Multivalent Plague, Anthrax Vaccines Using Bacteriophage T4 Display

描述（由申请人提供）：我们假设抗原（Ags）在T4噬菌体上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。本提案的主要目标是验证这一假设，并开发三价鼠疫和多价鼠疫-炭疽疫苗，这些疫苗可以保护人们免受炭疽芽孢杆菌和鼠疫耶尔森菌这两种A类生物防御病原体的侵害。我们开发了一种明确的体外系统，用于在噬菌体T4纳米颗粒（120 x 86 nm）上高密度排列大型外源Ags。全长90kDa的炭疽毒素与两种非必需噬菌体T4外衣壳蛋白Soc（小外衣壳蛋白，870拷贝）和Hoc（高抗原性外衣壳蛋白，155拷贝）融合，在大肠杆菌中过表达，纯化至均质，通过体外结合在hoc_soc_衣壳上展示。多个ag和大的异寡聚配合物可以显示到饱和，拷贝数可以控制。该空间暴露和对称排列的T4-rPA颗粒不含任何佐剂，具有高度免疫原性，可激发强pa特异性和致命毒素中和抗体（Ab）滴度，可完全保护家兔抵御100 LD50炭疽芽孢杆菌Ames孢子的攻击。最好的疫苗配方正在恒河猴的临床前试验中进行测试。结合我们在生化、分子遗传学、结构和气溶胶挑战方面的专业知识，我们设计了一种新的三价t4 -鼠疫疫苗，该疫苗由三种耶尔森氏菌Ags组成，即荚膜Ag， Caf1，低钙反应V Ag， LcrV和耶尔森氏菌分泌复合物因子YscF。YscF是III型分泌系统中高度保守的针状亚基，对所有致病性耶尔森菌种都至关重要。纳入YscF有望产生一种优越且广泛有效的三价鼠疫疫苗。对衣壳结合鼠疫Ags的结构配置和拷贝数进行优化。将对t4 -鼠疫Ags、佐剂、给药途径（包括皮肤贴片）的组合进行Ab和细胞免疫应答以及对最致命鼠疫毒株之一鼠疫耶尔森氏菌荚膜菌株CO92攻击的保护作用的测试。保护效果将在一种新的褐挪威大鼠气溶胶攻击模型中进行评估。最好的t4 -炭疽疫苗和t4 -鼠疫疫苗将结合在一起，形成一种新的多价炭疽-鼠疫疫苗。该疫苗对吸入性炭疽和肺鼠疫的保护效果将在恒河猴气溶胶攻击模型中进行试验。

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.