PA22-176, SBIR, Phase I, Development of a SARS-CoV-2 emerging variant infectivity and immune evasion panel to quantify the efficacy of vaccine booster induced neutralizing antibodies.

总结 随着COVID 19大流行和疫苗部署的继续，迫切需要监测新出现的 变体的感染性和逃避先前感染和疫苗诱导的中和抗体的能力。 目前，还没有办法快速量化新出现的变体的感染性和免疫逃避能力。的 定量病毒感染性的标准包括使用收获的病毒分离物并进行定量 噬菌斑测定，这是繁琐和耗时的。此外，使用传染性SARS-CoV-2需要 BSL-3设施中的高水平密封限制了其在普通临床和研究实验室中的应用。 假病毒已被广泛用于模拟SARS-CoV-2感染性和抗体中和。 假病毒，如从慢病毒和水泡性口炎病毒衍生的假病毒，可以模仿进入过程 SARS-CoV-2然而，这些假病毒体主要由非冠状病毒结构蛋白组成，并且需要 2-3几天就能产生结果。最近，一种新的杂合甲病毒-SARS-CoV-2假病毒粒子（HA-CoV-2）已被发现， 是私家侦探发明的（Hetrick）用于快速（3-6小时）和准确定量病毒感染性， 对中和抗体的敏感性 HA-CoV-2颗粒是非复制型SARS-CoV-2病毒样颗粒 (VLP)仅由SARS-CoV-2结构蛋白（S、M、N和E）和来自SARS-CoV-2的RNA报告基因组组成， 来自快速表达甲病毒载体。我们的初步研究表明，HA-CoV-2检测 可以快速量化接种疫苗和既往感染过的个体的中和抗体差异 特异的SARS-CoV-2变异体。维龙吉最近从乔治梅森那里获得了HA-CoV-2技术的许可 并计划开发商业试剂盒，其中包含代表SARS的HA-CoV-2颗粒组- CoV-2变异体目前在流通中具有最大的传染性和免疫逃逸的潜力。为病毒性 变种继续出现，迫切需要监测传染性并评估 新出现的变种的疫苗。HA-CoV-2系统将提供一个强大的平台， 病毒变体感染性和中和抗体应答的定量。我们建议发展一个医管局- 基于CoV-2假病毒的面板，用于检查感染性和中和抗体应答，具有两个特定目的。 具体目标1是筛选50种最流行的SARS-CoV-2变异体刺突蛋白， 使用HA-CoV-2假病毒平台，新出现的具有最高感染性的变体。具体目标二是 对循环中的每种COVID变体进行抗体中和试验，并确定 每种变体对疫苗和感染诱导的抗血清和恢复期血浆都有效。在这项研究中，WHO 抗SARS-CoV-2抗体的国际标准品和参比品将沿着进行筛选， 标准重组RBD抗体。在初步筛选后，将选择前5个变体进行开发 用于疫苗和加强剂开发人员评估其药物效力的商业面板试剂盒。一部分 在这个筛选过程中，维龙吉计划与约翰霍普金斯大学和其他非营利公共机构合作， 卫生组织提供本研究中获得的所有数据。HA-CoV-2假病毒目前 在Virongy公司网站（Virongy.com）上销售，仅供研究使用。

Summary As the COVID19 pandemic and vaccine deployment continue there is an urgent need to monitor emerging variants for their infectivity and ability to escape both prior infection and vaccine induced neutralizing antibodies. Currently, there is no way to rapidly quantify emerging variants infectivity and immune evading capacity. The standard for quantifying viral infectivity involves the use of harvested viral isolates and performing quantitative plaque assays, which are tedious and time consuming. Additionally, the use of infectious SARS-CoV-2 requires high-level containment in BSL-3 facilities that limits its application in common clinical and research laboratories. Pseudoviruses have been widely used to model SARS-CoV-2 infectivity and antibody neutralization. Pseudoviruses, such as those derived from lentivirus and vesicular stomatitis virus, can mimic the entry process of SARS-CoV-2. However, these pseudovirions consist of mostly non-coronavirus structural proteins and require 2-3 days to generate results. Recently, a novel hybrid alphavirus-SARS-CoV-2 pseudovirion (HA-CoV-2) has been developed by the P.I. (Hetrick) for rapid (3-6 hours) and accurate quantification of viral infectivity and sensitivity to neutralizing antibodies. The HA-CoV-2 particle is a non-replicating SARS-CoV-2 virus-like particle (VLP) composed of only SARS-CoV-2 structural proteins (S, M, N, and E) and a RNA reporter genome derived from a fast expressing alphavirus vector. Our preliminary studies have demonstrated that the HA-CoV-2 assay can rapidly quantify the differences in neutralizing antibodies from vaccinated and previously infected individuals to specific SARS-CoV-2 variants. Virongy has recently licensed the HA-CoV-2 technology from George Mason University, and plans to develop commercial kits with panels of HA-CoV-2 particles that represent the SARS- CoV-2 variants currently in circulation with the greatest infectivity and potential for immune escape. As viral variants continue to emerge there is an urgent need to monitor the infectivity and evaluate the effectiveness of the vaccines on emerging variants. The HA-CoV-2 system would provide a robust platform for rapid and accurate quantification of viral variant infectivity and the neutralizing antibody response. We propose to develop a HA- CoV-2 pseudovirus-based panel to examine infectivity and neutralizing antibody response with two specific aims. Specific Aim 1 is to screen the 50 most prevalent SARS-CoV-2 variant spike proteins in circulation and identify the emerging variants with the highest infectivity using the HA-CoV-2 pseudovirus platform. Specific aim 2 is to conduct antibody neutralization assays of each COVID variant in circulation and determine the sensitivity of each variant to both vaccine and infection induced anti-serum and convalescent plasma. For this study the WHO International Standard and Reference Panel for anti-SARS-CoV-2 antibody will be screened along with a standard recombinant RBD antibody. Following the initial screening the top 5 variants will be selected to develop a commercial panel kit for vaccine and booster developers to evaluate the efficacy of their prophylactics. As part of this screening process Virongy plans to collaborate with John Hopkins university and other non-profit public health organizations to provide all the data acquired in this study. The HA-CoV-2 pseudovirus are currently marketed on Virongy’s company website (virongy.com) for research use only.