Multivalent nanocluster universal influenza vaccine given by microneedle patch

流感是一种主要的公共卫生风险。目前的季节性流感疫苗对 保护健康成人免受密切匹配的病毒的侵害。因为连续的基因变化发生在 但是，季节性流感疫苗存在重大局限性，包括需要生产新疫苗 每个季节，选择疫苗株的不确定性，以及无法预防新的流感大流行。一 通用流感疫苗将克服这些挑战。在我们先前和初步的研究中， 通过去溶剂化流感病毒M2的保守胞外域， 蛋白（M2e）的纳米颗粒作为核心和交联甲型流感三聚体血凝素（HA）茎抗原 核心表面上。我们还将流感病毒内部核蛋白（NP）去溶剂化为颗粒核心， 交联M2e以产生双层纳米团簇。由此产生的层状纳米团簇诱导交叉 保护免受甲型流感两个系统发育组的病毒，包括大流行性潜在禽流感病毒 菌株 甲型流感和B型流感均可引起流行病。在这个建议中，我们将开发一个多价双层 纳米簇通用流感疫苗，由新设计的、保守的、来自两种病毒的抗原蛋白组成， 流感A和B，以及分子佐剂。这种纳米簇疫苗将诱导较强的交叉免疫 在不同的实验室动物模型中对甲型流感和B流感的保护。最佳纳米团簇 制剂将被封装到可溶解的微针（MN）贴剂中，以开发一种无刺，无痛， 热稳定的、自我给药的皮肤接种的通用流感疫苗。这三个具体目标是： 目标1.产生来自流感B的三聚体HA茎抗原和来自两者的保守NP的构建体 流感A和B，从这些和先前设计纳米团簇制造和表征 保守抗原蛋白。我们已经从这两种蛋白质中产生了结构稳定的HA茎蛋白。 甲型流感病毒（hrH1和hrH3）和四聚体M2e的系统发育组。我们将制造新型的双层 来自先前和新设计的保守流感抗原蛋白的纳米簇。 目标二。测试层状纳米团簇或多价最佳组合是否会诱导保护 在小鼠中对抗跨越甲型流感和B流感的病毒。我们将优化疫苗配方 在小鼠中诱导广泛反应性免疫应答和交叉保护，并在目的3中进一步研究。 目标3：将最佳多价纳米簇制剂封装到可溶性MN贴片中， 在小鼠和雪貂中测试基于MN的皮肤疫苗接种的保护范围。可溶性MN 与传统的注射器注射相比，基于贴片的皮肤流感疫苗接种具有许多优点， 无痛、无针、自我给药和冷链独立配送。 总的来说，我们的研究将开发出一种具有广泛交叉保护性的通用流感疫苗

SUMMARY Influenza is a major public health risk. Current seasonal influenza vaccines are effective in protecting against closely matched viruses in healthy adults. Because continuous genetic changes occur in influenza, there are major limitations to seasonal influenza vaccines including the need to produce new vaccines every season, uncertainty in selecting vaccin strains, and the inability to prevent novel influenza pandemics. A universal influenza vaccine will overcome these challenges. In our previous and preliminary studies, we have produced double-layered protein nanoclusters by desolvating the conserved ectodomain of the influenza M2 protein (M2e) into nanoparticles as cores and crosslinking influenza A trimeric hemagglutinin (HA) stalk antigens onto the core surfaces. We have also desolvated influenza internal nucleoprotein (NP) into particulate cores and cross linked M2e to generate double-layered nanoclusters. The resulting layered nanoclusters induced cross protection against viruses from both phylogenetic groups of influenza A, including pandemic potential avian strains. Both influenza A and B can cause epidemics. In this proposal, we will develop a multivalent double-layered nanocluster universal influenza vaccine composed of newly designed, conserved, antigenic proteins from both influenza A and B, and a molecular adjuvant. This nanocluster vaccine will induce strong cross immune protection against both influenza A and B in different laboratory animal models. The optimal nanocluster formulation will be encapsulated into dissolvable microneedle (MN) patches to develop a syringe-free, painless, thermostable, and self-administered skin-given universal influenza vaccine. The three specific aims are: Aim 1. Generate constructs of trimeric HA stalk antigen from influenza B and conserved NPs from both influenza A and B, fabricate and characterize nanoclusters from these and previously designed conserved antigenic proteins. We have generated structure-stabilized HA stalk proteins from both phylogenetic groups of influenza A (hrH1 and hrH3) and tetrameric M2e. We will fabricate novel double-layered nanoclusters from previous and new designed conserved influenza antigenic proteins. Aim 2. Test whether the layered nanoclusters or a multivalent optimal combination will induce protection against viruses spanning both influenza A and influenza B in mice. We will optimize a vaccine formulation inducing broadly reactive immune responses and cross protection in mice and further studies in Aim 3. Aim 3. Encapsulate the optimal multivalent nanocluster formulation into dissolvable MN patches and test the breadth of protection of the MN-based skin vaccination in both mice and ferrets. Dissolvable MN patch-based skin influenza vaccination has many advantages over conventional syringe injection including painless, needle-free, self-administration and cold chain-independent distribution. Overall, our research will develop a broadly cross-protective universal influenza vaccine