Combination Adjuvants to Program Durable Immunity to Respiratory Viral and Fungal Pathogens

摘要：病毒和真菌引起的呼吸道感染是全球主要的公共卫生问题。 除流感病毒外，目前还没有针对病毒或真菌的许可疫苗。人们普遍认为 T细胞记忆的诱导对于防御呼吸道中的病毒和真菌是至关重要的。我们和其他人 已经表明，诱导组织驻留记忆（TRM）CD 8和CD 4 T细胞和系统迁移记忆 CD 4 T细胞分别对预防甲型流感病毒（IAV）和吸入性真菌至关重要。然而，在这方面， TRM细胞和系统记忆性CD 4 T细胞都经历了磨损，导致短暂的免疫， 尤其是对于TC 1/TH 1细胞。因此，诱导持久的T细胞免疫对免疫系统构成了重大挑战。 疫苗学家与TH 1细胞相比，TH 17细胞显示出干细胞性、耐久性和细胞周期性的受欢迎属性。 功能可塑性我们建议定制组合佐剂以利用T17编程并诱导持久的免疫应答。 以及保护肺TRM细胞和迁移记忆CD 4 T细胞对抗病毒和真菌。 我们发现，Adjuplex，一种纳米乳剂佐剂，当与TLR 4激动剂吡喃葡萄糖结合时， 脂质A（GLA）在肺中引起抗原特异性CD 8和CD 4 T细胞应答，所述应答是：（i）持久的， 多方面的（TC 1/TC 17/TH 1/TH 17），和（ii）赋予针对IAV的持续>400天的异亚型免疫。 我们进一步发现，将这些佐剂与真菌的β-葡聚糖配体芽生菌内切葡聚糖酶2（Bl-Eng 2; Dectin-2激动剂）和β-葡聚糖颗粒（Dectin-1激动剂）增强抗病毒TC 17/TH 17/TC 1/TH 1和抗肿瘤活性 迁移性记忆T细胞可以预防真菌性肺炎。通过单细胞RNAseq，我们发现， 联合佐剂诱导表达ICOS的记忆性抗病毒和抗真菌CD 8和CD 4 T细胞簇 （可诱导的T细胞共刺激分子）、转录因子c-Maf和促进组织驻留的转录组， 干细胞性和非致病性T17编程。环状二核苷酸也促进T17编程， 肺因此，我们假设编程性干细胞样、功能可塑性、非致病性TC 17/TH 17 使用我们的组合佐剂（参与TLR-4，Dectin-1/2和STING途径）的记忆细胞将促进 对肺部病毒和真菌病原体的持久保护性免疫。我们的具体目标将测试三个 假设：目的1：诱发TC 17/TH 17干细胞样功能可塑性TRM或 系统迁移记忆将产生对呼吸道病毒和真菌病原体持久免疫力;目标2： TC 17/TH 17记忆的功能可塑性对于针对病毒和真菌的保护性免疫是重要的; ICOS/c-Maf途径是建立和/或维持持久疫苗诱导的保护性免疫应答的必要条件。 对呼吸道病毒和真菌感染的免疫力。拟议的工作意义重大，影响深远，因为 它有可能创造一个易于处理的佐剂系统/工具包，将促进疫苗的配制： 靶向粘膜或肠胃外给药;（ii）设计用于诱导TRM或全身性T细胞记忆;和（iii） 定制以引发CD 8和CD 4 T细胞，以保护免受病毒和真菌等多种病原体的侵害。

ABSTRACT: Respiratory infections with viruses and fungi constitute major public health problems globally. Except for influenza virus, there are no licensed vaccines against viruses or fungi. It is generally agreed that induction of T-cell memory is critical for defense against viruses and fungi in the respiratory tract. We and others have shown that induction of tissue-resident memory (TRM) CD8 and CD4 T cells and systemic migratory memory CD4 T cells are essential for protection against influenza A virus (IAV) and inhaled fungi, respectively. However, both TRM cells and systemic memory CD4 T cells undergo attrition, leading to short-lived immunity, which is especially true for TC1/TH1 cells. Therefore, induction of durable T-cell immunity poses major challenges for vaccinologists. As compared to TH1 cells, TH17 cells display sought-after attributes of stem-ness, durability and functional plasticity. We propose to tailor combination adjuvants to harness T17 programming and induce durable and protective lung TRM cells and migratory memory CD4 T cells against viruses and fungi. We find that Adjuplex, a nano-emulsion adjuvant, when combined with the TLR4 agonist glucopyranosyl lipid A (GLA), evokes antigen-specific CD8 and CD4 T-cell responses in the lung that are: (i) durable and multifaceted (TC1/TC17/TH1/TH17), and (ii) confer heterosubtypic immunity against IAV that persists >400 days. We further find that combining those adjuvants with fungal CLR ligands Blastomyces endoglucanase 2 (Bl-Eng2; Dectin-2 agonist) and b-glucan particles (Dectin-1 agonist) augments antiviral TC17/TH17/TC1/TH1 and elicits migratory memory T cells that protect against fungal pneumonia. By single-cell RNAseq, we found that our combined adjuvants induce memory antiviral and antifungal CD8 and CD4 T-cell clusters that express ICOS (Inducible T Cell Co-stimulator), the transcription factor c-Maf, and a transcriptome that fosters tissue residency, stem cell-ness and non-pathogenic T17 programming. Cyclic dinucleotides also promote T17 programming in lungs. This, we postulate that programming stem cell-like, functionally plastic, non-pathogenic TC17/TH17 memory cells with our combination adjuvants (that engage TLR-4, Dectin-1/2 and STING pathways) will foster durable protective immunity to viral and fungal pathogens in the lung. Our specific aims will test three hypotheses: Aim 1: Combination adjuvants that evoke TC17/TH17 stem cell-like functionally-plastic TRM or systemic migratory memory will engender durable immunity to respiratory viral and fungal pathogens; Aim 2: Functional plasticity of TC17/TH17 memory is important for protective immunity to viruses and fungi; Aim 3: The ICOS/c-Maf pathway is integral to establishment and/or maintenance of durable vaccine-induced protective immunity to respiratory viral and fungal infections. The proposed work is significant and of high impact because it has the potential to create a tractable adjuvant system/tool kit that will advance the formulation of vaccines: (i) targeted for mucosal or parenteral administration; (ii) designed to induce TRM or systemic T-cell memory; and (iii) tailored to elicit CD8 and CD4 T cells that protect against diverse pathogens such as viruses and fungi.