A Multicomponent Anthrax Vaccine using Phage T4 Display

描述（由申请人提供）：本申请的目标是开发一种易于施用的多组分炭疽疫苗，诱导持久的高抗体滴度，并提供对炭疽芽孢杆菌感染的保护。三种新型平台技术，无针皮肤贴片经皮免疫（TCI），噬菌体T4多组分展示，脂质体和乳剂佐剂配方，将对开发有效的炭疽疫苗产生影响。该研究将由两个互补的实验室进行，一个在T4噬菌体颗粒的遗传学和制造以及T4表面蛋白质表达方面具有高度技能，另一个在TCI、免疫原性脂质体、乳剂、疫苗配方和疫苗临床试验方面具有广泛经验，并与炭疽毒素生物学和生产方面处于领先地位的第三个实验室密切合作。所有三种炭疽毒素成分，保护性抗原（PA），致死因子（LF）和水肿因子（EF）的突变形式将以单独的蛋白或作为噬菌体T4的高抗原外衣壳蛋白（Hoc）的n端融合表达。带有n端六组氨酸标签的毒素- hoc融合蛋白将被大量纯化，并使用体外组装系统装载到噬菌体T4二十面体表面。TCI和肌内注射（IM）途径将可溶性蛋白或T4显示抗原与脂质体和乳剂佐剂配方结合，将使用小鼠模型并行评估，以产生保护性抗体滴度。免疫后的小鼠将接受(i)炭疽毒素、（ii）斯特恩菌株和（iii）艾姆斯菌株的攻击，以确定疫苗的效力。将在豚鼠模型中测试诱导小鼠抵抗毒性炭疽芽孢杆菌攻击的最佳组合。免疫应答将被描述为特征，并将对毒性炭疽菌株进行挑战实验，以选择诱导持久免疫的最佳疫苗产品。最有希望的炭疽候选疫苗将在非人类灵长类动物模型中进行测试，除了免疫反应的表征外，免疫的猕猴将受到炭疽芽孢杆菌雾化孢子的挑战。将评估保护的持续时间以及病理变化。

DESCRIPTION (provided by applicant): The goal of this application is to develop a multicomponent anthrax vaccine that can be easily administered, induces Iong-lasting high antibody titers, and provides protection against Bacillus anthracis infection. Three novel platform technologies, needle-free skin patch transcutaneous immunization (TCI), phage T4 multicomponent display, and liposome and emulsion adjuvant formulations, will be brought to bear on developing an efficacious anthrax vaccine. The research wilt be carried out by two complementary laboratories, one highly skilled in the genetics and manufacture of T4 bacteriophage particles and expression of proteins on the surface of T4, and the other with broad experience with TCI, immunogenic liposomes, emulsions, vaccine formulations, and vaccine clinical trials, in close collaboration with a third laboratory that is a leader in anthrax toxin biology and production. Mutant forms of all three anthrax toxin components, protective antigen (PA), lethal factor (LF) and edema factor (EF), will be expressed either as individual proteins or as N-terminal fusions of the highly antigenic outer capsid protein (Hoc) of bacteriophage T4. The toxin-Hoc fusion proteins with an N-terminal hexa-histidine tag will be purified in large quantities and loaded onto the phage T4 icosahedral surface using an in vitro assembly system. The TCI and intramuscular (IM) routes of delivery for combinations of soluble proteins or T4 displayed antigens with liposome and emulsion adjuvant formulations will be evaluated in parallel tracks, using the mouse model, for generation of protective antibody titers. The immunized mice will be challenged with (i) anthrax toxin, (ii) Sterne strain, and (iii) Ames strain, to determine the efficacy of the vaccines. The best combinations that induce protection in mice against challenge with virulent Bacillus anthracis will be tested in a guinea pig model. Immune responses will be characterized, and challenge experiments with the virulent anthrax strain will be performed to select the best vaccine product(s) that induce long-lasting immunity. The most promising anthrax vaccine candidates will be tested in a nonhuman primate model, in addition to characterization of immune responses, the immunized macaques will be challenged with the aerosolized spores of Bacillus anthracis. The duration of protection as well as pathological changes will be assessed.