Distinct longitudinal patterns of urine tumor DNA in patients undergoing surveillance for bladder cancer

Abstract

Cystoscopy is the gold standard for surveillance of non-muscle invasive bladder cancer (NMIBC), but the procedure is invasive and has suboptimal accuracy. The aim of this study was to investigate the potential of analyzing urine samples for the presence of urine tumor DNA (utDNA) to replace cystoscopy for surveillance of bladder cancer recurrence. In this longitudinal, prospective, and observational study, 47 patients were followed for recurrence for 2 years, involving analysis of utDNA using the BladMetrix DNA methylation biomarker test at each cystoscopy. In total, utDNA was detected in 21/23 recurrences (91% sensitivity), including 5/5 T1, T2, and carcinoma in situ (CIS) tumors (100%) and 10/12 Ta tumors (83%), with < 1% false-negative test results. Importantly, utDNA analysis showed the potential to reduce the number of cystoscopies by 55%, benefitting 79% of the patients. Eleven of 23 recurrences (48%) were detected earlier with utDNA than with cystoscopy, and distinct patterns of residual utDNA post-surgery indicated minimal residual disease (MRD) or field effect in 6% and 15% of the patients, respectively. In conclusion, utDNA analysis shows high sensitivity to detect tumor recurrence, potential to reduce the number of cystoscopies, and promise to guide patient-specific surveillance regimens.

Keywords: BladMetrix; DNA methylation; biomarker; field effect; minimal residual disease; recurrence.

Fig. 1

The overall performance of urine tumor DNA (utDNA) for surveillance of bladder cancer patients. Each line represents one patient (y‐axis), and each dot represents a longitudinal surveillance event (in months; x‐axis). The color of the dot indicates the performance of the utDNA analysis compared to the standard reference (i.e., histology if available, or else cystoscopy). Of the 261 surveillance controls utDNA correctly identified 21 recurrences (true positives; green dots) and misclassified 71 non‐recurrences as recurrences (false positives; orange dots). The utDNA analysis was correctly negative at 143 surveillance events with no recurrence (true negatives; blue dots), while two recurrences were missed by utDNA (false negatives; red dots). In 24 cases, utDNA analysis revealed early recurrence detections (green triangles). If treatment was administered to the patient during the study course, this is indicated to the left on the figure by a star (BCG), cross (mitomycin), or a diamond (adjuvant chemotherapy).

Fig. 2

Longitudinal urine tumor DNA (utDNA) analysis shown for five bladder cancer patients representing different clinical scenarios. (A) patient with recurrences, (B) patient without recurrence, (C) patient with early recurrence detection, (D) patient with MRD, (E) patient with field effect. The eight DNA methylation biomarkers (i.e., the BladMetrix test) used for the utDNA analysis are shown vertically in different colors with individual y‐axis values. The dotted lines indicate the biomarker‐specific methylation scoring threshold. A biomarker is scored positive if the DNA methylation level is above the threshold (closed circles) and negative if the DNA methylation level is below the threshold (open circles). The results of the utDNA analysis, cystoscopy, and histology for each surveillance control are shown below the chart. The numbers of methylated biomarkers are indicated inside the squares. The utDNA analysis is scored positive if ≥ 2/8 biomarkers are positive and negative if < 2/8 biomarkers are positive. MRD, minimal residual disease; NA, not available; R, recurrence; utDNA, urine tumor DNA.

Fig. 3

Urine tumor DNA (utDNA) identifies various clinical scenarios. Illustrative examples of utDNA results (positive: green plus sign, negative: red minus sign) at diagnosis, post‐surgery, during surveillance and recurrence in the setting of (A) regular recurrence detection, (B) early detection, (C) MRD, and (D) field effect. (A) utDNA correctly detects a recurrence, and the utDNA level drops below threshold post‐surgery. (B) A recurrence is detected with utDNA before it is detected by cystoscopy in the clinic. Post‐surgery, the level of utDNA drops below threshold. (C) Residual tumor cells are left after surgery, and the utDNA test is positive at all surveillance points until the recurrence is surgically removed. After surgery, with the successful removal of the entire recurrent tumor, the utDNA analysis is negative. (D) The tumor at diagnosis imposes cancer‐specific methylation changes to the surrounding normal‐appearing bladder tissue (indicated by pink tissue color), predisposing that tissue to undergo later malignant transformation. Consequently, the utDNA analysis is positive at all surveillance points, also after surgical removal of the recurrent tumor. MRD, minimal residual disease; utDNA, urine tumor DNA. Created with BioRender.