Multivalent Plague, Anthrax Vaccines Using Bacteriophage T4 Display

描述（由申请人提供）：我们假设T4噬菌体上抗原（Ag）的体外组装可以作为构建下一代多价生物防御疫苗的通用平台。该提案的主要目标是测试这一假设，并开发三价鼠疫和多价炭疽疫苗，可以提供对炭疽杆菌和鼠疫耶尔森氏菌，两个A类生物防御病原体的保护。我们已经开发了一种确定的体外系统，以高密度在噬菌体T4纳米颗粒（120 × 86 nm）上排列大的外源Ag。全长炭疽毒素，大到90 kDa，融合到两个非必需的噬菌体T4外衣壳蛋白，Soc（小外衣壳蛋白; 870个拷贝）和Hoc（高抗原性外衣壳蛋白; 155个拷贝），在E. coli，纯化至均一，并通过体外结合展示在hoc_pepsin_ capsid上。多个Ag和大的异源寡聚复合物可以被展示至饱和，并且拷贝数可以被控制。空间暴露且对称排列的T4-rPA颗粒在没有任何佐剂的情况下具有高度免疫原性，可引发强烈的PA特异性和致命毒素中和抗体（Ab）滴度，为兔子提供针对100 LD 50 B的完全保护。炭疽艾姆斯孢子攻击。最好的疫苗配方正在用恒河猴进行临床前试验。结合我们的生物化学，分子遗传学，结构和气溶胶挑战的专业知识，战略的目的是开发一种新的三价T4鼠疫疫苗组成的三个耶尔森氏菌抗原，荚膜Ag，Caf 1，低钙反应V Ag，LcrV，和耶尔森氏菌分泌复合因子，YscF。YscF是所有致病耶尔森氏菌属物种所必需的III型分泌系统的高度保守的针形成亚基，预期包含YscF将产生上级且广泛有效的三价鼠疫疫苗。将优化captain结合的鼠疫Ag的结构分布和拷贝数。将检测T4鼠疫抗原、佐剂、给药途径（包括皮肤贴剂）的组合的抗体和细胞免疫应答以及对荚膜菌株Y攻毒的保护作用。鼠疫CO 92，最致命的鼠疫菌株之一。将在新的Brown Norway大鼠气溶胶激发模型中评估保护效力。最好的T4炭疽疫苗和T4鼠疫疫苗将被结合起来，以创造一种新的多价炭疽鼠疫疫苗。将在恒河猴气溶胶激发模型中检测该疫苗对吸入性炭疽和肺鼠疫的保护效力。

DESCRIPTION (provided by applicant): We hypothesize that in vitro assembly of antigens (Ags) on bacteriophage T4 can be developed as a common platform for construction of next generation multivalent biodefense vaccines. The major goal of this proposal is to test this hypothesis and develop trivalent plague and multivalent plague-anthrax vaccines that can confer protection against Bacillus anthracis and Yersinia pestis, two Category A biodefense pathogens. We have developed a defined in vitro system to array large foreign Ags at high density on phage T4 nanoparticle (120 x 86 nm). Full-length anthrax toxins, as large as 90kDa, are fused to the two non-essential phage T4 outer capsid proteins, Soc (small outer capsid protein; 870 copies) and Hoc (highly antigenic outer capsid protein; 155 copies), over-expressed in E. coli, purified to homogeneity, and displayed on hoc_soc_ capsid through in vitro binding. Multiple Ags and large hetero-oligomeric complexes can be displayed to saturation and the copy number can be controlled. The spatially exposed and symmetrically arrayed T4-rPA particles are highly immunogenic without any adjuvant, eliciting strong PA-specific and lethal toxin neutralizing Antibody (Ab) titers, conferring complete protection to rabbits against 100 LD50 B. anthracis Ames spore challenge. The best vaccine formulations are being tested in a preclinical trial using rhesus macaques. Combining our biochemical, molecular genetic, structural, and aerosol challenge expertise, strategies are designed to develop a novel trivalent T4-plague vaccine comprised of three Yersinia Ags, the capsular Ag, Caf1, the low calcium response V Ag, LcrV, and the Yersinia secretory complex factor, YscF. Inclusion of YscF, a highly conserved needle-forming subunit of type III secretion system that is essential to all virulent Yersinia species, is expected to generate a superior and broadly effective trivalent plague vaccine. The structural disposition and copy number of the capsid-bound plague Ags will be optimized. Combinations of T4-plague Ags, adjuvants, route of delivery (including skin patch), will be tested for Ab and cellular immune responses and protection against challenge with the capsular strain Y. pestis CO92, one of the most lethal plague strains. Efficacy of protection will be assessed in a new Brown Norway rat aerosol challenge model. The best T4-anthrax and T4-plague vaccines will be combined to create a novel multivalent anthrax-plague vaccine. The efficacy of this vaccine for protection against both inhalation anthrax and pneumonic plague will be tested in rhesus macaque aerosol challenge in model.