Developing a Multivalent Subunit Particle Vaccine Against Tuberculosis

项目概要 可能需要一种有效的疫苗来减轻全球结核病（TB）负担。开发一 一直很困难，部分原因是结核分枝杆菌 (Mtb) 能够长期驻留 在宿主抗原 (Ag) 呈递细胞 (APC) 内，它表达多种 Ag，其中许多具有 已成为疫苗研发的目标。重组抗原一直是众多疫苗的焦点， 然而，临床测试的结果并不理想。此 R61/R33 提案响应计划 公告 RFA-AI-21-007：“结核病疫苗发现的创新”将利用下一个- 产生，疫苗佐剂系统，诱导重组抗原的无缝颗粒形成，以评估 多价结核分枝杆菌疫苗。将提出一种新的免疫范式，基于组合（通过简单的 混合）充分表征的可溶性蛋白 Mtb Ag 与疫苗佐剂，诱导自发性 纳米脂质体-Ag颗粒化。含有少量钴卟啉-磷脂的脂质体 (CoPoP) 通过将 his 标签自发插入双层来与带有 his 标签的 Ag 结合。这种方法给出 产生在生物介质中稳定的快速颗粒化。重组 Ag 与 CoPoP 简单混合 疫苗接种时的脂质体（无需进一步纯化）将完全转化以展示在表面上 ~100 nm 颗粒。 CoPoP最近完成I期并进入II期临床测试作为关键 COVID-19 疫苗的成分 (NCT04783311)。 CoPoP 还利用短时间诱导有效的细胞反应 小鼠中纳克肽剂量的 MHC-I 限制性肽免疫原；降低多个数量级 与传统疫苗佐剂的 Ag 剂量相比的剂量。初步数据表明 用已建立的 Mtb Ags Ag85A、CFP10、ESAT-6、HSP-X 和 Mpt64 修饰的多价颗粒 诱导小鼠的保护性细胞免疫反应。基本假设是多价、粒子 基于 Mtb Ags 将诱导比卡介苗更有效的强效保护性免疫反应 – Guérin (BCG) 在多种结核感染小动物模型中的应用。 R61阶段的提案将使用鼠标 结核病感染模型： (1) 开发特征明确的多价 Mtb Ag 颗粒； (2) 评估粒子如何 参数影响功能免疫原性； (3)比较鼻内和肌内注射的疗效 颗粒管理。基于 A) 提出了雄心勃勃且定量的 R61-R33 过渡里程碑 B) 多价银可生成具有良好储存稳定性的特征良好的颗粒，提供卓越的性能 相对于最好的单一银颗粒的保护作用，C) 优于小鼠中的传统 BCG 免疫。 该项目的 R33 阶段将涉及 (4) 在豚鼠模型中评估疫苗，以及 (5) 开发疫苗 并评估冻干热稳定疫苗的功效和毒性。

PROJECT SUMMARY An effective vaccine is likely required to reduce the global burden of tuberculosis (TB). Development of one has been difficult, in part due to the ability for Mycobacterium tuberculosis (Mtb) to reside for long periods within host antigen (Ag) presenting cells (APCs), where it expresses a multitude of Ags, many of which have been targeted for vaccine development. Recombinant Ags have been the focus of numerous vaccines, however outcomes to clinical testing have been less than ideal. This R61/R33 proposal in response to program announcement RFA-AI-21-007: “Innovation for Tuberculosis Vaccine Discovery” will leverage a next- generation, vaccine adjuvant system that induces seamless particle-formation of recombinant Ags to assess a multivalent Mtb vaccine. A new immunization paradigm will be advanced, based on combining (via simple mixing) well-characterized soluble protein Mtb Ags with a vaccine adjuvant that induces spontaneous nanoliposome-Ag particleization. Liposomes that contain small amounts of cobalt porphyrin-phospholipid (CoPoP) bind to his-tagged Ags via spontaneous insertion of the his-tag into the bilayer. This approach gives rise to rapid particleization that is stable in biological media. Recombinant Ags are simply admixed with CoPoP liposomes at the time of vaccination (without further purification) to be fully converted for display on the surface of ~100 nm particles. CoPoP recently completed phase I and entered phase II clinical testing as a key component of a COVID-19 vaccine (NCT04783311). CoPoP also induces potent cellular responses using short MHC-I restricted peptide immunogens at nanogram peptide dosing in mice; multiple orders of magnitude lower dosing compared to Ag doses with conventional vaccine adjuvants. Preliminary data demonstrates that multivalent particles decorated with the established Mtb Ags Ag85A, CFP10, ESAT-6, HSP-X, and Mpt64 induce protective cellular immune responses in mice. The underlying hypothesis is that multivalent, particle- based Mtb Ags will induce a potently protective immune responses more effective than Bacillus Calmette– Guérin (BCG) in multiple small animal models of TB infection. The R61 phase of the proposal will use mouse models of TB infection to: (1) Develop well-characterized, multivalent Mtb Ag particles; (2) Assess how particle parameters impact functional immunogenicity; and (3) Compare the efficacy of intranasal and intramuscular particle administration. Ambitious and quantitative R61-R33 transitional milestones are proposed based on A) generating well-characterized particles with storage stability with B) multivalent Ags providing superior protection relative to the best single Ag particles with C) superiority to conventional BCG immunization in mice. The R33 phase of the project will involve (4) assessing the vaccine in the guinea pig model, and (5) developing and assessing the efficacy and toxicity of a lyophilized thermostable version of the vaccine.