Designing mammarenavirus live vaccines with unbreachable attenuation

项目摘要/摘要 乳头瘤病毒拉萨(LASV)是西非特有的病毒，在那里它感染了数十万人 每年个人导致拉沙热(LF)病例数量较多，与高发病率和 死亡率。目前还没有美国FDA批准的LASV疫苗，目前的抗LASV治疗仅限于标签外 使用疗效有限的利巴韦林。LF已被列入世界卫生组织修订的优先疾病清单 研发蓝图，强调迫切需要疫苗来对抗LF。流行病学研究表明， 减毒活疫苗(LAV)是目前控制LF最可行的方法。哺乳动物病毒是 具有双片段负链RNA基因组的包膜病毒。每个基因组片段包含两个开放的 阅读由非编码基因间隔区(IGR)分隔的帧。大片段(L)编码核糖核酸 依赖的核糖核酸聚合酶、L蛋白和Z基质蛋白，而小片段(S)编码 表面糖蛋白前体(GPC)和核蛋白(NP)。我们已经记录了重组(R)形式 原型性淋巴细胞性脉络膜脑膜炎病毒(LCMV)表达密码子去优化 (CD)S段和L段均含有S段IGR的GPC或rLCMV/IGR(S-S)是稳定的 在小鼠体内完全减毒，但能够在一次给药后提供完全保护，防止 随后WT LCMV的致命挑战。重要的是，我们已经在LASV中复制了这些发现，因此 验证LCMV的使用，以生成概念验证结果，以指导LASV研究。的中心目标是 这个应用是为了检验这样一个假设，即rLCMV结合了CD GPC和S-IGR在 S和L的基因组片段，以下简称rLCMV/IGR-CD，都将具有良好的安全性和保护性 功能，如LAV，和牢不可破的衰减。为了检验我们的假设，我们将评估安全性， RLCMV/IGR-CD(Aim 1)的免疫原性和保护性，并检测rLCMV/IGR-CD 在联合感染的小鼠中，防止产生毒力增加的LCMV重组体(目标2)，以及 评估rLCMV/IGR-CD在FDA批准生产的人多轮感染期间的稳定性 Vero E6细胞和体内疫苗(目标3)。

Project Summary/Abstract The mammarenavirus Lassa (LASV) is endemic to West Africa where it infects several hundred thousand individuals yearly resulting in a high number of Lassa fever (LF) cases associated with high morbidity and mortality. There are no US FDA-approved LASV vaccines and current anti-LASV therapy is limited to an off-label use of ribavirin that has limited efficacy. LF has been included on the revised list of priority diseases for the WHO R&D Blueprint, underscoring an urgent need for vaccines to combat LF. Epidemiological studies indicate that a live-attenuated vaccine (LAV) represents the most feasible approach to control LF. Mammarenaviruses are enveloped viruses with a bi-segmented negative strand RNA genome. Each genome segment contains two open reading frames separated by a non-coding intergenic region (IGR). The large (L) segment encodes the RNA dependent RNA polymerase, L protein, and the Z matrix protein, whereas the small (S) segment encodes the surface glycoprotein precursor (GPC) and nucleoprotein (NP). We have documented that recombinant (r) forms of the prototypic mammarenavirus lymphocytic choriomeningitis virus (LCMV) expressing a codon deoptimized (CD) GPC or containing the IGR of the S segment in both the S and L segments rLCMV/IGR(S-S) are stable and fully attenuated in mice but able to provide complete protection, upon a single administration, against a subsequent lethal challenge with WT LCMV. Importantly, we have replicated these findings with LASV, thus validating the use of LCMV to generate proof of concept results to guide studies with LASV. The central goal of this application is to test the hypothesis that an rLCMV combining the features of a CD GPC and the S-IGR in both S and L genome segments, hereinafter rLCMV/IGR-CD, will have excellent safety and protective efficacy features as LAV, and an unbreachable attenuation. To test our hypothesis, we will assess the safety, immunogenicity, and protective efficacy of rLCMV/IGR-CD (Aim 1) and examine whether rLCMV/IGR-CD prevents, in co-infected mice, the generation of LCMV reassortants with increased virulence (Aim 2), as well as evaluate rLCMV/IGR-CD stability during multiple rounds of infection in FDA approved for production of human vaccines Vero E6 cells and in vivo (Aim 3).