Abstract
OBJECTIVES: This study aims to develop and clinically evaluate EarlyTect BCD Plus, a urine-based assay measuring two methylation sites of the PENK gene, and to assess its diagnostic performance and clinical utility according to haematuria risk stratification. MATERIALS AND METHODS: A dual-target quantitative methylation-specific PCR assay was optimized using the PENK gene and evaluated in 892 patients with haematuria from Korea and the United States who underwent cystoscopy and histopathology. Urine DNA was analysed for two PENK methylation markers, and test results were interpreted using a combined algorithm. Diagnostic accuracy was assessed, and clinical utility was further analysed for patients stratified by the 2025 AUA/SUFU haematuria guideline risk categories. RESULTS: In the pooled cohort (gross haematuria, n = 509; microhaematuria, n = 366; unspecified haematuria, n = 17), EarlyTect BCD Plus achieved a sensitivity of 87.7% (95% CI, 81.5%-92.5%), specificity of 82.5% (95% CI, 79.6%-85.2%) and negative predictive value (NPV) of 97.0% (95% CI, 95.5%-98.0%). Sensitivity for Ta-LG tumours improved to 60.5% compared with the original single-marker assay, while high-grade tumours were detected with 96.6% sensitivity. In the intermediate-risk group, NPVs were 99.1% for all BC and 100% for high-grade BC. CONCLUSIONS: EarlyTect BCD Plus significantly enhances detection of Ta-LG bladder cancer while maintaining high specificity. Its high NPV supports use as a non-invasive adjunct and triage tool, allowing deferral of cystoscopy in selected haematuria patients.