Validation of a Circulating Tumor DNA Blood Test in Muscle-Invasive Bladder and Upper Tract Urothelial Cancer

Overview

This prospective observational study evaluates a blood-based circulating tumor DNA (ctDNA) test in patients with muscle-invasive bladder cancer or upper tract urothelial cancer. ctDNA consists of small fragments of tumor DNA that can sometimes be detected in the blood. In this study, DNA from tumor tissue obtained during routine diagnostic or surgical procedures will be analyzed to identify tumor-specific mutations. Blood samples collected during routine clinical care will then be tested to determine whether the same mutations can be detected in plasma ctDNA. The main purpose is to assess how well the internally developed ctDNA assay detects tumor-specific mutations compared with tumor tissue sequencing. The study will also explore whether ctDNA results after treatment are associated with recurrence or remission during two years of follow-up, and whether changes in ctDNA levels during treatment are associated with pathological response. This study does not assign participants to a treatment and does not require additional blood draws beyond routine care. Treating physicians will remain blinded to ctDNA results during the study, so the test results will not influence clinical decisions.

Muscle-invasive bladder cancer and muscle-invasive upper tract urothelial carcinoma are associated with a relevant risk of recurrence or progression after treatment. Current follow-up mainly relies on imaging, pathology, and clinical assessment. Circulating tumor DNA (ctDNA) may provide additional information on tumor burden, minimal residual disease, and treatment response, but clinical implementation remains limited by cost, logistics, and the need for local assay validation. This study evaluates an internally developed, tumor-informed ctDNA workflow in patients treated at AZ Sint-Lucas Gent. Tumor DNA will be extracted from available formalin-fixed paraffin-embedded tumor tissue, such as tissue obtained during transurethral resection of bladder tumor, upper tract biopsy, nephro-ureterectomy, cystectomy, or biopsy of metastatic disease. Targeted next-generation sequencing will be used to identify patient-specific somatic mutations in the tumor tissue. When tumor-specific mutations are identified, matched plasma samples collected during routine clinical care will be analyzed for the same mutations. Cell-free DNA will be extracted from plasma and ctDNA detection will be performed using a tumor-informed approach, primarily by digital droplet PCR for known tumor-specific mutations. Mutations that cannot be assessed by digital droplet PCR may be further evaluated using next-generation sequencing. Blood samples will be obtained only as part of routine diagnostic, staging, treatment, or follow-up blood draws. The study does not assign participants to a specific treatment and does not require additional study-specific therapeutic interventions. Patients will receive standard clinical care according to institutional practice. Treating physicians will remain blinded to the ctDNA results during the study. The ctDNA results are investigational and will not guide treatment decisions during the study period. Clinical, pathological, treatment, molecular, and follow-up data will be collected to assess the performance of the assay and to explore associations between ctDNA status, treatment response, recurrence, progression, and remission over two years of follow-up. The study is intended to support internal validation of a local ctDNA assay and to determine whether this approach may be feasible for future integration into clinical workflows for urothelial cancer monitoring.