Multivalent nanocluster universal influenza vaccine given by microneedle patch

流感是一项重大的公共卫生风险。目前的季节性流感疫苗在以下方面有效 在健康成年人中保护免受密切匹配的病毒。因为持续的基因变化发生在 流感，季节性流感疫苗有很大的局限性，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。一个 通用流感疫苗将克服这些挑战。在我们之前和初步的研究中，我们有 通过解离流感M2的保守胞外结构域制备双层蛋白质纳米簇 以甲型流感病毒三聚体血凝素(HA)茎抗原为核心的蛋白质(M2e)纳米粒为核心 在岩心表面上。我们还将流感内部核蛋白(NP)分解成颗粒状核心，并 交联化的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 对甲型流感两个系统发育组的病毒，包括可能发生大流行的禽类的保护 菌株。 甲型和乙型流感都能引起流行病。在这个提案中，我们将开发一种多价双层 由新设计的、保守的、来自两种疫苗的抗原蛋白组成的纳米簇通用流感疫苗 甲型和乙型流感以及一种分子佐剂。这种纳米簇疫苗将诱导很强的交叉免疫 在不同的实验动物模型中对甲型和乙型流感的保护。最优的纳米团簇 该制剂将被封装成可溶解的微针(MN)贴片，以开发一种无注射器、无痛、 耐热、自行皮下注射的通用流感疫苗。这三个具体目标是： 目的1.从乙型流感病毒中制备三聚体HA杆状抗原，并从两者中获得保守的NPs 流感A和B，从这些和先前设计制造和表征纳米簇 保守的抗原蛋白。我们已经从这两种物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的系统发育群。我们将制作新颖的双层结构 来自以前和新设计的保守流感抗原蛋白的纳米簇。 目标2.测试层状纳米簇或多价最优组合是否会产生保护 在小鼠身上对抗跨越甲型和乙型流感的病毒。我们将优化疫苗配方 在小鼠中诱导广泛的反应性免疫反应和交叉保护以及在目标3中的进一步研究。 目标3.将最优的多价纳米簇配方封装成可溶的MN贴片和 测试以MN为基础的皮肤疫苗在小鼠和雪貂身上的保护范围。可溶MN 基于贴片的皮肤流感疫苗接种比传统注射器注射有许多优点，包括 无痛、无针、自主给药、冷链独立配送。 总体而言，我们的研究将开发一种广泛交叉保护的通用流感疫苗

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