Mechanisms Involved in, and Development of, FcR-Enhanced Mucosal Vaccination

摘要：图拉氏杆菌是一种革兰氏阴性的A类细胞内粘膜病原菌。细胞免疫是 而抗体(Abb)则延缓了感染的进程。瞄准 抗原提呈细胞(APC)上的Fc受体(FCR)抗原可增强体液免疫和细胞免疫。 我们假设以感染性疾病的抗原为靶点，如灭活的图拉氏F菌(IFT)，以粘膜FCR为靶点 网站将加强对黏膜挑战的保护。在目标1中，我们将调查预成型的能力 单抗-IFT复合体和单抗+IFT混合物，通过以下方式增强IFT的结合、内化和呈递 抗原特异性T细胞是启动保护性免疫反应的关键事件。我们将：1)研究 MAb：IFT比例对mAb-IFT结合、内化和小鼠APC提呈抗原的影响；2)测定， 使用小鼠APC，如果mAb和IFT的混合物可以用来代替预制的mAb-IFT；3)确定是否 上述FCR靶向策略也增强了IFT的结合、内化和人APC的呈递。在……里面 目的2，我们将在小鼠体内测定mAb-IFT复合体和/或mAb+IFT混合物的能力 I.N.、I.D.、I.M.或S.C.以加强对I.N的保护。或用活的图拉氏丝虫挑战身份，并识别 对观察到的保护至关重要的体液和/或细胞成分。我们将：1)验证最优FCR中介 在目标1中观察到的结合、内化和呈现与由 FCR靶向免疫原注射；2)确定CD8和CD4T细胞，B细胞，FCR，抗体， 和干扰素-γ在FCR依赖保护中的作用，使用缺乏这些免疫成分的小鼠；3)确定 FCR靶向IFT优先定位于淋巴组织，而不是非靶向IFT；4)确定是否有保护作用 对I.N.的指控。还有身份证。质询可以在外围设备(I.D.、I.M.或S.C.)之后生成免疫接种；5) 确定是否包含CTB(I.N.)和明矾(I.D.、I.M.或S.C.)进一步增强FCR产生的保护- 靶向免疫原。在目标3中，我们将验证该疫苗的灵活性和多种病原体的潜力。 站台。具体地说，我们将产生一种针对肺炎链球菌的FCR靶向亚单位疫苗(FC-PSPA)，以及 重大公共卫生问题的细胞外粘膜病原体。PSPA是S的表面组分。 肺炎，在佐剂存在的情况下产生抗体依赖的保护。我们将调查 含有不同量PSPA的单价和多价FC-PSPA结合物的能力，并给予 通过粘膜和外周途径，预防I.N。挑战肺炎链球菌。的重要意义。 上述研究是实质性的：1)新的和更安全的疫苗平台，产生体液和 需要细胞免疫；2)后者在黏膜疫苗的情况下尤其明显；3)有 迫切需要一种有效的图拉氏丝虫黏膜疫苗，以及一种更有效的疫苗 肺炎链球菌；4)了解FCR在粘膜免疫中的作用，以及产生保护作用 抗粘膜病原体，是缺乏的。拟议的研究将填补上述所有领域的重大空白。

SUMMARY: F. tularensis is a gram-negative Category A intracellular mucosal pathogen. Cellular immunity is critical for protection against this organism, while antibodies (Abs) delay the progression of infection. Targeting antigen (Ag) to Fc receptors (FcR) on Ag presenting cells (APC) can enhance humoral and cellular immunity. We hypothesize targeting infectious disease Ag, such as inactivated F. tularensis (iFt), to FcR at mucosal sites will enhance protection against mucosal challenge. In Aim 1, we will investigate the ability of preformed mAb-iFt complexes and mAb plus iFt mixtures, to enhance binding, internalization, and presentation of iFt by APC to Ag-specific T cells, key events in initiating a protective immune response. We will: 1) Examine the impact of mAb:iFt ratio on mAb-iFt-binding, internalization, and Ag presentation by mouse APC; 2) Determine, using mouse APC, if mAb plus iFt mixtures can be used in place of preformed mAb-iFt; 3) Determine if the above FcR-targeting strategies also enhance IFt-binding, internalization, and presentation by human APC. In Aim 2, we will determine in mice, the ability of mAb-iFt complexes and/or mAb plus iFt mixtures administered i.n., i.d., i.m., or s.c. to enhance protection against i.n. or i.d challenge with live F. tularensis, and identify the humoral and/or cellular components critical to the observed protection. We will: 1) Verify optimal FcR-mediated binding, internalization, and presentation observed in Aim 1, correlates with optimal protection generated by FcR-targeted immunogens administered i.n.; 2) Determine the role of CD8 and CD4 T cells, B cells, FcR, Ab, and IFN-gamma in FcR-dependent protection, using mice lacking these immune components; 3) Determine if FcR-targeted iFt preferentially localizes to lymphoid tissues, versus non-targeted iFt; 4) Determine if protection against i.n. and i.d. challenge can be generated following peripheral (i.d., i.m., or s.c.) immunization; 5) Determine if inclusion of CTB (i.n.) and Alum (i.d., i.m., or s.c.) further enhances protection generated by FcR- targeted immunogens. In Aim 3, we will validate the flexibility and the multi-pathogen potential of this vaccine platform. Specifically, we will generate an FcR-targeted subunit vaccine (Fc-PspA) against S. pneumoniae, an extracellular mucosal pathogen of significant public health concern. PspA is a surface component of S. pneumoniae, which generates Ab-dependent protection in the presence of adjuvant. We will investigate the ability of mono- and multivalent Fc-PspA conjugates containing variable amounts of PspA, and administered via mucosal and peripheral routes, to protect against i.n. challenge with S. pneumoniae. The significance of the above studies is substantial: 1) New and safer vaccine platforms, which generate both humoral and cellular immunity are needed; 2) The latter is particularly evident in the case of mucosal vaccines; 3) There is an urgent need for an effective mucosal vaccine against F. tularensis, and a more efficacious vaccine against S. pneumoniae; 4) Knowledge regarding the role of FcR in mucosal immunity, and the generation of protection against mucosal pathogens, is lacking. The proposed studies will fill significant gaps in all the above areas.