A novel live-attenuated Zika vaccine with a modified 5'UTR

摘要 寨卡病毒(ZIKV)是一种由蚊子传播的黄病毒，已导致毁灭性的先天性寨卡综合征 (CZS)，包括小头畸形、先天畸形和新生儿死亡 流行病。此外，ZIKV感染可导致格林-巴利综合征(GBS)和脑膜脑炎 成年人。目前，ZIKV的发病机制尚不完全清楚，也没有得到批准的治疗或 有疫苗可用。尽管自2017年以来人类感染寨卡病毒的人数有所下降，但ZIKV很可能会成为 地方性疾病，可以预防CZS和成人寨卡病毒疾病的疫苗仍然迫切需要准备 未来的疫情。我们的研究小组最近通过以下方式开发了一种新的活体减毒ZIKV株(命名为Z7) 在流行前ZIKV的5‘非翻译区插入50-RNA-核苷酸(NT)发夹 柬埔寨菌株FSS13025，在神经毒力、免疫拮抗和蚊子方面减弱 与美国流行分离株的传染性比较。我们的初步数据表明，Z7可以复制 高效地在Vero细胞中产生高滴度，而不会引起明显的细胞病变(CPE)或失去 甚至在连续十段之后插入序列。有趣的是，我们在NS2B中发现了一个突变S1417A 第7代后Z7中的蛋白质，表明Z7可能在传代过程中适应了一种适应度突变。 重要的是，Z7诱导杀菌免疫，完全预防高剂量攻击后的病毒血症。 一种美国流行毒株(PRVABC59)在小鼠身上。这些结果表明，ZIKV 5‘非编码区的修饰 一种新的策略来开发ZIKV和潜在的其他黄病毒的减毒活疫苗候选。在……里面 在这一应用中，我们将首先在小鼠模型中表征Z7诱导的体液免疫和细胞免疫 (靶标1)，然后评价Z7对小鼠的安全性(靶标2)。在具体目标3中，我们将 确定Z7减毒的作用机制。我们认为这个项目意义重大，因为 它将表征新型减毒ZIKV候选疫苗Z7的免疫力和安全性，该疫苗曾被 首次将发夹环引入病毒5‘非编码区。此外，这个R15项目还将 为大学本科生和研究生提供宝贵和独特的培训机会 位于密西西比州南部。

Summary Zika virus (ZIKV) is a mosquito-transmitted flavivirus that has caused devastating congenital Zika syndromes (CZS), including microcephaly, congenital malformation, and fetal demise in human newborns in the 2015 - 2016 epidemics. In addition, ZIKV infection can cause Guillain-Barré syndrome (GBS) and meningoencephalitis in adults. Currently, ZIKV pathogenesis is incompletely understood, and there is no approved therapeutic or vaccine available. Although the number of human Zika cases has dropped since 2017, ZIKV will likely become endemic, and a vaccine that can prevent CZS and adult Zika diseases remains urgently needed to prepare for future outbreaks. Our research group recently developed a new live-attenuated ZIKV strain (named Z7) by inserting a 50-RNA-nucleotides (nt) hairpin into the 5’ untranslated region (UTR) of a pre-epidemic ZIKV Cambodian strain, FSS13025, which is attenuated in neurovirulence, immune antagonism, and mosquito infectivity compared with the American epidemic isolates. Our preliminary data demonstrate that Z7 replicates efficiently and produces high titers in Vero cells without causing apparent cytopathic effects (CPE) or losing the insert sequence even after ten consecutive passages. Interestingly, we identified a mutation S1417A in NS2B protein in Z7 after the 7th passage, indicating Z7 may have adapted a fitness mutation through the passaging. Importantly, Z7 induces sterilizing immunity that completely prevents viremia after a challenge with a high dose of an American epidemic strain (PRVABC59) in mice. These results suggest that modification of ZIKV 5’UTR is a novel strategy to develop live-attenuated vaccine candidates for ZIKV and potentially for other flaviviruses. In this application, we will first characterize Z7-induced humoral and cell-mediated immunity in a mouse model (Specific Aim 1), and then evaluate the safety features of Z7 in mice (Specific Aim 2). In Specific Aim 3, we will determine the mechanisms of action of Z7 attenuation. We believe that this project is highly significant because it will characterize the immunity and safety of a novel live-attenuated ZIKV vaccine candidate, Z7, which was developed for the first time by introducing a hairpin loop into a viral 5’ UTR. In addition, this R15 project will also provide a valuable and unique training opportunity to both undergraduate and graduate students at the University of Southern Mississippi.