Bladder cancer is the sixth most common cancer in the U.S., has one of the highest recurrence rates of all solid cancers, and is the most expensive cancer to treat from diagnosis to death. Veterans are at increased risk for bladder cancer due higher rates of tobacco use and environmental exposure. There are significant unmet needs for biomarkers and molecular diagnostic tools to better inform decision making across all stages of bladder cancer. For patients with early stage disease, current diagnostic approaches are either invasive (e.g. cystoscopy) or lack sensitivity (e.g. urine cytology). We propose to develop molecular diagnostics that will lead to more effective and personalized therapeutic strategies for patients with localized bladder cancer. Tumor-derived nucleic acids in biological fluids such as urine represent promising biomarkers for non-invasive measurement of disease burden and response to therapy. Previously, we developed a pipeline for bladder cancer biomarker discovery, validation, and technology integration for clinical translation. We applied high throughput RNA sequencing of urinary sediments as a biomarker discovery tool and identified a urinary mRNA (u-mRNA) panel with promising diagnostic performance. In parallel, we validated an integrated microfluidics cartridge capable of “sample-in, answer-out” within 90 minutes using a separate u-mRNA panel. Our recent preliminary data indicate further improvement of diagnostic performance by combining the two panels within the integrated cartridge. Based on these promising preliminary data, we propose three specific aims: 1) to validate the optimized mRNA panel for bladder cancer surveillance and risk stratification; 2) to validate the optimized u- mRNA panel to improve bladder cancer screening and assess risk stratification; and 3) to validate the optimized u-mRNA panel for response assessment and monitoring in patients with high risk NMIBC treated with BCG immunotherapy. Our team is led by experienced academic VA investigators with complimentary expertise in molecular diagnostics (Liao), urologic oncology (Liao, Leppert), and clinical validation study design (Liao, Yu). Successful completion of the studies proposed here will serve as a foundation for incorporating urine-based biomarkers in bladder cancer surveillance and into prospective clinical trials to assess therapeutic interventions. We foresee that our approach will allow personalization of treatment strategies to improve outcomes for BC patients in both veterans and the general population.