Effect of presentation methods on the molecular mechanism of combinatorial adjuvants

 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已得到充分控制，在某些情况下已从人类文明中消除，但许多其他疾病仍然难以通过疫苗干预措施来治疗。我们有能力更好地了解天然传染性病原体如何通过激活多种免疫途径和产生组合反应来触发和控制哺乳动物免疫，这将极大地帮助设计和临床转化更有效的疫苗来对抗新出现的传染病，艾滋病毒/艾滋病，癌症等。刺激免疫系统产生针对抗原的保护性或治疗性免疫的分子来自病毒、细菌、真菌或寄生虫的病原体/病原体相关分子模式（PAMP/DAMP）。在自然感染中，多种佐剂和Ag被携带在颗粒状结构内，并且先天免疫细胞经历佐剂和抗原作为单个颗粒中的组合实体，其用于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先天性免疫细胞并串联控制适应性免疫的分子机制;（B）体内呈现佐剂组合的最谨慎方式是通过模拟病原体的颗粒载体（病原体样颗粒，PLPs）。这些载体的性质（例如大小、电荷、组成）的变化将影响佐剂如何与先天免疫细胞相互作用并调节所得的免疫应答;这一方面也可能发生在自然感染中。我们的首要假设是，两种临床相关佐剂CpG和单磷酰脂质A（MPLA）的组合体内效应可以（a）通过呈现模式（可溶性，与不同物理性质的颗粒载体相比）精确调节，以及（B）当作为颗粒载体一起递送时，通过体外测定更好地预测。为了检验这些，我们提出以下目标。目的1：开发和表征各种尺寸和佐剂密度的CpG/MPLA共负载的病原体样颗粒（PLP）佐剂。目的2：在体外研究各种DC亚群对联合佐剂制剂的反应的分子机制。目的3：在体内鉴定对组合佐剂制剂的协同全身免疫应答背后的分子机制。总的来说，所提出的研究将（a）极大地推进我们对介导通过组合佐剂的获得性免疫的有效引发的分子机制的理解，（B）确定控制组合佐剂和PLP的佐剂性的关键理化参数，以及（c）导致开发具有立即翻译潜力和临床用途的基于组合佐剂的人疫苗的新平台技术和试剂。

 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.