VACCINE DEVELOPMENT FOR ALPHAVIRUSES

这个子项目是利用资源的许多研究子项目之一 由NIH/NCRR资助的中心拨款提供。子项目的主要支持 而子项目的主要调查员可能是由其他来源提供的， 包括其它NIH来源。 为子项目列出的总成本可能 代表子项目使用的中心基础设施的估计数量， 而不是由NCRR赠款提供给子项目或子项目工作人员的直接资金。 基孔肯雅热（CHIK）是由基孔肯雅病毒（一种致病性甲病毒）引起的媒介传播病毒性疾病。 最初，我们在实验性非人灵长类动物模型中重现了病毒性疾病（M。fascicularis）（n=3）。 这种发病率模型模拟了临床综合征，并显示了人类感染中注意到的自然疾病的特征。 建立该模型是为了提供一个先进的监测系统，用于测试针对CHIK开发的候选疫苗的效力。 正在开发的CHIK疫苗是基于含有小核糖核酸病毒内部核糖体进入位点（IRES）的构建体，以取代用于表达CHIK病毒（CHIKV）结构蛋白的亚基因组启动子。这种方法高度减毒病毒，防止蚊子感染，但保留免疫原性。 使用CHIK非人灵长类动物模型在单独的实验中测试两种版本的疫苗（CHIK/IRESv 1，v2）。 在动物体内植入远程无线电遥测技术，用于连续监测临床体征（核心体温、呼吸频率、心率、ECG），并在接种疫苗后和攻毒后采集样本进行病毒血症、抗体和病毒血症测定。 每组4只动物（N=16）通过皮下（SQ; CHIK/IRESv 1）或皮内（ID; CHIK/IRESv 2）途径以5.5^log单次剂量进行初免，或用生理盐水进行假疫苗接种。 接种后约45天，用6.0log野生型CHIKV SQ攻击所有动物。 CHIK/IRESv 2疫苗组产生的中和抗体滴度最高，CHIK/IRESv 1（ID）和CHIK/IRESv 1（SQ）组的平均滴度略低。 接种后观察到很少或没有病毒瘤，通过植入式遥测测量的接种后临床应答不显著。 在用WT CHIKV攻毒后，直至感染后+45天，没有接种疫苗的动物发生病毒血症、核心温度升高或指示CHIK疾病的任何其他临床标志。 相比之下，假疫苗接种的对照急性地（PI +1 - 3d）发展病毒血症，并且显示核心温度、心率和心电图测量的显著变化。 CHIK/IRES候选疫苗是安全的、高度免疫原性的，并且保护非人灵长类动物免受使用WT CHIKV的稳健实验攻击模型的攻击。 可探索疫苗剂量和接种途径的优化，以评价与该疫苗产品相关的长期免疫力。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Chikungunya (CHIK) is a vectorborne viral disease caused by chikungunya virus, a pathogenic alphavirus. Initially we recapitulated the viral disease in an experimental nonhuman primate model (M. fascicularis) (n=3). This morbidity model emulated the clinical syndrome and showed the hallmarks of natural disease noted in human infections. The model was established to provide an advanced monitoring system for testing the efficacy of candidate vaccines developed against CHIK. The CHIK vaccines under development are based upon constructs containing a picornavirus internal ribosomal entry site (IRES) to replace the subgenomic promoter for expression of the CHIK virus (CHIKV) structural proteins. This approach highly attenuates the virus and prevents mosquito infection, but preserves immunogenicity. The CHIK nonhuman primate model was used to test two versions of the vaccine (CHIK/IRESv1, v2) in separate experiments. Animals were implanted with remote radiotelemetry for continuous monitoring of clinical signs (core temperature, respiratory rate, heart rate, ECG) and samples were taken for viremia, antibody, and viremia assays following vaccination and after challenge. Groups of four animals (N=16) prime vaccinated with a single dose of 5.5^log by either the subcutaneous (SQ; CHIK/IRESv1) or intradermal (ID; CHIK/IRESv2) route, or sham vaccinated with saline. Approximately 45 days postvaccination, all animals were challenged SQ with 6.0log of wild-type CHIKV. The highest neutralizing antibody titers were generated by the CHIK/IRESv2 vaccine group, with slightly lower mean titers by CHIK/IRESv1 (ID) and CHIK/IRESv1 (SQ) groups. Little or no virema was noted after vaccination, and clinical response after vaccination as measured by implantable telemetry was unremarkable. Upon challenge with WT CHIKV, no vaccinated animals developed viremia, elevation in core temperature, or any other clinical hallmarks indicative of CHIK disease up to +45 days postinfection. In contrast, sham-vaccinated controls developed viremia acutely (+1-3d PI) and showed dramatic changes in core temperature, heart rate, and electrocardiogram measurements. The CHIK/IRES vaccine candidates are safe, highly immunogenic, and protect nonhuman primates from a robust experimental challenge model using WT CHIKV. Refinements in vaccine dose, as well as route of vaccination may be explored to evaluate the long term immunity associated with this vaccine product.