Novel Influenza A Nanovaccines for Broad Cross Protection

描述(申请人提供)：流感是一个世界性的公共卫生问题。尽管目前的流感疫苗在对抗密切匹配的病毒方面是有效的，但目前疫苗的主要限制是需要每个季节都生产新的疫苗，选择正确的毒株存在不确定性，以及无法防止新的流感大流行。迫切需要诱导针对多种A型流感病毒的广泛保护性免疫反应的改进疫苗，不仅用于季节性流感，而且用于预防大流行性流感。保守的表位是此类疫苗的有效免疫原。应综合考虑包括改进抗原、新的抗原传递技术和疫苗接种方案在内的综合战略，以克服目前在开发通用流感疫苗方面的障碍。在该项目中，我们建议生成编码保守的流感抗原的结构，这些抗原在自然病毒感染和季节性疫苗接种中通常不被宿主免疫系统感知。我们将使用最新的纳米技术方法，从产生的抗原蛋白中直接自组装出纳米簇，作为通用流感疫苗候选。我们将研究这些纳米疫苗在小鼠和豚鼠模型中诱导的交叉保护的广度，以及交叉保护的免疫相关性。1.构建构象稳定的HA茎结构域(CSHA-STRK)和四聚体M2e串联重复序列(TM2ET)重组蛋白的表达载体。HA中相对保守的茎结构域在病毒粒子中被其球状头部结构域所屏蔽，并且不能被免疫细胞有效地感知。没有球形头部的CSHA茎将克服这一限制，并保持其天然构象。此外，随着表位密度的增加，tM2et将保留M2e的四聚体结构。2.制备CSHA-STEK和tM2e抗原自组装的纳米团簇。这些纳米尺寸的颗粒将直接从产生的抗原性重组蛋白中自我组装，并在摄取后释放完整的抗原。这些颗粒将最大限度地增加抗原有效载荷，控制空间抗原递呈，并调节DC的释放。我们将对颗粒的大小、抗原含量、分布和释放以及DC的内化进行表征。3.研究上述纳米簇诱导的免疫反应和保护性免疫的广度，以及广泛交叉保护的免疫相关性。基于来自两个系统发育组的HA茎结构域的组合的疫苗可以预防所有的甲型流感病毒。TM2et的加入将进一步增加 有可能受到广泛的保护。我们将研究不同纳米疫苗组合诱导的免疫反应，一组甲型流感病毒株对攻击的交叉保护，以及交叉保护的免疫相关性。

DESCRIPTION (provided by applicant): Influenza is a worldwide public health problem. Although current influenza vaccines are effective in battling closely matched viruses, major limitations of current vaccines are the need to produce new vaccines every season, the uncertainty in choice of the correct strains, and the inability to prevent a new influenza pandemic. Improved vaccines inducing broadly protective immune responses against multiple type A influenza viruses are urgently needed, not only for seasonal influenza but also for pandemic influenza prevention. Conserved epitopes are potent immunogens for such vaccines. Combined strategies including improved antigens, new antigen delivery techniques and vaccination regimens should be considered together to overcome current obstacles in the development of a universal influenza vaccine. In the project we propose to generate constructs encoding conserved influenza antigens which are not usually sensed the by host immune system in natural viral infection and seasonal vaccination. We will use the newest nanotechnology approach to produce nanoclusters self-assembled directly from the resulting antigenic proteins as universal influenza vaccine candidates. We will investigate the breadth of cross-protection induced by these nanovaccines in mouse and guinea pig models, and the immune correlate of the cross protection. The three specific aims to be investigated are: 1. Generation of constructs expressing conformation-stabilized HA stalk domain (csHA-stalk) and tetrameric M2e tandem repeat (tM2et) recombinant proteins. The relatively conserved stalk domain in HA is shielded by its globular head domain in virions and is less effectively sensed by immune cells. A csHA-stalk without the globular head will overcome this limitation, and retain its native conformation. Also, tM2et will retain the tetrameric structure of M2e with increased epitope density. 2. Production of nanoclusters self-assembled from csHA-stalk and tM2e antigens. These nano-size particles will be directly self-assembled from the resulting antigenic recombinant proteins, and release intact antigens after uptake. These particles will maximize antigenic payload, control spatial antigen presentation, and regulate release in DCs. We will characterize the size of the particles, antigen content, distribution and release, and internalization by DCs. 3. Investigation of immune responses and the breadth of protective immunity induced by the above nanoclusters and of immune correlates of broad cross-protection. Vaccines based on a combination of HA stalk domains from the two phylogenetic groups may protect against all influenza A viruses. The addition of tM2et will further increase the potential for broad protection. We will study immune responses induced by different nanovaccine combinations, the cross protection to challenge by a panel of influenza A viral strains, and immune correlates of the cross protection.