Development of a founder-based CMV vaccine to interrupt Ebola virus transmission

描述（由申请人提供）：埃博拉病毒（EBOV）是线状病毒家族的阴性单链RNA病毒，被列为NIAID a类制剂。该项目的长期目标是开发基于巨细胞病毒（CMV）的疫苗，以干扰野生类人猿种群中的EBOV传播。近年来，西非人间暴发的埃博拉病毒相关严重出血热不断增加，导致高发病率和死亡率，死亡率接近90%1。人类的自然宿主和传播方式尚未确定，但蝙蝠和类人猿（大猩猩和黑猩猩）似乎在传播中发挥了作用。在啮齿类动物和非人灵长类动物（NHP）模型中，没有一种候选疫苗能够有效地从最初的疫苗接受者传播到人群的其他成员1,7-16。因此，人口免疫要求大多数人接种疫苗。这种无法传播的情况给针对西非类人猿人群的EBOV疫苗带来了困难。使用基于cmv的载体表达EBOV抗原的疫苗可能是这种难以接近的野生种群的理想疫苗。巨细胞病毒具有显著的再感染和在人群中传播的能力，无论先前是否有巨细胞病毒免疫17-19。基于cmv的疫苗也具有高度的免疫原性17,20-24。在我们的战略中，接种“创始人”疫苗最初会在人群中传播疫苗，从而消除对所有个体进行免疫的需要。巨细胞病毒也垂直传播，因此基于巨细胞病毒的疫苗可以提供永久的解决方案，以防止EBOV的传播。巨细胞病毒很可能在野生大猩猩和黑猩猩中普遍存在25-27。基于巨细胞病毒的疫苗仅代表感染了一种良性的、普遍存在的病毒，区别仅在于ebov衍生抗原的表达。EBOV作为五种抗原性不同的物种存在。我们建议构建基于cmv的载体，分别表达EBOV糖蛋白（GP）（体液免疫反应的主要但高度可变的靶标）和EBOV核蛋白（NP）（一种含有多个T细胞表位的高度保守的蛋白）。同时靶向GP和NP可诱导异源（跨ebov种）和同源保护。这项初步研究旨在确定以创始人为基础的EBOV疫苗的原则，非常适合R21资助。在小鼠中建立MCMV的“原理验证”将支持R01应用，以评估在NHP模型中表达GP和np的基于恒河猴cmv的载体所提供的保护，其中也可以评估交叉保护水平。特异性目的1：将构建和表征体外分别表达EBOV GP或NP（分别指定为MCMV/EBOV-GP和-NP）的两种小鼠巨细胞病毒（MCMV）载体构建。特异性目标2：确定MCMV/EBOV载体在小鼠体内诱导和维持EBOV GP和np特异性体液和T细胞反应的能力。具体目标3：将在小鼠模型中确定MCMV/EBOV载体抵御小鼠适应性EBOV攻击的能力。

DESCRIPTION (provided by applicant): Ebola virus (EBOV) is a negative single-stranded RNA virus of the filovirus family that is listed as a NIAID Category A agent. The long-term aim of this project is to develop cytomegalovirus (CMV)-based vaccines to interfere with EBOV transmission in wild Great Ape populations. Human outbreaks of severe EBOV-associated hemorrhagic fever in Western Africa have been increasing in recent years, resulting in high morbidity and mortality with a death rate approaching 90%1. The natural reservoir and mode of transmission to humans is not defined, however bats and Great Apes (gorillas and chimpanzees) appear to play a role in transmission. None of the vaccine candidates that have been shown to protect against EBOV in rodent and non-human primate (NHP) models have the ability to spread efficiently from initial vaccine recipients to other members of the population1,7-16. Thus, population immunization requires that most individuals are vaccinated. This inability to disseminate poses difficulties when targeting EBOV vaccines to inaccessible Western African Great Ape populations. A vaccine using CMV-based vectors expressing EBOV antigens may be ideal for such inaccessible wild populations. CMV possesses the remarkable ability to re-infect and disseminate through the population regardless of prior CMV immunity17-19. CMV-based vaccines are also highly immunogenic17,20-24. In our strategy, vaccination of 'founders' would initially spread the vaccine through the population, eliminating the need for immunization of all individuals. CMV is also transmitted vertically, thus a CMV-based vaccine could afford a permanent solution to preventing EBOV transmission. CMV is most likely ubiquitous in wild gorillas and chimpanzees25-27. A CMV-based vaccine merely represents infection with a benign, ubiquitous virus, differing only by expression of EBOV-derived antigens. EBOV exists as five antigenically distinct species. We propose to construct CMV-based vectors individually expressing EBOV glycoprotein (GP) (a major, but highly variable target of the humoral immune response) and EBOV nucleoprotein (NP) (a more highly conserved protein containing multiple T cell epitopes). Targeting both GP and NP may induce heterologous (cross-EBOV species) as well as homologous protection. This initial study to establish the principle of a founder-based vaccine for EBOV is ideally suited to R21 funding. Establishment of 'proof-of-principle' with MCMV in mice will support an R01 application to assess protection afforded by GP and NP-expressing rhesus CMV-based vectors in the NHP model, wherein levels of cross-protection can also be assessed. Specific Aim 1: will construct and characterize in vitro two murine CMV (MCMV) vector constructs individually expressing either EBOV GP or NP (designated MCMV/EBOV-GP and -NP, respectively). Specific Aim 2: will determine the ability of MCMV/EBOV vectors to induce and maintain an EBOV GP and NP-specific humoral and T cell response in mice. Specific Aim 3: will determine the capacity of MCMV/EBOV vectors to protect against mouse-adapted EBOV challenge in a mouse model. PUBLIC HEALTH RELEVANCE: Ebola virus (EBOV) is a NIAID Category A agent that causes outbreaks of lethal haemorrhagic fever in endemic areas of Western Africa. The frequency of outbreaks has been on the rise, which combined with the potential for accidental and deliberate release into non-endemic nations, makes EBOV a major health concern. The long-term aim of this project is to develop cytomegalovirus (CMV)-based vaccines to interfere with EBOV transmission in wild Great Ape populations, and thereby block this infectious conduit for EBOV transmission to Man.