A Multiplex Protein Biomarker-Based Immunoassay for the Early Detection of Bladder Cancer and its Implications in Tumor Biology

Over 83,000 Americans will be diagnosed with bladder cancer in 2021 with over 17,000 dying of the disease during this period. Unfortunately, both the absolute numbers of cases and deaths from bladder cancer have increased by 57 and 41%, respectively, since 2000. When detected early (i.e., NMIBC or stage 1), the 5-yr survival rate is >90%, compared to a significant reduction in survival if the disease is noted to be MIBC (stage 2; 50% 5-yr survival) or metastatic (stages 3 and 4; <20% 5-yr survival). Thus, the prevailing idea is that early detection of bladder cancer in high risk individuals (i.e., individuals exposed to certain carcinogens) will likely be the best modality to address advanced bladder cancer’s dismal outcomes. Currently, the evaluation of at risk individuals remains a challenge, and as such, there are no modalities available to effectively screen this high risk population. Previously, we have a) identified a bladder cancer-associated diagnostic “signature” comprised of 10 biomarkers, b) developed a multiplex immunoassay to query the “signature” in voided urine samples and c) performed analytical validation of the multiplex immunoassay. Using the multiplex immunoassay, we have generated encouraging preliminary data from a cohort of 362 subjects (46 cancers) (AUC 0.95; sensitivity 0.93, specificity 0.93, positive predictive value 0.65 and negative predictive value 0.99). Thus, for the first time, we possess a robust assay that can be used to non-invasively detect bladder cancer. Utilizing this assay in our ongoing prospective study surveilling patients with a history of bladder cancer, we have noted an elevation of our ‘signature’ as early as 18 months prior to the clinical diagnosis of cancer and an actual positive multiplex immunoassay in all cancer patients 12 months prior to the clinical diagnosis of cancer. Furthermore, we have evidence that 9 of our 10 biomarkers within the “signature” are expressed in relevant carcinogen induced mouse bladder cancer model. Hypothesis: A bladder cancer-associated signature exists that can be leveraged to indicate the presence of bladder cancer from a single voided urine sample months to years prior to the clinical presentation and diagnosis of bladder cancer. Specific Aims: 1) To perform a pilot study to evaluate the multiplex immunoassay’s ability to early detect bladder cancer and 2) To use a relevant carcinogen induced mouse bladder cancer model to identify early changes within the tumor microenvironment which could serve as biomarkers candidates for testing in human samples. Significance This research will open the door for improving on the non-invasive methods for the early detection of bladder cancer, and as such, it will have a marked impact on patient survival. Methodology We will conduct a prospective pilot study (n=150) to demonstrate the feasibility of identifying, following and testing high risk individuals for bladder cancer. Then utilizing a relevant carcinogen induced mouse bladder cancer model, we will study the spatial and temporal association of our “signature” and link it to key changes within the tumor microenvironment, identifying novel biomarkers for future clinical development. Expected Results There exists an unmet clinical need for reliable biomarkers to early detect bladder cancer when its more treatable with improved survival rates.