Multivalent Plague, Anthrax Vaccines Using Bacteriophage T4 Display

描述(申请人提供)：我们假设T4噬菌体上的抗原体外组装(AGS)可以作为构建下一代多价生物防御疫苗的通用平台。这项提议的主要目标是检验这一假设，并开发三价鼠疫和多价鼠疫-炭疽疫苗，以提供对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保护。我们已经开发了一个明确的体外系统，可以在噬菌体T4纳米颗粒(120×86 nm)上以高密度排列大量的外源AGS。将长达90 kDa的炭疽全长毒素与两个非必需的噬菌体T4外衣壳蛋白Soc(870个拷贝)和Ad(高抗原性外衣壳蛋白155个拷贝)在大肠杆菌中高效表达，纯化至均一，并通过体外结合展示于HOC_SOC_衣壳蛋白上。多个AGS和大的杂寡聚复合体可以显示到饱和，拷贝数可以控制。空间暴露和对称排列的T4-RPA颗粒具有高度的免疫原性，不需要任何佐剂，可诱导出强大的PA特异性和致死性毒素中和抗体(Ab)效价，对100 LD50炭疽杆菌孢子攻击给予完全保护。最好的疫苗配方正在用恒河猴进行临床前试验。结合我们的生化、分子遗传学、结构和气溶胶挑战专业知识，我们设计了一种新型三价T4鼠疫疫苗，由三个耶尔森氏菌AGS、衣壳抗原、CAF1、低钙反应V抗原、LcrV和耶尔森氏菌分泌复杂因子YscF组成。YscF是III型分泌系统的一个高度保守的针形亚单位，对所有毒力强的耶尔森菌物种都是必不可少的，纳入YscF有望产生一种优质和广泛有效的三价鼠疫疫苗。将优化衣壳包围型鼠疫AGS的结构配置和拷贝数。T4-鼠疫AGS、佐剂、传递途径(包括皮肤贴片)的组合将接受抗体和细胞免疫反应测试，并对最致命的鼠疫菌株之一--鼠疫耶尔森氏菌CO92的攻击提供保护。保护效果将在一种新的布朗挪威大鼠气溶胶挑战模型中进行评估。最好的T4-炭疽疫苗和T4-鼠疫疫苗将结合在一起，创造出一种新的多价炭疽-鼠疫疫苗。该疫苗对吸入性炭疽和肺鼠疫的预防效果将在恒河猴气雾剂攻击模型中进行测试。

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.