Mechanisms of DDO Adjuvancy

摘要 能够诱导持久的1型细胞免疫的佐剂，包括Th1 CD4+T细胞和 细胞毒性CD8+T细胞在细胞内病原体疫苗的研制中受到高度重视 它们可以躲避抗体。我们已经确定了一种有效的新型佐剂，它可以引起保护性的1型偏向 回应。这些佐剂是从缺陷病毒基因组(Ddos)中提取的人工合成的rna寡核苷酸。 并通过与细胞Rig-I样受体(RLR)结合来触发强烈的免疫反应。DDO包含唯一的 免疫刺激基序，我们认为对他们触发RLR信号的能力是必要的。在小鼠中，DDoS 诱导I型IFN和其他细胞因子的局部表达并促进DC在 引流淋巴结节。此外，DDoS促进依赖于I型干扰素IgG2b/c偏向抗体反应 能够保护小鼠免受致命病毒攻击，并诱导产生干扰素γ抗原的特异性CD_4和CD_8+T细胞 细胞。此外，DDoS与基于角鲨烯的佐剂AddaVax协同作用，提供强大的类型-1 对AddaVax+DDoS佐剂疫苗的免疫偏向。这些数据支持我们的核心假设 DDoS代表了一类新的佐剂，刺激RLR/I型干扰素信号轴驱动 最佳的长寿1型体液和细胞免疫。 该方案中的实验使用了洛佩兹和 Scott实验室评估DDO诱导的T细胞反应的质量、寿命和保护能力 在疫苗接种过程中，并表征负责指导 DDO治疗后的1型免疫应答。具体地说，在目标1中，我们将使用我们的能力来跟踪 DDoS在体内，与一系列报告和转基因小鼠一起鉴定DDoS的早期相互作用 与免疫系统的细胞，特别是树突状细胞，并确定潜在的关键目标的发展 第一类有偏见的反应。在目标2中，我们将评估T辅助1细胞的发育和质量，细胞毒性T 细胞、T滤泡辅助细胞和组织驻留T细胞对单用DDoS佐剂疫苗的反应 或与AddaVax结合使用，并评估I型IFN在建立这些反应中的作用。在《目标3》中， 我们将测试DDoS诱导对细胞内原生动物寄生虫的CD4+介导的保护的能力 利什曼原虫，因为对这种寄生虫的保护依赖于CD4+Th1细胞，并独立于 抗体，我们将使用一个模型直接评估DDoS诱导保护性T细胞反应的能力 雪貂接种流感疫苗。

SUMMARY Adjuvants able to induce long lasting type-1 cellular immunity, including both Th1 CD4+ T cells and cytotoxic CD8+ T cells, are highly sought out for the development of vaccines against intracellular pathogens that evade antibodies. We have identified a powerful new class of adjuvant that elicits protective type-1 biased responses. These adjuvants are synthetic RNA oligonucleotides derived from defective viral genomes (DDOs) and trigger strong immune responses by engaging cellular RIG-I-like receptors (RLRs). DDOs contain a unique immunostimulatory motif that we identified as essential for their ability to trigger RLR signaling. In mice, DDOs induce localized expression of type I IFNs and other cytokines and promote accumulation of DCs in the draining lymph node. In addition, DDOs promote a type I IFN-dependent IgG2b/c-biased antibody response able to protect mice from lethal virus challenge and induce IFNγ-producing antigen-specific CD4+ and CD8+ T cells. Moreover, DDOs synergize with the squalene-based adjuvant AddaVax, providing strong type-1 immunity bias to vaccines adjuvanted with AddaVax+DDOs. These data support our central hypothesis that DDOs represent a new class of adjuvants that stimulate the RLR/type I IFN signaling axis to drive optimal long-lived type-1 humoral and cellular immunity. Experiments in this proposal use the combined expertise and unique set of tools of the Lopez and Scott laboratories to assess the quality, longevity, and protective capacity of DDO-induced T cell responses during vaccination, and to characterize specific molecular and cellular mechanisms responsible for directing the type-1 immune response after DDO administration. Specifically, in Aim 1 we will use our ability to track DDOs in vivo, together with a series of reporter and transgenic mice, to identify the early interactions of DDOs with cells of the immune system, in particular DCs, and to identify potential key targets for the development of type -1 biased responses. In Aim 2, we will assess the development and quality of T helper 1 cells, cytotoxic T cells, T follicular helper cells, and tissue resident T cells in response to a vaccine adjuvanted with DDOs alone or in combination with AddaVax, and assess the role of type I IFNs in establishing these responses. In Aim 3, we will test the ability of DDOs to induce CD4+-mediated protection against the intracellular protozoan parasite Leishmania, since protection against this parasite is dependent upon CD4+ Th1 cells and is independent of antibodies, and we will directly assess the ability of DDOs to induce protective T cell responses using a model influenza vaccine in ferrets.