Effect of presentation methods on the molecular mechanism of combinatorial adjuvants

 说明(申请人提供)：疫苗接种是人类历史上最成功的公共卫生干预措施之一。尽管许多不治之症和高死亡率疾病得到了完全控制，在某些情况下从人类文明中消失了，但许多其他疾病仍然难以得到疫苗干预。我们能够更好地了解自然感染病原体如何通过激活多条免疫途径并产生组合反应来触发和控制哺乳动物的免疫，这将极大地帮助设计和临床翻译针对新出现的传染病、艾滋病毒/艾滋病、癌症等的更有效的疫苗。我们在刺激疫苗佐剂方面的大部分知识来自病毒、细菌、真菌或寄生虫的病原体/危险相关分子模式(PAMP/DAMP)。在自然感染中，多个佐剂和Ag被携带在一个颗粒状结构中，天然免疫细胞作为单个颗粒中的一个结合实体经历佐剂和抗原，其作用是定位和集中一组协同刺激信号。因此，我们认为：(A)为了开发更有效的疫苗，我们必须了解多种佐剂作用于天然免疫细胞并协同控制获得性免疫的分子机制；以及(B)在体内呈现佐剂组合的最谨慎的方式是通过模仿病原体(病原体样颗粒，PLP)的颗粒载体。这些载体的性质(如大小、电荷、成分)的变化将影响佐剂与先天性免疫细胞相互作用的方式，并调节由此产生的免疫反应；这一方面也可能发生在自然感染中。我们的主要假设是，两种临床相关的佐剂CpG和单磷脂A(Mpla)的体内组合效应可以(A)被呈现方式(可溶的，与不同物理性质的颗粒载体相比)精确地调节，以及(B)当作为颗粒载体一起输送时，通过体外试验更好地预测。为了测试这些，我们提出了以下目标。目的1：制备不同大小、不同佐剂密度的CpG/MPLA共负载致病素样颗粒(PLP)佐剂，并对其进行表征。目的2：体外研究不同DC亚群对联合佐剂制剂反应的分子机制。目的3：在体内鉴定联合佐剂制剂协同全身免疫反应背后的分子机制。总之，拟议的研究将极大地促进我们对联合佐剂有效启动适应性免疫的分子机制的理解，(B)确定控制联合佐剂和PLP佐剂的关键物理化学参数，以及(C)导致具有即时翻译潜力和临床应用的基于组合佐剂的人类疫苗的新平台技术和试剂的开发。

 DESCRIPTION (provided by applicant): Vaccination is one of the most successful public health interventions in human history. Despite the fact that many incurable and high-mortality diseases are fully controlled, and in some cases eliminated from human civilization, many other diseases remain elusive to vaccine interventions. Our ability to better understand how natural infectious pathogens trigger and control mammalian immunity by activating multiple immune-pathways and generating a combinatorial response, will greatly help the design and clinical translation of more effective vaccines against emerging infectious diseases, HIV/AIDS, cancer, etc. Most of our knowledge in stimulating vaccine adjuvants, molecules that stimulate the immune system to generate protective or therapeutic immunity against antigens, comes from pathogen/danger-associated molecular patterns (PAMPs/DAMPs) of viruses, bacteria, fungi or parasites. In natural infections, multiple adjuvants and Ag are carried inside a particle-like structure and innate immune cells experience adjuvant and antigens as a combination entity in single particles, which acts to localize and concentrate a set of synergistic stimulatory signals. Thus, we argue, that (a) in order to develop more efficacious vaccines we must understand the molecular mechanisms by which multiple adjuvants act on innate immune cells and in tandem, control adaptive immunity; and (b) the most prudent way of presenting combinations of adjuvants in vivo is through particulate carriers that mimic pathogens (Pathogen-like particles, PLPs). Variation of these carriers' properties (e.g. size, charge, composition) will affect how adjuvants interact with innate immune cells and modulate the resulting immune response; an aspect that likely occurs in natural infections as well. Our overarching hypotheses are that the combinatorial in vivo effects of two clinically-relevant adjuvants, CpG and Monophosphoryl Lipid A (MPLA), can be (a) precisely modulated by the mode of presentation (soluble, vs particulate carriers of different physical properties) and (b) better predicted by in vitro assays when delivered together as particulate carriers. To test these, we propose the following aims. Aim 1: Develop and characterize CpG/MPLA co-loaded pathogen-like-particle (PLP) adjuvants of various sizes and adjuvant- density. Aim 2: Investigate in-vitro, the molecular mechanisms involved in how various DC subsets respond to combination adjuvant formulations. Aim 3: Identify in vivo, the molecular mechanisms behind synergistic systemic immune-responses to combination adjuvant formulations. Collectively, the proposed studies will (a) greatly advance our understanding of molecular mechanisms that mediate potent priming of adaptive immunity by combination-adjuvants, (b) identify key physico-chemical parameters that control adjuvanticity of combination-adjuvants and PLPs, and (c) lead to the development of new platform technologies and reagents for combinatorial-adjuvant based human vaccines with immediate translational potential and clinical use.