MP59-03: A urine-based mutation-methylation assay for the surveillance of high risk non-muscle invasive bladder cancer patients

Joep de Jong*, Florus de Jong, Angelique Ziel-van der Made, Ties Hagdorn, Egbert Boevé, Rotterdam, Netherlands, Hossain Roshani, The Hague, Netherlands, Eric Oomens, Breda, Netherlands, Rob Pelger, Ewout Steyerberg, Leiden, Netherlands, Joost Boormans, Chris Bangma, Tahlita Zuiverloon, Ellen Zwarthoff, Rotterdam, Netherlands

Introduction: High-risk non-muscle invasive bladder cancer (HR-NMIBC) patients require life-long follow-up, including frequent cystoscopy and urine cytology. Previously, we developed a non-invasive diagnostic urine assay for the follow-up of HR-NMIBC patients to detect methylation of OTX1 and mutations in FGFR3 and TERT genes. Importantly, this assay showed promising performance in the development setting, with a strong anticipatory effect. Here, we aimed to validate the diagnostic performance of the assay and evaluate its utility in an unselected prospectively collected cohort of HR-NMIBC patients under surveillance.

Methods: We prospectively included 204 HR-NMIBC patients from five centers. A total of 736 urine sample pairs (evening and morning urines) were collected prior to cystoscopy during regular visits and mutation/methylation status of the three genes was analysed on cellular DNA from voided urines. A test was considered positive if =1 alteration was detected in at least one urine sample. The primary end point was tumor recurrence. Cross-sectional analyses were performed to assess sensitivity and specificity. Gene marker scores were evaluated in generalised mixed effects models to adjust for within-patient correlation. Further longitudinal analyses assessed the anticipatory effect of the urine assay in a Cox proportional hazard model with time-dependent effects to calculate the hazard rate (HR) for developing a recurrence over time.

Results: In total, 63 recurrences were diagnosed for which we had concomitant assay results. On cross-sectional analyses, the assay detected 75% (95% CI 62.1%-84.7%) of recurrences, with a specificity of 70% (95% CI 66.4%-73.5%). The cross-sectional sensitivity of concomitant assays for detecting =T1 disease (histologically confirmed) was 100%, detecting 5 out of 5 histological =T1 recurrences. Furthermore, mixed effects model analyses revealed OTX1 (P=0.005) and TERT (P=0.004) as significant predictors for disease recurrence. With a median follow-up of 25.3 months (IQR 18.6 – 30.7), 29 additional tumors were diagnosed without available concomitant urine samples, including recurrences detected during follow-up after the urine collection period. On longitudinal analyses, a positive urine assay showed a significant (P <0.001) HR of 3.5 for predicting a recurrence over time. Furthermore, having a recurrence during the study period also appeared to be a significant (P <0.001) predictor for additional tumor recurrence in the future.

Conclusions: This study validates the performance of a previously developed urine assay in an unselected cohort of HR-NMIBC patients under surveillance. With a robust sensitivity/specificity, together with a strong anticipatory effect, this assay proves a useful adjunct ready for evaluation in a future randomized controlled trial.

Source of Funding: ZonMw Efficiency Grant