Monitoring Recurrent Bladder Cancer with Electro-Phage Biosensors

项目摘要 这个项目的首要目标是用一种独特的方法检测复发的膀胱癌 能够定量检测患者尿液中肿瘤特异性生物标志物的技术平台。如果 该测试平台的成功，将改变目前监测复发的范式 膀胱癌。膀胱癌治疗后，患者面临高达60%的复发 通常每隔几个月通过细胞学检查和单一生物标志物进行监测 使用昂贵的中央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，患者的依从性是一个重要的问题。 我们的目标是将PI和Co-I实验室的一项新技术转化为临床 它将为患者提供快速、接近患者、敏感、特异、非侵入性和廉价的检测 复发性膀胱癌的检测。 Weiss和Penner实验室最近描述了电噬菌体生物传感器 使用定制病毒(噬菌体或噬菌体)作为生物标记亲和试剂。这个 生物传感器可以结合和测量合成尿液和尿液中癌症生物标志物的浓度 匿名捐赠者的尿样。我们把病毒和电子病毒结合在一起 导电聚合物，允许对坚固的 尿液中前列腺癌生物标志物亚硝胺浓度的检测和定量。 PM检测灵敏度符合临床检测要求。这种敏感性是 通过将基因编码和化学合成的噬菌体包裹在一起来实现 可能的癌症生物标记物的结合剂。 拟议的项目在先前研究的基础上扩展到包括多种癌症生物标记物 并确定复发膀胱癌的分子“指纹”。我们假设一个多- 分析方法将为监测膀胱复发提供更好的敏感性和特异性 癌症。靶向癌症生物标记物将被过度表达、纯化和重新折叠；然后 噬菌体展示将被用来鉴定肿瘤特异性结合蛋白。同时，实验将 优化噬菌体电极的结构以最大限度地提高信噪比，并提高 多分析物分析的敏感性和特异性。这样的优化将用于指导 开发一种多通道传感器，由工业合作伙伴PhageTech制造。 临床试验将分两个阶段进行，重点是分析和临床 电噬菌体生物传感器检测膀胱癌复发的有效性。在……里面 第一阶段，将分析10-20名患者的尿样，以比较敏感性和 使用FDA批准的两个生物标记物的测试进行量化；此外，阶段1将收集 有足够的数据指导第二阶段临床试验的设计。在这项规模更大的试点临床试验中， 近200例各病理分期、组织学分级的膀胱癌患者 将被登记；将从患有其他疾病的患者那里获得同等数量的对照样本 生殖器-尿路恶性肿瘤和健康患者。 因此，这项研究将比较目前对复发膀胱癌的监测方式。 利用新的生物传感器阵列方法，基于尿液的独特分子指纹诊断 治疗膀胱癌复发。这一护理要点，易于执行，不需要标签和试剂 传感方法将允许非侵入性、成本更低、更频繁的监控和 因此更早发现复发的膀胱癌。

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.