Alternative approaches for NALT-based immunity to respiratory pathogens

描述（由申请方提供）：鼻咽道是衰弱和潜在致死病原体（包括炭疽杆菌和流感病毒）的主要入口。除了一般血流之外，能够促进粘膜组织中的抗体和细胞毒性T细胞应答的粘液瘤疫苗比经典的注射疫苗更有效地防止呼吸道病原体。因此，需要安全的粘膜佐剂和疫苗递送系统。这项资助的先前工作集中在将小鼠鼻相关淋巴网状组织（NALT）定义为针对鼻咽道的免疫应答的诱导位点。我们使用霍乱毒素和开发新的衍生物缺乏ADP核糖基转移酶活性，以规避这种肠毒素佐剂的反应原性。此外，我们还评估了炭疽毒素对小鼠NALT的作用。这些研究还包括炭疽保护性抗原的抗体（Ab）的表征，特别强调保护粘膜组织的潜力。共有五个最初的具体目标得到成功解决。在我们的研究过程中，一个主要的和意想不到的发现是，炭疽杆菌保护性抗原与cAMP诱导炭疽杆菌水肿因子的突变体一起鼻内共给药增强了针对这两种分子的粘膜和全身免疫力。此外，我们发现水肿毒素（EdTx或保护性抗原加水肿因子）衍生物增强了对共同施用的不相关抗原如重组鼠疫耶尔森氏菌F1-V抗原的粘膜和全身免疫。与神经节苷脂结合肠毒素霍乱毒素不同，EdTx及其衍生物在鼻腔应用后都不靶向中枢神经系统组织。在这项更新资助中，我们将解决总体假设，即舌下应用EdTx衍生物将诱导基于NALT的免疫力并保护免受呼吸道病原体的侵害，而不会产生通常与鼻内应用肠毒素相关的不良反应。具体目标之一是建立通过舌下途径与保护性抗原（PA）共同施用的水肿因子衍生物的佐剂活性，以增强对炭疽毒素组分的免疫力。具体目标二将表征用EdTx衍生物作为佐剂的舌下免疫所提供的对呼吸道病毒感染的保护性免疫。具体目标三的研究将确定用于产生分泌型伊加（SIgA）Ab的诱导位点和用EdTx衍生物舌下免疫后的粘膜免疫。最后，具体目标四将确定EdTx衍生物作为舌下佐剂诱导SIgA反应的分子信号。这项资助将揭示EdTx衍生物作为舌下疫苗佐剂并诱导基于NALT的免疫的机制。我们还将验证一种新的粘膜疫苗递送途径，以及新的基于PA的粘膜佐剂，用于诱导针对呼吸道病原体的免疫力。公共卫生相关性：迄今为止，经鼻递送疫苗被认为是触发免疫应答的诱导位点以发展针对呼吸道病原体的最佳免疫的最有效手段。然而，鼻用疫苗中使用的肠毒素佐剂可诱导严重的副作用，这是由于它们靶向中枢神经系统并维持炎症反应的能力。该基金将用于探索水肿毒素衍生物作为舌下疫苗佐剂的有效性。完成后，我们将验证一种新的疫苗输送系统，以及新的粘膜佐剂，用于诱导对呼吸道病原体的免疫力。

DESCRIPTION (provided by applicant): The nasopharyngeal tract is a major portal entry of debilitating and potentially lethal pathogens including Bacillus anthracis and influenza virus. Mucosal vaccines capable of promoting antibody and cytotoxic T cell responses in mucosal tissues, in addition to the general bloodstream, protect more effectively against respiratory pathogens than classical injected vaccines. Therefore, there is a need for safe mucosal adjuvants and vaccine delivery systems. Previous work on this grant focused at defining murine nasal-associated lymphoreticular tissues (NALT) as inductive sites for immune responses targeting the nasopharyngeal tract. We used cholera toxin and developed novel derivatives lacking ADP ribosyl transferase activity to circumvent the reactogenicity of this enterotoxin adjuvant. In addition, we have assessed the action of anthrax toxins on murine NALT. These studies have also included the characterization of antibodies (Abs) to anthrax protective antigens with special emphasis on the potential to protect mucosal tissues. A total of five original Specific Aims were successfully addressed. A major and unexpected finding during the course of our studies was the fact the nasal co-administration of Bacillus anthracis protective antigen together with a mutant of the cAMP-inducing Bacillus anthracis edema factor enhanced mucosal and systemic immunity against these two molecules. Furthermore, we found that the edema toxin (EdTx or protective antigen plus edema factor) derivative enhanced mucosal and systemic immunity to co-administered unrelated antigens such as recombinant Yersinia pestis F1-V antigen. Unlike the ganglioside-binding enterotoxin cholera toxin, neither EdTx nor its derivatives target central nervous system tissues after nasal application. In this renewal grant, we will address the overall hypothesis that sublingual application of EdTx derivatives will induce NALT-based immunity and protect against respiratory pathogens, without the adverse effects often associated with nasal application of enterotoxins. Specific aim one will establish the adjuvant activity of edema factor derivatives co-administered with protective antigen (PA) via the sublingual route for enhanced immunity to anthrax toxin components. Specific aim two will characterize protective immunity to respiratory viral infection afforded by sublingual immunization with EdTx derivatives as adjuvant. Studies in specific aim three will identify inductive sites for the generation of secretory IgA (SIgA) Abs and mucosal immunity after sublingual immunization with EdTx-derivatives. Finally, specific aim four will determine molecular signals underlying the induction of SIgA responses by EdTx-derivatives as sublingual adjuvant. This grant will unravel the mechanisms by which EdTx derivatives act as adjuvant for sublingual vaccines and induce NALT-based immunity. We will also validate a new route for mucosal vaccine delivery, as well as new PA-based mucosal adjuvant(s) for the induction of immunity against respiratory pathogens. PUBLIC HEALTH RELEVANCE: Thus far, nasal delivery of vaccines was believed to be the most effective mean to trigger inductive sites of immune responses for the development of optimal immunity against respiratory pathogens. However, enterotoxin adjuvants employed in nasal vaccines could induce severe side effects resulting from their ability to target the central nervous system and sustain inflammatory responses. This grant will explore the efficacy of edema toxin derivatives as adjuvant for sublingual vaccines. Upon completion, we will have validated a new vaccine delivery system, as well as new mucosal adjuvant(s) for the induction of immunity against respiratory pathogens.