HA Surface Presented Yeast Flu Vaccine and Its Enhancement by CD154 Codisplay

说明(申请人提供)：接种疫苗是目前唯一能有效阻止流感病毒在人与人之间传播的方法。传统的以鸡蛋为基础的流感疫苗技术受到影响，因为如果发生全球疫情，它们无法大规模和快速地生产针对流行性流感的疫苗。在此，基于PI以前在蛋白质表面展示方面的工作，提出了一种新型的有效流感疫苗。这项新技术的核心是在酵母表面展示一种病毒蛋白，如禽流感H5N1血凝素(HA)，使这些重组酵母可以用作流感疫苗。众所周知，酵母能够表达功能性透明质酸。透明质酸的表面展示可以显著促进宿主免疫系统对抗原的识别，并与酵母细胞膜成分介导免疫佐剂效应。尽管与哺乳动物细胞相比，酵母具有不同的糖基化能力，但这种有限的糖基化实际上可能是研制酵母疫苗的一个好处。据报道，天然流感病毒在人类体内和标准疫苗中的糖基化实际上通过阻止抗体进入HA表面而干扰免疫诱导。这一好处将在这项工作中得到彻底的调查。或者，将糖化酵母用于构建HA表面展示的酵母疫苗，以评估这些疫苗的免疫变化。为了提高酵母疫苗的效力和免疫原性，将采用双蛋白表面展示技术将HA与CD154共展示。HA与CD154的共展示将进一步刺激抗原提呈细胞的成熟，促进获得性免疫反应。综上所述，提出了一种假设，即HA表面展示的酵母疫苗可以诱导对流感的强大保护性免疫。此外，这些有效的疫苗可以通过与CD154共展示HA来增强。最近在Pi的实验室进行的一项动物疫苗研究支持了这一假设。为了进一步验证这一假设，本项目提出了三个目标：目标1，制备HA表面呈现的酵母流感疫苗；目标2，研究酵母疫苗在动物(小鼠)中诱导的体液和细胞免疫反应；目标3，展示酵母疫苗对小鼠的致死性禽流感的保护作用。该项目的长期目标是将动物研究转化为临床前研究，确定这些酵母疫苗对人类的免疫原性，并加强这项技术，以开发其他疫苗来治疗许多与病毒感染相关的疾病，如艾滋病和癌症。与基于病毒的疫苗不同，酵母疫苗在人类身上使用是安全的，疫苗存储不需要冷藏。此外，它们可以以低成本大规模、快速地生产。此外，这些疫苗不需要病毒通过卵子，为疫苗种子株的开发提供了一种可能性，使其与原始的“野生”病毒更接近，并有可能转化为更好的免疫原性和有效的反应。与公共卫生相关：在没有任何控制措施(疫苗接种或药物)的情况下，据估计，在美国，3-2级4大流行可能影响15%至35%的美国人口，经济影响可能在710亿至1670亿美元之间。这项拟议的技术推进了疫苗生产技术，不仅可用于大规模快速制造禽流感疫苗，还可用于其他疫苗，如癌症疫苗。因此，这项技术有可能广泛影响我们治疗和预防病毒相关疾病的能力，具有无数潜在的社会效益和巨大的商业潜力。

DESCRIPTION (provided by applicant): Vaccination is currently the only method that can effectively impede the spread of influenza viruses among people. Traditional egg-based influenza vaccine technologies suffer from their incapability of massive and rapid production of vaccines against circulating influenza if a global outbreak occurs. Here, a new type of potent influenza vaccines is proposed based on the PI's previous works on protein surface display. The core of this new technology is to display a viral protein such as avian influenza H5N1 hemagglutinin (HA) on yeast surface in a manner such that these recombinant yeasts can serve as vaccines against influenza. Yeast is known of being able to express functional HA. The surface display of HA can significantly facilitate the recognition of antigens by host immune systems and mediate an immunoadjuvant effect with yeast cell membrane components. Although yeast has different glycosylation capability as compared to mammalian cells, this limited glycosylation may actually be a benefit to formulating a yeast vaccine. It has been reported that the glycosylation of natural influenza viruses in humans and in the standard vaccine actually interferes with induction of immunity by preventing access of antibodies to the HA surface. This benefit will be investigated thoroughly in this work. Alternatively, glycosylated yeast will be employed to construct HA surface-displayed yeast vaccines for evaluating the alteration of immunity of these vaccines. To enhance the efficacy and immunogenicity of the yeast vaccines, a dual-protein surface display technique will be adopted to codisplay the HA with CD154. The codisplay of HA with CD154 will further stimulate the maturation of antigen presenting cells and promote the adaptive immune response. Taken together, a hypothesis was developed that HA surface-displayed yeast vaccines can elicit strong and protective immunity against influenza. Furthermore, these potent vaccines can be enhanced by co-displaying HA with CD154. A recent animal vaccination study conducted in PI's lab supports this hypothesis. This project is proposed to further verify this hypothesis, three aims are proposed: Aim 1, Generate HA surface presented yeast influenza vaccines; Aim 2, Characterize both humoral and cell-mediated immune responses induced by yeast vaccines in animals (mice); Aim 3, Demonstrate the protection of mice from lethal avian influenza using yeast vaccines. The long-term goals of this project are to translate the animal studies into preclinical studies, determine the immunogenicity of these yeast vaccines in humans, and to augment this technology to develop other vaccines for treating many virus- infection related diseases such as AIDS and cancers. Unlike virus-based vaccines, yeast vaccines are safe for use in humans, and vaccine storage does not require refrigeration. Moreover, they can be massively and rapidly produced at a low cost. In addition, these vaccines do not require the passage of virus through eggs, offering a possibility for vaccine seed strain development that more closely matches the original "wild" virus and translating potentially into a better immunogenic and effective response. PUBLIC HEALTH RELEVANCE: In the absence of any control measures (vaccination or drugs), it has been estimated that in the United States a 3medium2level4 pandemic could affect between 15% and 35% of the U.S. Population, and the economic impact could range between $71 and $167 billion. The proposed technology advances vaccine production technology, which can be used for massive and rapid manufacturing not only avian influenza vaccines but also other vaccines such as cancer vaccines. Thus, this technology has the potential to impact broadly on our ability to treat and prevent virus-related diseases, stating with innumerable potential society benefits and large commercial potential.