (SOC) groups. Among 390 eligible patients, 135 (34.6%) with low-risk were subsequently randomized to either the CxbT-informed decision or SOC groups. Overall, 63% of CxbT test results were negative. All patient groups underwent conventional flexible white-light cystoscopy per SOC. Among non-low-risk patients, 82% underwent cystoscopy. In low-risk patients, cystoscopy was performed in 67% of the SOC group and 27% of the CxbT marker group [relative risk: 0.41, 95% confidence interval (CI): 0.27–0.61], corresponding to a 59% reduction. Compared with cystoscopy, CxbT demonstrated 90% sensitivity, 56% specificity, and 99% NPV for detecting UC. The authors concluded that the clinical utility of CxbT in safely reducing unnecessary cystoscopies in low-risk patients with microhematuria could theoretically lower patient morbidity and discomfort, improve access to care, and reduce environmental impact.

Clinical utility of CxBladder tests

CxBladder tests are commercially available in Australia, New Zealand, Singapore, the United States, Israel, Argentina, Uruguay, Venezuela, Vietnam, the Philippines, Malaysia, and Brunei. These tests comprise four options tailored to different clinical needs and situations: CxBladder Detection (CxbD), CxbT, CxBladder Resolve (CxbR), and CxBladder Monitoring. CxbD and CxbT were developed to detect and rule out BCa in patients presenting with hematuria (7). CxbR is designed for use following CxbD and CxbT. In cases where CxbT yields positive results, it reflects the positivity to CxbR, enabling identification of patients with a high probability of high-impact tumors, such as high-grade Ta, Tis, or T1–3 late-stage BCa. The sensitivity and specificity of the test were 92.4% (95% CI: 83.3–96.7%) and 93.8% (95% CI: 86.8–97.2%), respectively, when selecting exclusively for high-grade or late-stage tumors (8). This enhances clinical decision-making and facilitates the accurate segregation of patients with high-impact tumors. Additionally, CxBladder Monitoring reliably rules out recurrent UC. The sensitivity and NPV were 93% and 97%, respectively, outperforming all other Food and Drug Administration (FDA)-approved urine tests in excluding patients with a low risk of recurrent UC (9). The CxBladder Monitor reduces the need for routine cystoscopy and other costly invasive procedures, allowing urologists to concentrate on patients at higher risk of recurrence who require more intensive clinical evaluation.

Other urine-based biomarkers for screening and post-treatment monitoring of BCa

In several decades, numerous efforts have been made to develop useful urine-based tests using various types of biomaterials, target markers, and assays to decrease or omit cystoscopies. Table 1 lists several urine-based tests, including urine cytology, immunoCyt/uCyt+, UroVysion, NMP-22, and bladder tumor antigen (BTA), that have been approved by FDA (10-12). The sensitivities of urine-based biomarkers are superior to that of urine cytology, whereas the specificities are inferior in most of the biomarkers. A systematic review and meta-analysis compared the diagnostic performance among CxBladder, AssureMDx, BTA, Nuclear Matrix Protein 22 (NMP22), UroVysion, and ImmunoCyt/uCyt+ in the evaluation of primary hematuria for screening of BCa (13). A total of 18 studies were included in the analysis, demonstrating the sensitivity ranged from 65.9% to 97.3%, and the specificity ranged from 57.7% and 83.3% across the six tests. AssureMDx, CxBladder, and ImmunoCyt/uCyt+ had better diagnostic performance based on their sensitivity, specificity, diagnostic odds ratio, and positive and negative likelihood ratios. Current diagnostic capability of the FDA-approved urine-based tests remain insufficient for wide application as a triage test for cystoscopy in patients with primary hematuria.

Another mRNA-based test, the Xpert^Ò Bladder Cancer (Cepheid, Sunnyvale, CA, USA) quantifies five mRNA targets (ABL1, CRH, IGF2, UPK1B, and ANXA10) by real-time PCR. All the five genes are overexpressed in BCa cells. Xpert Bladder Cancer encompasses two tests to meet different clinical needs and situations: Xpert^Ò Bladder Cancer Detection and Monitor. The first prospective study that included 155 urine samples from 140 patients with a history of non-muscle-invasive bladder cancer (NMIBC) showed high sensitivity (84%) and specificity (91%) of the Xpert^Ò Bladder Cancer Monitor (14). Only few reports are available on Xpert Bladder Cancer Detection for screening and initial detection, and one of which shows a high sensitivity of 100% and low specificity of 4.5% (15). It might be necessary to determine an appropriate risk calculation algorism for this diagnostic setting.

Summary

Patients and physicians have long preferred reducing the number of cystoscopies for hematuria screening and post-treatment monitoring. DNA alteration- and mRNA-based urinary tests have demonstrated high diagnostic accuracy for detecting UC, including low-grade tumors, as reported in previous studies. In the STRATA trial (5), researchers validated the clinical utility of CxbT in safely reducing unnecessary cystoscopies in low-risk patients with microhematuria. However, this study has limitations, including the absence of patient quality of life (QOL) assessment and economic impact analysis of CxbT in patients with microhematuria. Additionally, external validation using diverse cohorts is currently lacking. Further post-hoc analyses of the STRATA trial and prospective studies designed to confirm the high cost-effectiveness of these tests are needed to support their widespread adoption.

Acknowledgments

None.

Footnote

Provenance and Peer Review: This article was commissioned by the editorial office, Translational Andrology and Urology. The article has undergone external peer review.

Peer Review File: Available at https://tau.amegroups.com/article/view/10.21037/tau-2025-27/prf

Funding: None.

Conflicts of Interest: The author has completed the ICMJE uniform disclosure form (available at https://tau.amegroups.com/article/view/10.21037/tau-2025-27/coif). The author has no conflicts of interest to declare.

Ethical Statement: The author is accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved.

Open Access Statement: This is an Open Access article distributed in accordance with the Creative Commons Attribution-NonCommercial-NoDerivs 4.0 International License (CC BY-NC-ND 4.0), which permits the non-commercial replication and distribution of the article with the strict proviso that no changes or edits are made and the original work is properly cited (including links to both the formal publication through the relevant DOI and the license). See: https://creativecommons.org/licenses/by-nc-nd/4.0/.

References

1. Sung H, Ferlay J, Siegel RL, et al. Global Cancer Statistics 2020: GLOBOCAN Estimates of Incidence and Mortality Worldwide for 36 Cancers in 185 Countries. CA Cancer J Clin 2021;71:209-49. [Crossref] [PubMed]
2. Matsuda T, Okuyama A. Incidence rate for bladder cancer in Japanese in Japan and in the United States from the Cancer Incidence in Five Continents. Jpn J Clin Oncol 2017;47:284-5. [Crossref] [PubMed]
3. Barocas DA, Boorjian SA, Alvarez RD, et al. Microhematuria: AUA/SUFU Guideline. J Urol 2020;204:778-86. [Crossref] [PubMed]
4. Woldu SL, Ng CK, Loo RK, et al. Evaluation of the New American Urological Association Guidelines Risk Classification for Hematuria. J Urol 2021;205:1387-93. [Crossref] [PubMed]
5. Lotan Y, Daneshmand S, Shore N, et al. A Multicenter Prospective Randomized Controlled Trial Comparing Cxbladder Triage to Cystoscopy in Patients With Microhematuria: The Safe Testing of Risk for Asymptomatic Microhematuria Trial. J Urol 2024;212:41-51. [Crossref] [PubMed]
6. Kavalieris L, O’Sullivan PJ, Suttie JM, et al. A segregation index combining phenotypic (clinical characteristics) and genotypic (gene expression) biomarkers from a urine sample to triage out patients presenting with hematuria who have a low probability of urothelial carcinoma. BMC Urol 2015;15:23. [Crossref] [PubMed]
7. Harvey JC, Cambridge LM, Ellen CW, et al. Analytical Validation of Cxbladder(®) Detect, Triage, and Monitor: Assays for Detection and Management of Urothelial Carcinoma. Diagnostics (Basel) 2024;14:2061. [Crossref] [PubMed]
8. Raman JD, Kavalieris L, Konety B, et al. The Diagnostic Performance of Cxbladder Resolve, Alone and in Combination with Other Cxbladder Tests, in the Identification and Priority Evaluation of Patients at Risk for Urothelial Carcinoma. J Urol 2021;206:1380-9. [Crossref] [PubMed]
9. Lotan Y. Clinical comparison of noninvasive urine tests for ruling out recurrent urothelial carcinoma. Urol Oncol 2017;35:531.e15-22. [Crossref] [PubMed]
10. Miyake M, Owari T, Hori S, et al. Emerging biomarkers for the diagnosis and monitoring of urothelial carcinoma. Res Rep Urol 2018;10:251-61. [Crossref] [PubMed]
11. Flores Monar GV, Reynolds T, Gordon M, et al. Molecular Markers for Bladder Cancer Screening: An Insight into Bladder Cancer and FDA-Approved Biomarkers. Int J Mol Sci 2023;24:14374. [Crossref] [PubMed]
12. Tomiyama E, Fujita K, Hashimoto M, et al. Urinary markers for bladder cancer diagnosis: A review of current status and future challenges. Int J Urol 2024;31:208-19. [Crossref] [PubMed]
13. Soputro NA, Gracias DN, Dias BH, et al. Utility of urinary biomarkers in primary haematuria: Systematic review and meta-analysis. BJUI Compass 2022;3:334-43. [Crossref] [PubMed]
14. van Valenberg FJP, Bridge JA, Mayne D, et al. Validation of a mRNA-based urine test for bladder cancer detection in patients with hematuria. Eur Urol Suppl 2017;16:e190-1. [Crossref]
15. Pycha S, Trenti E, Mian C, et al. Diagnostic value of Xpert® BC Detection, Bladder Epicheck®, Urovysion® FISH and cytology in the detection of upper urinary tract urothelial carcinoma. World J Urol 2023;41:1323-8. [Crossref] [PubMed]