Rational design of transferrin binding protein-based vaccines to combat gonorrhea

人类病原体淋病奈瑟菌（Ngo）能够利用人转铁蛋白作为唯一来源 铁。从人转铁蛋白（hTf）的有效铁转运需要表达TonB依赖的，整合的 外膜转运蛋白（TbpA）和表面暴露脂蛋白（TbpB）。Tbps的表达式为： 在男性志愿者研究中，淋球菌建立人类感染是必要的，这意味着淋球菌在人类感染中的关键作用。 这些蛋白质在人类宿主中。因为所有的淋球菌都表达这种转运系统， 由于这些组分是很保守的，因此将在本提案中探索它们作为疫苗靶点的潜力。的 在拟议研究中要检验的总体假设是，合理设计的疫苗由以下组成： 工程化的TbpA和TbpB抗原与实验选择的佐剂组合，将提供 针对无症状定植和致病性炎症免疫交叉保护 多样的N.淋病菌株。该提案的具体目标如下：目标1。合理设计 TBP疫苗在这个目标中，淋球菌Tbps的结构将被定义有和没有hTf。这些 结构将指导诱变工作，生产配体结合缺陷且缺乏的疫苗抗原 具有最小结构破坏的高变序列。目标二。优化人源化疫苗递送 下生殖道感染的雌性小鼠模型。佐剂、递送途径、剂量和时间表将在 在女性下生殖道感染模型中测试了最佳保护作用， hTf。免疫学相关因素和保护的决定因素将通过广泛测试体液和 细胞免疫因子目标3. Tbp疫苗对新型人源化感染的保护效果试验 模型将测试优化的抗原、佐剂、途径和时间表在人源化小鼠中的保护作用。 盆腔炎、男性尿道炎和鼻咽感染模型。同样，免疫学 将描述这些新的动物感染模型中保护作用的相关因素和决定因素。目标4。 评价合理设计的疫苗在所有感染模型中的交叉保护作用。优化，合理- 设计的疫苗配方将测试对广泛的Ngo菌株的保护作用，包括 耐抗生素的“超级细菌”菌株。保护的免疫学相关性将通过这些验证 在所有人源化小鼠感染模型中，这些研究意义重大，因为它们可能导致 开发针对已产生耐药性的顽固病原体的有效疫苗 现有的治疗方法。这些研究具有创新性，因为它们将采用结构导向疫苗 设计开发配体结合缺陷疫苗抗原，假设其表现出增强的 与野生型蛋白质相比，它们具有更高的免疫原性。此外，这些抗原将被测试用于保护， 将定义新的人源化小鼠感染模型和保护的免疫相关性， 这是我们对非政府组织免疫力理解上的一个关键缺口。

The human pathogen, Neisseria gonorrhoeae (Ngo) is capable of utilizing human transferrin as the sole source of iron. Efficient iron transport from human transferrin (hTf) requires expression of a TonB-dependent, integral outer membrane transporter (TbpA), and a surface-exposed lipoprotein (TbpB). Expression of the Tbps is necessary for the gonococcus to establish human infection in male volunteer studies, implying a critical role for these proteins in the human host. Because this transport system is expressed by all gonococci and the protein components are well conserved, their potential as vaccine targets will be explored in this proposal. The overarching hypothesis to be tested in the proposed study is that a rationally-designed vaccine consisting of engineered TbpA and TbpB antigens, combined with an experimentally selected adjuvant, will provide immunologic cross-protection against both asymptomatic colonization and pathogenic inflammation caused by diverse N. gonorrhoeae strains. The specific aims of the proposal are as follows: Aim 1. Rational design of Tbp-based vaccines. In this aim, structures of the gonococcal Tbps will be defined with and without hTf. These structures will guide mutagenesis efforts to produce vaccine antigens defective in ligand binding and lacking hypervariable sequences with minimal structure disruption. Aim 2. Optimizing vaccine delivery in humanized female mouse model of lower genital tract infection. Adjuvants, delivery routes, doses and schedules will be tested for optimal protection in the female lower genital tract infection model using humanized mice expressing hTf. Immunological correlates and determinants of protection will be defined by broadly testing humoral and cellular immunity factors. Aim 3. Test for protective efficacy of Tbp-based vaccines in new humanized infection models. Optimized antigens, adjuvants, routes and schedules will be tested for protection in humanized mouse models of pelvic inflammatory disease, male urethritis and nasopharyngeal infection. Again, immunological correlates and determinants of protection in these new animal models of infection will be characterized. Aim 4. Evaluate rationally designed vaccines for cross protection in all models of infection. Optimized, rationally- designed vaccine formulations will be tested for protection against a broad group of Ngo strains, including antimicrobial resistant “superbug” strains. Immunological correlates of protection will be validated with these strains in all of the humanized mouse models of infection. These studies are significant since they may lead to the development of an efficacious vaccine against a recalcitrant pathogen that has developed resistance to existing therapeutic methods. These studies are innovative because they will employ structure-guided vaccine design to develop ligand-binding incompetent vaccine antigens, which are hypothesized exhibit enhanced immunogenicity compared to the wild-type proteins. Moreover, these antigens will be tested for protection in novel humanized mouse models of infection and immunological correlates of protection will be defined, closing a key gap in our understanding of immunity against Ngo.