Development and initial evaluation of a urine DNA methylation assay for pan-urinary cancer detection

Cancers affecting the urinary system, encompassing malignancies of the bladder, ureter, renal pelvis, kidney, and prostate, represent a major global health challenge. DNA methylation biomarkers for multi-cancer detection in the urinary tract have been evaluated in limited contexts. This study aimed to develop a non-invasive urine-based assay utilizing DNA methylation markers for the simultaneous detection of multiple urinary system cancers. In this retrospective case–control study of 609 participants, a four-gene urine DNA methylation model (H4C6, SIX6, SEPTIN9, and SHOX2) was developed in a training set (n = 425) and evaluated in a held-out split-sample internal validation set (n = 184). The pan-urinary cancer detection (PUCD) model achieved an area under the curve of 0.95 (95% confidence interval (CI) 0.913–0.987) in the validation set. Notably, it demonstrated high sensitivity for early-stage (Stage I/II) cancers (88.2%, 95% CI 78.5–93.9%) alongside the caveat that performance varies by cancer type. The assay provided discrimination between cancer and non-cancer samples even within subgroups positive for urine occult blood or protein. These findings support further prospective evaluation of the model as a potential non-invasive adjunctive triage tool for patients with hematuria or other suspicious urinary symptoms.