Preclinical Development of a Thermostable Trivalent Filovirus Vaccine

项目摘要 丝状病毒引起暴发性出血热，在人类暴发中病死率高达90%。 虽然丝状病毒（埃博拉病毒（EBOV）和马尔堡病毒（MARV））仅在中部某些地区流行， 非洲和菲律宾，它们的极端毒性和武器化的潜力导致了 确定这两个都是对美国国家安全和医疗对策的高度优先生物威胁， 被优先纳入国家战略储备。虽然被动策略 免疫治疗最近取得了重大进展，疫苗接种仍然是最经济， 技术上可行的方法，以保护更大的人群获得这些病毒感染之一。 虽然多种埃博拉候选疫苗最近在临床开发方面取得了重大进展， 路径，目前还不知道哪种丝状病毒可能会导致公共卫生紧急情况。因此， 需要开发一种普遍安全的、满足储存要求的多价丝状病毒疫苗。 本项目的总体目标是研制一种耐热、三价、重组亚单位丝状病毒疫苗 它可以保护处于危险中的人群免受EBOV（扎伊尔埃博拉病毒）的所有致病株的感染， SUDV（苏丹埃博拉病毒）和MARV。将通过冻干重组体来实现热稳定性。 允许在使用时重构的抗原。高度纯化的重组丝状病毒亚单位蛋白是 从稳定转化的昆虫细胞表达。该生产系统的一个关键优势是能够 始终如一地产生大量纯的、稳定的和正确折叠的病毒蛋白。免疫亲和 色谱法是高效生产的关键，并且通过使用植物表达的 克隆抗体与其他疫苗技术不同，重组亚单位方法允许包含 从不同的病原体抗原，以实现真正的广谱疗效。抗原剂量的微调， 免疫时间表和佐剂选择允许将新的或修饰的靶标快速包含到核心中 疫苗制剂。在拟议工作期间将进一步确定这一核心提法。独特的 我们的平台的优势最初是针对证明在候选人的可行性与保护 针对三种丝状病毒，具有仅通过使用高度纯化的亚单位蛋白质可实现的安全性特征。 本研究分为四个具体目标：在目标1中，将测试三价制剂的有效性 针对丝状病毒病的非人灵长类动物（NHP）模型中的三种丝状病毒中的每一种。在Aim 2中， 冻干抗原的免疫原性将在小鼠中得到证实， 与液体（非冻干）制剂相比，将对每种病毒的挑战进行检测 使用NHP模型。在目标3中，我们将确定最终剂量水平，并测试最终剂量的防护耐久性。 配方（长达12个月）。目的4是关注抗原生产方法的发展， 完全可规模化和受控的工艺，以生成三种抗原的良好表征的抗原批次。

PROJECT SUMMARY Filoviruses cause fulminant hemorrhagic fevers with a case-fatality rate of up to 90% in human outbreaks. Although filoviruses (Ebolavirus (EBOV) and Marburgvirus (MARV)) are endemic only to certain parts of central Africa and the Philippines, their extreme virulence and potential for weaponization have lead to the determination that both are high priority biothreats to US national security and medical countermeasures have been prioritized for acquisition into the Strategic National Stockpile. While strategies for passive immunotherapy have made significant progress recently, vaccination remains the most economical and technically feasible approach to protect larger groups of people from acquiring one of these viral infections. While multiple Ebola vaccine candidates have recently made significant progress on their clinical development path, it is unknown which of the filoviruses may next cause a public health emergency. Therefor a continuing need exists to develop a universally safe, multivalent filovirus vaccine that meets stockpiling requirements. The overall goal of this project is to develop a thermostable, trivalent, recombinant subunit filovirus vaccine that can protect at risk populations against infection by all pathogenic strains of EBOV (Zaire Ebolavirus), SUDV (Sudan Ebolavirus), and MARV. Thermostability will be achieved by lyophilization of the recombinant antigens allowing reconstitution at the time of use. The highly purified recombinant filovirus subunit proteins are expressed from stably transformed insect cells. A key advantage of this production system is the ability to consistently produce large quantities of pure, stable, and properly folded viral proteins. Immunoaffinity chromatography is key for the highly efficient production and is being facilitated by the use of plant-expressed monoclonal antibodies. Unlike other vaccine technologies, the recombinant subunit approach permits inclusion of antigens from diverse pathogens to achieve truly broad spectrum efficacy. Fine tuning of antigen dosing, immunization schedule, and adjuvant selection allow the rapid inclusion of new or modified targets into a core vaccine formulation. This core formulation will be further defined during the proposed work. The unique advantages of our platform are initially targeted at demonstrating feasibility in a candidate with protection against three filoviruses, having a safety profile only achievable with the use of highly purified subunit proteins. This research is divided into four Specific Aims: In Aim 1, the trivalent formulation will be tested for efficacy against each of the three filoviruses in non-human primate (NHP) models of filovirus disease. In Aim 2 the immunogenicity of lyophilized antigens will be confirmed in mice and then the lyophilized trivalent formulation will be tested against challenge with each of the viruses in comparison to a liquid (non-lyophilized) formulation using NHP models. In Aim 3, we will define the final dose level and test the durability of protection of the final formulation (up to 12 months). Aim 4 is focused on antigen production methods development establishing a fully scaleable and controlled process to generate well-characterized antigen lots of the three antigens.