An engineered meningococcal OMV vaccine for Africa against all capsular groups

描述(申请人提供)：脑膜炎奈瑟氏菌在撒哈拉以南非洲引起暴发性流行。大多数是由A组包膜菌株引起的。然而，W-135群和X群毒株也会在该地区引起流行，一旦引入A群多糖-蛋白质结合疫苗大规模免疫，这些毒株就可能出现。我们的目标是为非洲开发一种针对所有衣壳组菌株的脑膜炎双球菌疫苗。我们的方法将是使用为“B组疫苗”确定的新的蛋白质抗原，这种抗原也可以诱导出针对其他胶囊菌株的保护性抗体。这些“非常规”抗原将出现在简单的外膜囊泡(OMV)中，这种囊泡具有强大的天然佐剂。我们的研究将建立在以前洗涤剂提取的OMV疫苗经验的基础上，这种疫苗已被证明对人类安全有效。它们的主要限制是诱导主要针对PorA的血清杀菌抗体，而PorA在抗原性上是可变的。为了扩大对带有异源PorA的菌株的保护，我们准备了B组菌株的突变体，这些突变体被改造成过表达因子H结合蛋白(FHBP)，这是两种有前景的B组重组蛋白疫苗中的新抗原。通过在内毒素的生物合成中引入一个额外的突变，我们减弱了内毒素的活性。在小鼠中，由突变体制备的非洗涤剂处理的OMV疫苗可诱导针对遗传多样性的B组毒株以及来自非洲的A、W-135和X流行群的血清杀菌抗体反应。我们的假设是，由来自非洲的突变株制备的本地OMV疫苗将诱导针对来自非洲的株表达的PorA、fHBP和其他抗原的更广泛的杀菌抗体。此外，脂多糖的突变将消除对OMV洗涤剂提取的需要，OMV在传统OMV疫苗中用于降低LPS，但也会提取所需的抗原，如fHBP。在目标1中，我们将调查来自非洲不同地区的200株脑膜炎球菌的遗传谱系和编码fHBP、PorA和其他疫苗抗原的基因的序列多样性。在目标2中，我们将用定量捕获ELISA法检测抗原的表达，并用流式细胞术检测活菌表面抗原的可及性。在目标3中，我们将创建最近非洲流行毒株的突变株，这些突变株将被改造成表达不止一种PorA分子，过度表达fHBP，并已减弱内毒素。疫苗株还将被选为粘附素/侵袭素NADA天然高表达的疫苗株。我们将从突变体中制备天然的OMV疫苗，并通过检测与该疫苗体外孵育的人PBMC的炎性细胞因子反应来评估OMV的毒性。我们将对小鼠和婴儿灵长类动物进行免疫，并检测针对来自非洲的菌株的血清杀菌抗体反应。这一结果将提供原则性证据，证明OMV疫苗很可能在人类中具有良好的耐受性，并引发广泛的保护性免疫。这些发现将支持在人类身上测试OMV疫苗的应用，以控制由所有衣壳群菌株引起的撒哈拉以南地区的脑膜炎双球菌流行。 公共卫生相关性：脑膜炎球菌在撒哈拉以南非洲引起脑膜炎的爆炸性流行，在几个月内可能涉及10万多例病例。大多数疫情是由A组包膜菌株引起的，但来自其他包膜组的菌株也开始在该地区引起疫情。非洲正在开发一种针对A群疾病的多糖结合疫苗，但令人严重关切的是，一旦采用A群结合疫苗进行大规模免疫，可能会出现带有其他胶囊的菌株并导致流行病。我们建议从突变型脑膜炎球菌菌株开发一种外膜囊泡疫苗，该疫苗是为高表达有希望的蛋白疫苗抗原而设计的，作为非洲通用的脑膜炎球菌疫苗，对抗所有被囊群的菌株引起的疾病。

DESCRIPTION (provided by applicant): Neisseria meningitidis causes explosive epidemics in sub-Saharan Africa. Most are caused by capsular group A strains. However, group W-135 and X strains also cause epidemics in this region, and these strains may emerge once mass immunization with a group A polysaccharide-protein conjugate vaccine is introduced. Our goal is to develop a meningococcal vaccine for Africa that targets strains from all capsular groups. Our approach will be to use novel protein antigens identified for "group B vaccines", which also elicit protective antibodies against strains with other capsules. These "unconventional" antigens will be presented in simple outer membrane vesicles (OMV) that have potent natural adjuvants. Our studies will build on previous experience with detergent-extracted OMV vaccines, which are proven to be safe and effective in humans. Their major limitation is that they elicit serum bactericidal antibodies primarily directed at PorA, which is antigenically variable. To extend protection to strains with heterologous PorA, we prepared mutants of group B strains that were engineered to over-express factor H binding protein (fHbp), which is a novel antigen in two promising group B recombinant protein vaccines. By introducing an additional mutation in LPS biosynthesis, we attenuated endotoxin activity. In mice, non-detergent-treated OMV vaccines prepared from the mutants elicited serum bactericidal antibody responses against genetically diverse group B strains, as well as epidemic group A, W-135 and X strains from Africa. Our hypothesis is that a native OMV vaccine prepared from mutant strains from Africa will elicit even broader bactericidal antibodies directed at PorA, fHbp and other antigens expressed by strains from Africa. Further, the LPS mutation will eliminate the need for detergent extraction of the OMV, which is used to decrease LPS in conventional OMV vaccines, but also extracts desirable antigens such as fHbp. In Aim 1, we will investigate genetic lineages and sequence diversity of genes encoding fHbp, PorA and other vaccine antigens among 200 meningococcal isolates from a geographically diverse collection of strains from Africa. In Aim 2, we will measure antigen expression by a quantitative capture ELISA, and antigen surface-accessibility on live bacteria by flow cytometry. In Aim 3, we will create mutants of recent African epidemic strains, which will be engineered to express more than one PorA molecule, over-express fHbp, and have attenuated endotoxin. The vaccine strains also will be selected for naturally high expression of an adhesin/invasin, NadA. We will prepare native OMV vaccines from the mutants, and assess OMV toxicity by measuring inflammatory cytokine responses of human PBMCs incubated in vitro with the vaccine. We will immunize mice and infant primates and measure serum bactericidal antibody responses against strains from Africa. The results will provide proof of principle that the OMV vaccine is likely to be well-tolerated in humans and elicit broad protective immunity. These findings would support an application to test the OMV vaccine in humans for control of meningococcal epidemics in sub-Sahara caused by strains from all capsular groups. PUBLIC HEALTH RELEVANCE: Meningococci cause explosive epidemics of meningitis in sub-Sahara Africa that can involve more than 100,000 cases in a few months. Most epidemics have been caused by encapsulated group A strains but strains from other capsular groups also have begun to cause epidemics in the region. A polysaccharide conjugate vaccine against group A disease is being developed for Africa but there is grave concern that strains with other capsules may emerge and cause epidemics once mass immunization with the group A conjugate vaccine is introduced. We propose to develop an outer membrane vesicle vaccine from mutant meningococcal strains, engineered for over-expression of promising protein vaccine antigens, as a universal meningococcal vaccine for Africa against disease caused by strains from all capsular groups.