Preclinical Development of Flagellin as a Mucosal Adjuvant in Biodefense Vaccines

描述（由申请人提供）：革兰氏阴性鞭毛蛋白是一种非常有效的先天和适应性免疫反应激活剂。我们的工作以及其他人的工作已经确定鞭毛蛋白在小鼠中是一种有效的佐剂。我们还发现鞭毛蛋白在非人类灵长类动物中是一种有效的佐剂。鞭毛蛋白具有使其成为优良佐剂的特性，包括：效力；粘膜活跃;鞭毛蛋白免疫宿主的有效性；易于在细菌中生产；5)在蛋白质的高变区插入抗原序列而不丧失鞭毛蛋白的生物活性的能力。我们已经明确地确定，鞭毛蛋白是一种高效的粘膜佐剂，在小鼠和非人灵长类动物中产生针对鼠疫杆菌抗原的强特异性抗体反应，并促进对鼠疫杆菌致命呼吸道攻击的保护。由于鞭毛蛋白作为人类粘膜佐剂具有巨大的潜力，我们提出以下目标，将推进鞭毛蛋白用于人类的发展：确定鞭毛蛋白/Y的功效。鼠疫F1和V融合蛋白作为保护性粘膜疫苗。我们产生了一种重组蛋白，编码鞭毛蛋白和鼠疫杆菌的F1和V抗原，并确定该融合蛋白保留了鞭毛蛋白的生物活性。我们打算确定融合蛋白是否在小鼠和食蟹猴中促进抗f1和V IgG反应，并保护小鼠免受鼠疫杆菌CO92的呼吸攻击。具体目标2。设计并实施符合良好生产规范（GMP）的鞭毛蛋白/Y生产工艺。鼠疫疫苗可以很容易地用于临床前、临床和最终的商业用途。具体目标3。用鞭毛蛋白/Y进行GLP毒理学研究。鼠疫疫苗。一旦该工艺得到优化，将准备一个工程批次来研究其在GLP条件下的小鼠模型的安全性和免疫原性。该批号还将用于特性研究，并在临床生产准备中启动正式的稳定性研究。具体目标产生鞭毛/Y符合良好生产规范（GMP）条件的鼠疫疫苗。在特定目标2中制定的方案将用于在GMP条件下生产克数量的疫苗。这些目标的完成将为临床评价粘膜鞭毛蛋白/Y提供非常坚实的基础。人类鼠疫疫苗

DESCRIPTION (provided by applicant): Gram-negative flagellin is an extraordinarily potent activator of innate and adaptive immune responses. Our work as well as that of others has established that flagellin is a potent adjuvant in mice. We have also found that flagellin is a potent adjuvant in non-human primates. Flagellin possesses features that make it an excellent adjuvant including: potency; mucosally active; effectiveness in flagellin immune hosts; ease of production in bacteria; and 5) the ability to insert antigen sequences in the hypervariable region of the protein without loss of flagellin bioactivity. We have clearly established that flagellin is a highly effective mucosal adjuvant in the development of a strong, specific antibody response against Y. pestis antigens in mice and non-human primates and also promotes protection against a lethal respiratory challenge with Y. pestis. Since flagellin possesses tremendous potential as a mucosal adjuvant for use in humans, we propose the following aims that will advance the development of flagellin for human use: Specific Aim 1. To determine the efficacy of a flagellin/Y. pestis F1 and V fusion protein as a protective mucosal vaccine. We have generated a recombinant protein encoding flagellin and both the F1 and V antigens of Y. pestis and have established that this fusion protein retains flagellin biological activity. We propose to determine if the fusion protein promotes a robust anti-F1 and V IgG response in mice and cynomolgus monkeys and protection in mice against respiratory challenge with Y. pestis CO92. Specific Aim 2. To design and implement a Good Manufacturing Practice (GMP) compatible production process for a flagellin/Y. pestis vaccine that can be readily scaled for pre-clinical, clinical and ultimately commercial use. Specific Aim 3. To conduct GLP toxicology studies with the flagellin/Y. pestis vaccine. Once the process has been optimized, an engineering lot will be prepared to study its safety and immunogenicity under GLP conditions in a mouse model. This lot will also be used for characterization studies and to initiate formal stability studies in preparation for clinical manufacture. Specific Aim 4. To produce a flagellin/Y. pestis vaccine under Good Manufacturing Practices (GMP) conditions. The protocol developed in Specific Aim 2 will be used to produce gram quantities of the vaccine under GMP conditions. The completion of these aims will provide a very strong foundation for the clinical evaluation of the mucosal flagellin/Y. pestis vaccine in humans.