The Rigor and Clinical Utility of PSMA Enriched Extracellular Vesicles for Prostate Cancer Detection

前列腺癌筛查挽救了生命，但却导致绝大多数男性遭受 不必要的侵入性前列腺活检，以及过度诊断和治疗惰性癌症的风险。多个- 前列腺癌的参数磁共振成像(MpMRI)和分子生物标记物被用来评估临床风险。 重大前列腺癌(CsPCA)和需要活检，但它们在检测准确性上是有限的 CsPCa，导致许多男性仍然需要接受活组织检查，因为担心错过一个重要的肿瘤。这项建议 阐述提高cspca检测精确度以减轻前列腺负担的一个重要问题。 癌症筛查。由癌细胞释放到体液中的细胞外小泡(EV)很有希望 液体活组织检查的生物标志物，因为它们可以从血液和尿液中提取并携带分子成分 反映出亲代肿瘤。问题是选择性地提取前列腺癌释放的EV可能是 来自非前列腺源的EVS的挑战性和贡献性可能会降低检测率和特异性。在协作中 与Exosome Diagnostics的研发团队合作，他们是临床级领域的专家 EV生物标志物分析，我们开发了一种丰富表达前列腺特异性EV的方法 膜抗原(PSMA)表面蛋白。我们发现，PSMA EV捕获浓缩在 检测不同的、可能更具前列腺特异性的肠道病毒mRNAs和长的非编码RNA。 然而，尽管EV内容物被脂膜很好地保护，但收集和 表达PSMA表面蛋白的EVS的加工还没有严格的定义。在我们提议的 项目，我们将专注于开发一种对csPCA更具特异性的尿液PSMA EV检测方法。 目前可用于PCA的测试。具体地说，我们将检验PSMA获得的尿液EVS 浓缩可以提供一组EV RNA标志物，从而显著增加前列腺癌的风险 评估。在具体目标1中，我们制定了一项创新的多因素分析发展计划，以解决 先前对表面抗原捕获的严密性和重复性的限制并开发出最佳工作流程 用于PSMA浓缩。在具体目标2中，我们将对PSMA富集物进行RNAseq综合分析 以及在一项正在进行的迈阿密大学(MDSelect：NCT04240327)临床试验中招募250名患者的男性EV总数 接受活检以评估csPCA，以确定PSMA浓缩与总EVS的相加价值 用于csPCA检测。在具体目标3中，我们将开发和验证一种新的尿液EV特征，以增强 使用来自PSMA富集型和总电动汽车的RNAseq数据进行csPCA检测的准确性，以及多机构 正在进行的NCI EDRN(NCT03784924)临床试验中的验证。根据我们的初步数据和 结合我们研究团队的专业知识，我们可以很好地开发和提供基于EV的尿液 生物标志物分析，显著提高csPCA的检测能力，并在临床试验中得到验证。这项提议将 大幅增强csPCA检测，为未来基于EV的前列腺癌标志物奠定基础。

Screening for prostate cancer saves lives but results in an overwhelming number of men being subjected to unnecessary, invasive prostate biopsies, and the risk of over diagnosis and treatment of indolent cancer. Multi- parametric MRI (mpMRI) of the prostate and molecular biomarkers are used to evaluate the risk of clinically significant prostate cancer (csPCa) and need for biopsy, however they are limited in their accuracy for detecting csPCa, resulting in many men still needing to undergo biopsy for fear of missing a significant tumor. This proposal addresses an important issue in enhancing the precision of csPCa detection to reduce the burdens of prostate cancer screening. Extracellular Vesicles (EVs) that are released into body fluids by cancer cells are promising biomarkers for liquid biopsy since they can be extracted from blood and urine and carry molecular constituents reflecting the parent tumor. The problem is selectively extracting EVs released from prostate cancers can be challenging, and contribution by EVs of non-prostate origin can lessen detection and specificity. In collaboration with the research and development team at Exosome Diagnostics, who are experts in the field of clinical-grade EV biomarker analysis, we have developed a method to enrich for EVs expressing the Prostate Specific Membrane Antigen (PSMA) surface protein. We have found that PSMA EV capture enrichment results in the detection of a different, and potentially more prostate specific profile of EV mRNAs and long non-coding RNAs. However while EV contents are well protected by lipid membranes, optimal conditions for the collection and processing of EVs expressing PSMA surface protein have not yet been rigorously defined. In our proposed project, we will focus on development of a urine PSMA EV assay that is more specific for csPCa than the other currently available tests for PCa. Specifically, we will test the hypothesis that the urine EVs obtained by PSMA enrichment can provide a panel of EV RNA markers that can substantially enhance prostate cancer risk assessment. In Specific Aim 1, we have developed an innovative multi-factor assay development plan to address previous limitations in the rigor and reproducibility of surface antigen capture and develop an optimal workflow for PSMA enrichment. In Specific Aim 2, we will conduct RNAseq comprehensive profiling of PSMA enriched and total EVs from men in an ongoing U Miami (MDSelect: NCT04240327) clinical trial enrolling 250 patients undergoing biopsy for evaluation of csPCa to determine the additive value of PSMA enrichment over total EVs for csPCa detection. In Specific Aim 3, we will develop and validate a novel urinary EV signature to enhance the accuracy of csPCa detection using RNAseq data from PSMA enriched and total EVs, with multi-institutional validation in an ongoing NCI EDRN (NCT03784924) clinical trial. Based on our preliminary data and the combined expertise of our research team we are well positioned to develop and deliver an EV based urine biomarker assay that significantly enhances csPCa detection with validation in clinical trials. This proposal will substantially enhance csPCa detection and lay the foundation for future EV based prostate cancer markers.