Virus-like particle vaccines against respiratory syncytial virus

描述（由申请人提供）：我们的长期目标是开发一种安全、有效的疫苗来预防呼吸道合胞病毒（RSV）疾病。RSV是婴儿毛细支气管炎、病毒性肺炎、呼吸衰竭和机械通气的主要原因。在美国，它每年导致> 120，000名婴儿住院，是婴儿病毒性死亡的主要原因。在老年人中，RSV导致三分之一的肺炎住院。在儿童和成人中，RSV感染是哮喘急性加重的主要原因。没有许可的疫苗。已经开发了实验性RSV疫苗，包括灭活、减毒活疫苗、亚单位疫苗、病毒载体疫苗和DNA疫苗，并在动物模型和小型临床试验中进行了测试。然而，这些努力基本上都不成功。灭活疫苗和亚单位疫苗有引起免疫病理学的历史。RSV减毒活疫苗安全。然而，减毒RSV特征性地诱导瞬时/弱免疫，如野生型RSV。 我们正在开发新型病毒样颗粒（VLP）RSV疫苗。VLP是基因工程蛋白质的复合物，其颗粒状病毒样结构缺乏病毒遗传物质，因此不能复制。作为VLP呈递的病毒蛋白具有高度免疫原性，并诱导保护性体液、细胞和粘膜免疫应答。我们拥有在VLP上表达病毒蛋白和评估VLP诱导的免疫力的丰富经验。RSV融合体（F）和附着糖蛋白（G）含有所有已知的中和抗体表位和几个T细胞表位。我们假设表达RSV F、G或F和G表面的VLP将诱导强RSV特异性免疫应答和免疫力。 目的一：利用重组杆状病毒系统在昆虫细胞中表达VLP蛋白，并利用梯度离心法纯化VLP。我们拥有这些方法的经验，这些方法已被FDA批准用于人类使用，并可扩展用于生产。我们将通过共表达这些蛋白与流感基质M1蛋白作为VLP核心来产生RSV-G、RSV-F和RSV-GF VLP。共表达粒细胞-巨噬细胞集落刺激因子（GM-CSF）蛋白的VLP表现出增强的细胞和体液应答。我们将产生具有和不具有锚定的GM-CSF的RSV VLP。此外，我们将通过测试三种不同的联合给药佐剂，氢氧化铝（明矾），CpG寡脱氧核苷酸（CpG ODN）和单磷酰脂质（MPL），优化免疫原性。后两种佐剂刺激已知对于抗RSV应答重要的Toll样受体信号传导途径。在目的2中，我们将在小鼠模型中定义这些RSV VLP疫苗的免疫原性和有效性。将用VLP鼻内致敏和加强小鼠。免疫后将定量RSV特异性抗体（IgG和伊加）和T细胞应答。为了确定保护作用，我们将用已知在BALB/c小鼠中产生比实验室RSV毒株更高的肺病毒载量的重组嵌合RSV毒株攻击小鼠。总的来说，这些实验将推进RSV VLP疫苗的开发，并可能导致急需的RSV疫苗获得批准。 公共卫生相关性：呼吸道合胞病毒（RSV）每年在美国导致约120，000名婴儿住院，并且是婴儿细支气管炎和病毒性死亡的主要原因。尽管对传统疫苗方法进行了数十年的研究，但没有批准的RSV疫苗。我们正在推进RSV的病毒样颗粒（VLP）疫苗，我们预测RSV VLP将是预防RSV疾病的有效和安全的疫苗。

DESCRIPTION (provided by applicant): Our long term goal is to develop a safe, effective vaccine to prevent respiratory syncytial virus (RSV) disease. RSV is the leading cause of bronchiolitis, viral pneumonia, respiratory failure, and mechanical ventilation in infants. It causes >120,000 infant hospitalizations per year in the USA and is the leading cause of infant viral death. In the elderly, RSV causes one third of pneumonia hospitalizations. In children and adults, RSV infection is a major cause of acute asthma exacerbations. There is no licensed vaccine. Experimental RSV vaccines including inactivated, live attenuated, subunit, viral-vectored, and DNA have been developed and tested in animal models and small clinical trials. However, these have been largely unsuccessful. Inactivated and subunit vaccines have a history of causing immunopathology. Live attenuated RSV vaccines are safe. However, attenuated RSVs characteristically induce transient/poor immunity like wild-type RSV. We are developing novel virus-like particle (VLP) RSV vaccines. VLPs are genetically engineered complexes of proteins in a particulate virus-like structure that lacks viral genetic material and therefore cannot replicate. Viral proteins presented as VLPs are highly immunogenic and induce protective humoral, cellular, and mucosal immune responses. We have extensive experience expressing viral proteins on VLPs and evaluating VLP-induced immunity. The RSV fusion (F) and attachment glycoprotein (G) contain all known neutralizing antibody epitopes and several T cell epitopes. We hypothesize that VLPs expressing RSV F, G, or both F and G surface will induce strong RSV-specific immune responses and immunity. In Aim 1, we will generate VLPs using a recombinant baculovirus system to express VLP proteins in insect cells and gradient ultracentrifugation to purify VLPs. We have experience with these methods which are FDA approved for human use and scalable for production. We will generate RSV-G, RSV-F, and RSV-GF VLPs by co-expressing these proteins with influenza matrix M1 protein as the VLP core. VLPs co-expressing granulocyte-macrophage colony-stimulating factor (GM-CSF) proteins exhibit enhanced cellular and humoral responses. We will generate RSV VLPs with and without anchored GM-CSF. Also, we will optimize immunogenicity by testing three different co-administered adjuvants, aluminum hydroxide (alum), CpG oligodeoxynucleotides (CpG ODN), and monophosphoryl lipid (MPL). The latter two adjuvants stimulate toll- like receptor signaling pathways known to be important for anti-RSV responses. In Aim 2, we will define the immunogenicity and efficacy of these RSV VLP vaccines in a mouse model. Mice will be primed and boosted with VLPs intranasally. RSV-specific antibody (IgG and IgA) and T cell responses will be quantified after immunization. To determine protection, we will challenge mice with a recombinant, chimeric RSV strain known to give higher lung viral loads than laboratory RSV strain in BALB/c mice. Collectively, these experiments will advance RSV VLP vaccine development and may lead to a much needed approved RSV vaccine. PUBLIC HEALTH RELEVANCE: Respiratory syncytial virus (RSV) causes apprimately 120,000 infant hospitalizations in the US each year and is the leading cause of bronchiolitis and viral death in infants. Despite decades of research with traditional vaccine approaches, there are no approved RSV vaccines. We are advancing virus-like particle (VLP) vaccines for RSV, and we predict RSV VLPs will be effective and safe vaccines that prevent RSV disease.