Monitoring Recurrent Bladder Cancer with Electro-Phage Biosensors

项目摘要 该项目的总体目标是使用独特的方法检测复发性膀胱癌。 该技术平台能够定量患者尿液中的肿瘤特异性生物标志物。如果 如果成功，这一测试平台将改变目前监测经常性支出的模式， 膀胱癌膀胱癌治疗后，患者面临高达60%的复发率 率，并且通常每隔几个月通过细胞镜检和单一生物标志物监测一次 使用昂贵的中心实验室测试进行测量。不幸的是细胞学检查是侵入性的 并发症发生率高达15%因此，患者依从性是一个重要问题。 我们的目标是将PI和Co-I实验室的新技术转化为临床 它将提供快速，接近病人，敏感，特异，非侵入性和廉价的测试， 复发性膀胱癌的检测。 韦斯和彭纳实验室最近描述了电噬菌体生物传感器 其使用定制的病毒（噬菌体或噬菌体）作为生物标志物亲和试剂。的 生物传感器可以结合并测量合成尿中癌症生物标志物的浓度， 匿名捐赠者的尿样我们把病毒和电子 导电聚合物，它允许阻抗光谱测量的鲁棒 检测和定量尿中亚nM浓度的前列腺癌生物标志物。 pM检测试剂盒灵敏度在临床试验要求范围内。这种敏感性是 通过用基因编码和化学合成的 与推定的癌症生物标志物结合。 拟议的项目扩展了之前的研究，纳入多种癌症生物标志物 并确定复发性膀胱癌的分子“指纹”。我们假设一个多- 分析物方法将允许更好的灵敏度和特异性用于监测膀胱复发 癌靶向的癌症生物标志物将被过表达、纯化和重折叠;然后 噬菌体展示将用于鉴定肿瘤特异性结合物。与此同时，实验将 优化噬菌体电极的结构，以最大化信噪比，并提高 多分析物测定的灵敏度和特异性。这种优化将用于指导 开发由工业合作伙伴PhageTech制造的多通道传感器。 临床试验将分两个阶段进行，重点是分析和临床 用于检测膀胱癌复发的电噬菌体生物传感器的验证。在 第1阶段，将分析10-20名患者的尿样，以比较灵敏度和 使用FDA批准的两种生物标志物检测进行定量;此外，第1阶段将收集 有足够的数据来指导第2阶段临床试验的设计。在这个规模更大的临床试验中， 约200例所有病理分期和组织学分级的膀胱癌患者 将入组;将从其他患者中获得相同数量的对照样本。 泌尿生殖道恶性肿瘤和健康患者。 因此，该研究将比较目前复发性膀胱癌的监测方式 与新的生物传感器阵列的方法，一个独特的，尿为基础的，分子指纹诊断 治疗膀胱癌复发这种护理点，易于执行，无标签和试剂 传感方法将允许进行非侵入性、成本更低、更频繁的监测和 从而更早地发现复发性膀胱癌。

Project Summary The overarching goal of this project is to detect recurrent bladder cancer using a unique technology platform capable of quantitating tumor-specific biomarkers in patient urine. If successful, this testing platform will change the current paradigm for monitoring recurrent bladder cancer. Following treatment for bladder cancer, patients face up to a 60% recurrence rate, and are typically monitored every few months by cytoscopy and single biomarker measurements using expensive central laboratory testing. Unfortunately, cytoscopy is invasive, and has a complication rate as high as 15%. Patient compliance is therefore a significant issue. Our goal is to translate a new technology from the PI and Co-I's laboratories into the clinic where it will offer rapid, near patient, sensitive, specific, non-invasive and inexpensive testing for the detection of recurrent bladder cancer. The Weiss and Penner laboratories have recently described Electro-Phage biosensors that use customized viruses (bacteriophage or phage) as biomarker affinity reagents. The biosensor can bind and measure the concentrations of cancer biomarkers in synthetic urine and spiked urine samples from anonymous donors. We have combined viruses with an electrically conductive polymer, which allows impedance spectroscopic measurements for the robust detection and quantification of sub-nM concentrations of a prostate cancer biomarker in urine. The pM assay sensitivity is within the requirements for clinical testing. This sensitivity is achieved by coating the phage with both genetically encoded and chemically synthesized binders to the putative cancer biomarkers. The proposed project expands on previous studies to include multiple cancer biomarkers and identify a molecular “fingerprint” for recurrent bladder cancer. We hypothesize that a multi- analyte approach will allow better sensitivity and specificity for monitoring recurrence of bladder cancer. The targeted cancer biomarkers will be over-expressed, purified, and refolded; then phage display will be employed to identify tumor specific binders. In parallel, experiments will optimize the architecture of the Phage-Electrode to maximize signal-to-noise, and improve the sensitivity and specificity of the multi-analyte assay. Such optimization will be used to guide development of a multi-channel sensor, fabricated by the industrial partner PhageTech. The clinical trial will be conducted in two stages, and focus on analytical and clinical validation of the Electro-Phage biosensor for the detection of bladder cancer recurrence. In Stage 1, urine samples from 10-20 patients will be analyzed to compare sensitivity and quantification with FDA-approved tests for two biomarkers; in addition, Stage 1 will collect sufficient data to guide the design of the Stage 2 clinical trial. In this larger, pilot clinical trial, approximately 200 patients with bladder cancer of all pathologic stages and histologic grades will be enrolled; an equal number of control samples will be obtained from patients with other genito-urinary tract malignancies and healthy patients. Thus, the study will compare current monitoring modalities for recurrent bladder cancer with the novel biosensor array approach, a distinct, urine-based, molecular fingerprint diagnostic for bladder cancer recurrence. This point of care, easy to perform, label- and reagent-free sensing approach will allow for non-invasive, less costly, more frequent, monitoring and therefore earlier detection of recurrent bladder cancer.