Protective Activity of a Multi-Functional Immunogen

描述（由申请人提供）：需要现有人用佐剂的替代品。我们的研究表明，在没有佐剂的情况下，在Ag呈递细胞上靶向抗原（Ag）到人Fcgamma受体I型（hFcgammaRI）可以增强体外T细胞的活化和体内Ag特异性抗体（Ab）和细胞因子的产生。然而，还没有人创造出重组免疫原，从而可以在传染病模型中检查hFcgammaRI靶向方法，或对所涉及的机制进行深思熟虑的研究。肺炎链球菌是一种呼吸道病原体，其保护性银（PspA）已被确定。PspA是一种细菌表面蛋白，当与佐剂一起用作免疫原时，可在小鼠中产生对肺炎链球菌感染的保护作用。我们假设：1)用抗hfcgammari - pspa融合蛋白免疫可以增强（在没有佐剂的情况下）对肺炎链球菌感染的保护。2)增强的机制可能涉及到银在淋巴组织中的定位改变，以及银加工的改变。为了测试后者，我们将生成抗hfcgammari - hel融合蛋白。HEL是一种蛋白质Ag，通常用于进行Ag加工和提呈的机制研究，在体外和体内。在specific Aim 1中，我们将：A)去人源化已经生成的抗hfcgammari - pspa和抗hfcgammari - HEL融合蛋白。B)检测融合蛋白在体外和体内增强T细胞和B细胞反应的能力。C)在体内，确定hFcgammaRI靶向Ag是否部分通过增强Ag在淋巴组织的定位而起作用。在特异性目标2中，我们将：A)测试抗hFcgammaRI-PspA融合蛋白对肺炎链球菌攻击的保护能力。B)利用生化、免疫学和超微结构技术研究hFcgammaRI靶向对Ag加工和呈现的影响。拟议的研究将为进一步的hFcgammaRI靶向研究奠定基础，在小鼠和人类中使用各种传染病剂，消除对传统佐剂的需求，大大减少接种个体所需的Ag量（降低成本和潜在毒性），并提供一种同时增强体液和细胞免疫反应的疫苗递送系统。因此，这些研究和这种疫苗策略将适用于成人、儿童和免疫功能低下人群中针对多种病原体的疫苗。

DESCRIPTION (provided by applicant): Alternatives to current human adjuvants are needed. Our studies demonstrate that in the absence of adjuvant, targeting antigen (Ag) to human Fcgamma receptor type I (hFcgammaRI) on Ag presenting cells enhances T cell activation in vitro and Ag-specific antibody (Ab) and cytokine production in vivo. However, no one has created recombinant immunogens, which permit examination of the hFcgammaRI targeting approach in an infectious disease model, or the deliberate study of the mechanism(s) involved. Strepto- coccus pneumoniae is a respiratory pathogen for which a protective Ag (PspA) has been identified. PspA is a bacterial surface protein, which, when used as an immunogen with adjuvant, generates protection against S. pneumoniae infection in mice. We hypothesize: 1) That protection against S. pneumoniae infection can be enhanced (in the absence of adjuvant) by immunizing with an anti-hFcgammaRI-PspA fusion protein. 2) That the mechanism(s) of enhancement will involve alterations in Ag localization to lymphoid tissues, as well as alterations in Ag processing. To test the latter, we will generate an anti-hFcgammaRI-HEL fusion protein. HEL is a protein Ag commonly used to conduct mechanistic studies of Ag processing and presentation, in vitro and in vivo. In Speific Aim 1 we will: A) Dehumanize anti-hFcgammaRI-PspA and anti-hFcgammaRI- HEL fusion proteins already generated. B) Examine the ability of the fusion proteins to enhance T and B cell responses in vitro and in vivo. C) Determine, in vivo, if hFcgammaRI targeting of Ag works, in part, through enhanced localization of Ag to lymphoid tissues. In Specific Aim 2 we will: A) Test the ability of the anti- hFcgammaRI-PspA fusion protein to protect against challenge with S. pneumoniae. B) Study the influence of hFcgammaRI targeting on Ag processing and presentation utilizing biochemical, immunological and ultrastructural techniques. The proposed studies will lay the ground-work for further hFcgammaRI targeting studies using a variety of infectious disease agents in mouse and human, eliminate the need for traditional adjuvant, substantially reduce the amount of Ag required to vaccinate an individual (reducing cost and potential toxicity), and provide a vaccine delivery system, which simultaneously enhances humoral and cellular immune responses. Thus, these studies and this vaccine strategy will be applicable to vaccines against a wide variety of pathogens in adult, pediatric, and immunocompromised populations.