Development of an RSV Vaccine by Molecular Manipulation of the Viral Matrix Prot

人类呼吸道合胞病毒(HRSV)是导致儿童毛细支气管炎和 肺炎。据估计，全世界每年有10万儿童死亡，开发一种抗病毒药物 HRSV疫苗是当务之急。在目前的方法中，实时衰减是有吸引力的，因为不同于 灭活疫苗，它有望诱导广泛和平衡的免疫反应。然而，现场HRSV 到目前为止，疫苗还不能完全预防婴儿和儿童的潜在危险副作用。 这项研究的长期目标是克服活减毒HRSV的安全挑战 并提供HRSV生命周期的基本知识，以设计替代的抗HRSV 接近了。为了达到这些目标，这项建议集中在病毒基质的分子操作上 (M)增强减毒活疫苗安全性的蛋白质。M蛋白对复制和发挥作用是必不可少的 在病毒粒子组装过程中发挥着重要作用，其中许多过程与生产高度相关， 病毒颗粒的组成、释放，或许还有稳定性。因此对M函数有了更好的理解 为疫苗的进步提供了巨大的潜力。开发了一种新的系统，该系统基于一种传染病 缺乏M基因(M-NULL)的病毒，允许快速筛选和操纵M功能。通过提供 表达M突变体的质粒感染M-NULL病毒的细胞，初步筛选鉴定为M 有可能调节活病毒感染性后代生产水平的突变。这项建议 利用基于M-NULL的系统同样识别和操作与程序集相关的M函数并测试 活体内的翻译潜力，通过以下特定目的(略)：1)识别M功能 和突变，以及调节病毒组装、组成和释放的潜在机制。2) 基于M确定对传播缺陷的活病毒的免疫应答质量 体内和体外的突变。3)测试有希望的M突变病毒在攻击后保护小鼠的能力 野生型HRSV。这些目标将提高我们对HRSV复制和 测试M蛋白操纵的潜力，以帮助产生安全的减毒活疫苗。

Human Respiratory Syncytial Virus (HRSV) is the single largest viral cause of pediatric bronchiolitis and pneumonia. With an estimated mortality of >100,000 children per year woridwide, development of an anti- HRSV vaccine is a priority. Among the current approaches, live-attenuation is attractive because, unlike inactivated vaccines, it promises to induce a broad and balanced immune response. However, live HRSV vaccines have so far been unable to fully prevent potentially dangerous side-effects in infants and children. The long-term objectives of this research are to overcome the safety challenges of live-attenuated HRSV vaccines and to contribute fundamental knowledge ofthe HRSV life cycle to design alternative anti-HRSV approaches. To meet these objectives, this proposal focusses on molecular manipulation of the viral matrix (M) protein to enhance safety of live-attenuated vaccines. The M protein is essential for replication and plays a prominent role in virion-assembly processes, many of which are highly relevant for the production, composition, release, and perhaps stability of virus particles. A better understanding of M functions therefore offers significant potential for vaccine advancements. A novel system was developed based on an infectious virus lacking the M gene (M-null) which allows rapid screening and manipulation of M functions. By providing plasmids expressing M mutants to cells infected with the M-null virus, a preliminary screen identified M mutations with potential to regulate the level of infectious progeny production of a live virus. This proposal utilizes the M-null based system to likewise identify and manipulate assembly-relevant M functions and test the translational potential in vivo, through the following Specific Aims (abbreviated): 1) Identify M functions and mutations, and the underlying mechanisms, that regulate virus assembly, composition, and release. 2) Determine the quality ofthe immune response to live viruses with transmission-deficiencies based on M mutations, in vitro and in vivo. 3) Test promising M mutant viruses for ability to protect mice after challenge with wildtype HRSV. Together these aims will raise our fundamental understanding of HRSV replication and test the potential of M protein manipulation to contribute to the generation of a safe live-attenuated vaccine.