Rapid response to pandemic influenza via multi-antigen RNA-based vaccine

项目摘要 该提案旨在开发基于RNA的快速反应流感疫苗原型，以支持NIAID的呼吁 用于预防大流行性流感威胁的核酸疫苗。甲型H7N9禽流感病毒 严重的呼吸道疾病，死亡率很高。这种病毒的高度人畜共患病能力引起了人们的严重关注 这种病毒的风险可能与H5病毒株的风险相似。虽然进展 在H5流感疫苗的发展，H7产品已经落后。建议的H7N9疫苗 将在两个方面有所创新。1)H7N9疫苗通常集中在血凝素（HA）蛋白， 主要免疫原，但对HA H7的免疫应答虽然具有保护性，但比对其他HA的免疫应答弱 在人类身上。因此，Tiba将开发一种多抗原方法，特别是包括优化比例的 病毒体蛋白HA、NA、M1和NP。这种方法预期增加免疫原性和异型性。 疫苗的保护潜力。2)常规的脂质纳米颗粒（LNP），其是核酸纳米颗粒的支柱。 酸递送需要大比例的“结构”脂质，导致相对低的RNA含量。提巴有 开发了一种改进的树状聚合物纳米颗粒系统，最大限度地提高了递送的RNA质量含量， RNA降解，并使细胞在体内有效摄取。这里开发的原型组成 将作为一个平台，可以快速实施任何爆发抗原序列。Tiba会满足 近期目标是开发HA/NA/M1/NP原型疫苗，以推进活病毒攻击 通过完成三个第一阶段目标，在第二阶段进行实验和IND使能研究。一是验证 图10示出了基于H7 HA复制子RNA的表达构建体的体内性能。这将产生和制定 在BALB/c小鼠中以0.2 µg、2.0 µg和20 µg进行测试，以测量细胞和 体液免疫原性这些研究将确定后续实验所需的最低剂量。 在第二个目标中，Tiba将生成并测试NA、M1和NP mRNA和复制子的个体制剂 候选物，通过T细胞ELISpot以类似的增加剂量验证其体内性能，并且对于NA，还 通过ELISA来测量抗体应答。最终目的是构建多抗原疫苗原型 将HA复制子RNA与NA、M1和NP RNA的最佳平衡通过共包封这四种RNA来组合， 在单一纳米颗粒制剂中的有效载荷。这种多抗原候选疫苗将在男性和 雌性BALB/c小鼠，与仅HA和不相关的抗原编码对照疫苗进行比较，以确定 保留了针对所有组分的免疫原性，并且如果抗HA应答受益于包含免疫原性， 另外的病毒体蛋白。候选原型将与最先进的LNP配方进行比较， 与目前RNA疫苗的黄金标准相比，评估Tiba的递送技术的性能。

Project Summary The proposal is to develop an RNA-based, rapid-response influenza vaccine prototype, supporting NIAID’s call for nucleic acid-based vaccines that protect against pandemic influenza threats. Avian influenza A H7N9 causes severe respiratory illness with a high mortality rate. The virus’ high zoonotic capacity has raised serious concerns over the possibility of a pandemic, with the risk being potentially similar to that of H5 strains. While progress has been made in the development of H5 influenza vaccines, H7 products have lagged. The proposed H7N9 vaccine will be innovative in two respects. 1) H7N9 vaccines typically focus on the hemagglutinin (HA) protein as the main immunogen, but immune responses to HA H7, while protective, are weaker than those against other HAs in humans. Therefore, Tiba will develop a multi-antigen approach, specifically including optimized ratios of the virion proteins HA, NA, M1, and NP. This approach is expected to increase the immunogenicity and heterotypic protective potential of the vaccine. 2) Conventional lipid nanoparticles (LNPs), which are the mainstay of nucleic acid delivery, require a large proportion of “structural” lipid, resulting in a relatively low RNA content. Tiba has developed a modified dendrimer nanoparticle system that maximizes the delivered RNA mass content, protecting RNA from degradation, and enables efficient uptake by cells in vivo. The prototype composition developed here will serve as a platform into which any outbreak antigen sequences could be rapidly implemented. Tiba will meet the near-term goal of developing a prototype HA/NA/M1/NP vaccine to advance toward live virus challenge experiments and IND-enabling studies in Phase 2 by completing three Phase 1 Aims. The first is to validate performance of a H7 HA replicon RNA-based expression construct in vivo. This will be generated and formulated with Tiba’s delivery system and tested in BALB/c mice at 0.2 µg, 2.0 µg, and 20 µg to measure cellular and humoral immunogenicity. These studies will establish the minimum dose required for subsequent experiments. In the second Aim, Tiba will generate and test individual formulations of NA, M1, and NP mRNA and replicon candidates, validating their performance in vivo at similarly increasing doses by T cell ELISpots and, for NA, also by ELISA to measure antibody responses. The final Aim is to compose a multi-antigen prototype vaccine combining the optimal balance of HA replicon RNA with NA, M1, and NP RNAs by co-encapsulating the four payloads in a single nanoparticle formulation. This multi-antigen candidate vaccine will be tested in male and female BALB/c mice, in comparison to HA-only and irrelevant antigen-coding control vaccines to determine if immunogenicity is retained against all components, and if anti-HA responses benefit from inclusion of the additional virion proteins. The candidate prototypes will be compared to state-of-the-art LNP formulations to evaluate the performance of Tiba’s delivery technology compared to the current gold-standard for RNA vaccines.