Novel vaccine strategies for prevention of N. meningitidis group B disease

说明(申请人提供)：目前还没有广泛有效的疫苗可用于预防脑膜炎奈瑟氏菌B组菌株引起的疾病，脑膜炎奈瑟菌占脑膜炎双球菌疾病病例的50%或更多。由于B组胶囊是一种自身抗原，因此需要替代疫苗策略。我们正在研究H因子结合蛋白(FHBP)的疫苗潜力，fHBP是一种表面暴露的脂蛋白，以前称为GNA1870。血清抗fHBP抗体与细菌结合，激活补体介导的细菌溶解，也抑制人类补体下调调节蛋白H因子(Fh)的结合。在没有结合Fh的情况下，生物体更容易受到补体介导的细菌溶解的影响。FHBP作为疫苗的一个局限性是抗原可变性(3个组，组内有一些亚变异体)。变异组1中的fHBP产生的抗fHBP抗体对表达变异体2或3蛋白的菌株无杀菌作用。为了绕过这个问题，我们正在构建表达不同变异组表位的重组嵌合fHBP分子。在小鼠中，原型嵌合fHBP疫苗能诱导出针对表达fHBP变异体1、2或3的菌株的血清杀菌抗体。由于重组的fHBP可能不表达所有对激发广泛杀菌活性重要的表位，我们还在从高表达fHBP的脑膜炎奈瑟菌菌株中制备天然的外膜囊泡(OMV)疫苗。由于洗涤剂处理用于从传统OMV疫苗中去除内毒素提取fHBP，我们通过灭活脂A生物合成基因LpxL1来减弱内毒素活性。来自突变株的天然OMV在刺激人PBMC产生促炎细胞因子方面的活性比来自野生型株的OMV低1000至10,000倍。在小鼠中，来自过表达fHBP的LpxL1基因敲除突变体的天然OMV疫苗比对照重组fHBP或洗涤剂提取的OMV疫苗具有更广泛的杀菌活性。在这项应用中，我们将在这些遗传方法的基础上开发出更安全、更广泛有效的疫苗。在目标1中，我们将定义两个抗fHBP单抗的位置，因为这两个抗fHBP单抗的表位在fHBP变异群中是保守的，并且这两个单抗与第二个抗fHBP单抗介导协同杀菌活性。在目标2中，我们将构建第二代重组嵌合fHBP疫苗，旨在诱导比迄今制备的原型嵌合疫苗更广泛的杀菌抗体反应。在目标3中，我们将构建额外的脑膜炎奈瑟氏菌突变体，从不同的变异组过度表达fHBP，其中编码不想要的抗原的基因被灭活。这些突变体将被用于生产高度免疫原性和安全的OMV疫苗，该疫苗将在小鼠身上进行免疫原性评估，随后在非人类婴儿灵长类动物中进行免疫原性评估。拟议的研究将确定这些疫苗方法是否有可能预防包括B组在内的所有脑膜炎奈瑟氏菌疾病。由于其他病原体也与fh结合以绕过固有宿主防御，因此所学到的经验可能适用于其他疫苗的开发。

DESCRIPTION (provided by applicant): There are no broadly effective vaccines available for prevention of disease caused by group B strains of N. meningitidis, which account for 50% or more of cases of meningococcal disease. Because the group B capsule is an autoantigen, alternative vaccine strategies are needed. We are investigating the vaccine- potential of factor H binding protein (fHbp) a surface-exposed lipoprotein previously called GNA1870. Serum anti-fHbp antibodies bind to the bacteria, activate complement-mediated bacteriolysis, and also inhibit binding of the human complement down-regulatory protein, factor H (fH). In the absence of bound fH, the organism becomes more susceptible to complement-mediated bacteriolysis. One limitation of fHbp as a vaccine is antigenic variability (3 groups with some subvariants within groups). Anti-fHbp antibodies elicited by fHbp in variant group 1 are not bactericidal against strains expressing variant 2 or 3 proteins. To circumvent this problem, we are constructing recombinant chimeric fHbp molecules that express epitopes from different variant groups. In mice, a prototype chimeric fHbp vaccine elicited serum bactericidal antibodies against strains expressing fHbp variant 1, 2 or 3. Because recombinant fHbp may not express all epitopes important for eliciting broad bactericidal activity, we also are preparing native outer membrane vesicle (OMV) vaccines from N. meningitidis strains engineered to over-express fHbp. Since detergent treatments used to remove endotoxin from conventional OMV vaccines extract fHbp, we attenuate endotoxin activity by inactivating a lipid A biosynthesis gene, LpxL1. A native OMV from the mutant was 1000- to 10,000-fold less active in stimulating human PBMCs to produce proinflammatory cytokines than OMV from the wildtype strain. In mice, a native OMV vaccine from an LpxL1 knockout mutant with over-expressed fHbp elicited broader bactericidal activity than control recombinant fHbp or detergent-extracted OMV vaccines. In this application, we will build on these genetic approaches to develop even safer and more broadly effective vaccines. In Aim 1, we will define locations of epitopes of two anti-fHbp mAbs that are of special interest because the epitopes are conserved across fHbp variant groups and the mAbs mediate cooperative bactericidal activity with second anti-fHbp mAbs. In Aim 2, we will construct second-generation recombinant chimeric fHbp vaccines designed to elicit even broader bactericidal antibody responses than the prototype chimeric vaccines prepared to date. In Aim 3, we will construct additional N. meningitidis mutants with over-expressed fHbp from different variant groups, and in which genes encoding unwanted antigens are inactivated. The mutants will be used to produce highly immunogenic and safe OMV vaccines, which will be evaluated for immunogenicity in mice and, subsequently, in nonhuman infant primates. The proposed studies will determine whether these vaccine approaches have the potential to prevent all N. meningitidis disease, including group B. Since other pathogens also bind fH to circumvent innate host defenses, the lessons learned may be applicable to development of other vaccines.