Development of a Novel, Sensitive ZIKV-Specific Sero-diagnostic Assay Utilizing Biologically Inspired Synthetic Molecules

项目摘要/摘要 寨卡病毒(ZIKV)是一种蚊媒黄病毒，与登革热病毒(DENV)有基因上的亲缘关系，它会产生温和的 发热性疾病伴有斑丘疹，大约80%的患者没有表现出症状。直到 自1947年发现ZIKV以来，只报告了14例人类疾病。2015年，一场疫情 据估计，仅巴西目前就有100多万人感染。而ZIKV感染是 通常轻微，最近在成人和儿童中与格林-巴利综合征有关，以及 新生儿小头畸形率高得惊人。目前还没有商业化的疫苗或诊断化验 可用。用抗ZIKV抗体测量的ZIKV免疫力的准确检测将是 预防和控制程序。目前的ZIKV血清诊断方法通常显示大量交叉 与其他黄病毒试剂的反应性。我们最近开发了一种革命性的方法来发现 通过筛选一个非常大的组合来诊断病毒感染(HIV和DENV)的新抗体配体 生物启发的但非生物的分子形状或类肽的文库(108个分子变体)， 对抗感染后患者血清的面板。使用这种方法，我们分离出了几个特定的类肽 感染后血清中存在的结合抗体，在正常非免疫血清中不存在，在感染前血清中也不存在。 感染血清。我们的概念验证初步研究已经证明了类肽生物标志物能够 HIV抗体阳性和阴性血清以及DEN抗体阳性血清的鉴别 和DEN抗体阴性血清。此外，我们还发现了能够 区分最近和非最近的艾滋病毒感染。我们使用了这些新的非生物特异性的 配体作为检测HIV的基础，表现出很好的敏感性和特异性。在这笔赠款中，我们 建议利用这项新技术开发强大的、基于多肽的检测方法来专门检测寨卡病毒 抗体。我们将用累西腓患者的血清面板测试这项创新技术， 巴西寨卡病毒爆发的震中。直接访问临床队列和患者样本将我们的 团队很好地进行了ZIKV诊断开发。在鉴定出ZIKV反应性类肽后，我们将进行 针对所选类肽的免疫球蛋白反应动力学的详细表征。这个 对这些类肽生物标记物的鉴定将导致第二代 将进一步优化类肽活性的组合文库，并将确定其作用机制 ZIKV特异性类肽生物标记物与患者抗体之间的结合。本文件中描述的目标 该项目将导致开发一种高度优化、低成本、基于类肽的高通量检测方法，以 准确检测寨卡病毒抗体，从而诊断个别患者并测量受影响患者的发病率 人口。这种诊断化验和疾病监测工具的发展将大大增加 全球公共卫生机构监测具有全球重要性的传染病的能力。

PROJECT SUMMARY/ABSTRACT Zika virus (ZIKV), a mosquito-borne flavivirus genetically related to dengue virus (DENV), produces a mild febrile illness with maculo-papular rash with approximately 80% of patients demonstrating no symptoms. Until 2007 only 14 cases of human disease had been reported since ZIKV's discovery in 1947. In 2015, an epidemic exploded with estimations of over one million current infections in Brazil alone. While ZIKV infections are generally mild, recently they have been associated with Guillain-Barré syndrome in adults and children and an alarmingly high rate of microcephaly in newborns. There are no commercial vaccines or diagnostic assays available. Accurate detection of ZIKV immunity, as measured by anti-ZIKV antibodies, will be a critical tool in prevention and control programs. Current ZIKV sero-diagnostic methods typically show substantial cross- reactivity with other flavivirus reagents. We have recently developed a revolutionary approach to discover novel diagnostic antibody ligands for viral infections (HIV and DENV) by screening a very large combinatorial library (108 molecular variants) of biologically inspired, but non-biological, molecular shapes, or peptoids, against panels of post-infection patient sera. Using this method, we isolated several peptoids that specifically bound antibodies present in post-infection sera that are not present in normal non-immune sera nor pre- infection sera. Our proof-of-concept preliminary studies have demonstrated the ability of peptoid biomarkers to differentiate between HIV antibody positive and HIV antibody negative serum as well as DEN antibody positive and DEN antibody negative serum. Additionally, we have also identified peptoid biomarkers that can distinguish between recent and non-recent HIV infections. We have used these novel non-biological specific ligands as the bases for HIV ELISA assays which display very good sensitivity and specificity. In this grant, we propose to harness this novel technology to develop robust, peptoid-based assays to specifically detect ZIKV antibodies. We will test this innovative technology against panels of sera, assembled from patients in Recife, the epicenter of the Zika outbreak in Brazil. Direct access to clinical cohorts and patient samples positions our team well for a ZIKV diagnostic development. After identification of ZIKV reactive peptoids, we will perform a detailed characterization of the kinetics of the immunoglobulin responses against the selected peptoids. The identification of these peptoid biomarkers will then lead to the development of a second-generation combinatorial library where peptoid reactivity will be further optimized, and we will identify the mechanisms of binding between the ZIKV-specific peptoid biomarkers and the patient antibody. The goals described in this project will lead to the development of a highly optimized, low cost, peptoid-based high-throughput assay to accurately detect ZIKV antibodies and thereby diagnose individual patients and measure incidence in affected populations. The development of such diagnostic assay and disease surveillance tools will greatly increase the capacity of public health agencies worldwide to monitor infectious diseases of global importance.