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

项目摘要 该提议是开发一种基于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禽流感致病原因 严重的呼吸道疾病，死亡率高。该病毒的高人畜共患病能力引起了严重的关注。 对于大流行的可能性，其风险可能与H5毒株相似。在取得进展的同时 在H5流感疫苗的研发方面，H7产品一直处于滞后状态。拟议中的H7N9疫苗 将在两个方面有所创新。1)H7N9疫苗通常专注于血凝素(HA)蛋白，作为 主要免疫原，但对HA H7的免疫反应虽然具有保护性，但比对其他HAS的免疫反应弱 在人类身上。因此，TIBA将开发一种多抗原方法，具体包括优化的 病毒粒子蛋白HA、NA、M1和NP。这种方法有望增加免疫原性和异型性。 疫苗的保护潜力。2)传统的脂质纳米粒(LNPs)，它是核的支柱 酸输送，需要很大比例的“结构性”脂类，导致RNA含量相对较低。提巴有 开发了一种改进的树枝状大分子纳米粒子系统，使交付的RNA质量含量最大化，保护 RNA不会被降解，并能在体内被细胞有效摄取。在这里开发的原型组合 将作为一个平台，任何爆发的抗原序列都可以在其中快速实施。提巴将会遇到 开发HA/NA/M1/NP疫苗原型以应对活病毒挑战的近期目标 通过完成三个第一阶段目标，在第二阶段进行实验和IND-Enabling研究。第一个是验证 H7 HA复制子RNA表达载体在体内的表现。这将被生成和制定 在BALB/c小鼠身上测试0.2微克、2.0微克和20微克，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将产生和测试NA、M1和NP mRNA和复制子的单独配方 候选者，通过T细胞ELISpots以类似的增加剂量验证他们在体内的表现，对于NA，也 用酶联免疫吸附试验检测抗体反应。最终目标是合成一种多抗原原型疫苗。 将HA复制子RNA与NA、M1和NP RNA共同封装，使其达到最佳平衡 单一纳米颗粒配方中的有效载荷。这种多抗原候选疫苗将在男性和 雌性BALB/c小鼠，与仅HA和无关抗原编码对照疫苗进行比较，以确定 对所有成分保持免疫原性，如果抗HA反应受益于包含 额外的病毒粒子蛋白。候选原型将与最先进的LNP配方进行比较，以 与目前RNA疫苗的黄金标准相比，评估TIBA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.