Engineered Nanoparticles for Protective Subunit Vaccine Delivery and Discovery

项目总结 结核分枝杆菌是一种引起结核病的细菌病原体，结核病是一种折磨人的疾病。 20亿人，每年导致150万人死亡。全球结核病的负担是巨大的，而且还在不断增长， 而一种有效的疫苗将为这个问题提供最佳的长期解决方案。然而，这样的疫苗 仍然不存在。阻碍有效结核病疫苗开发的主要障碍包括 对哪些抗原决定簇赋予保护性免疫的认识不完全，而且缺乏 能够激发记忆CD8+和CD4+T细胞反应的疫苗接种策略 并清除感染。这个项目的目标是设计和评估一种创新的纳米颗粒(Np)。 经改造的疫苗可诱导强健持久的肺CD8+和CD4+Th1记忆T细胞反应 抗结核分枝杆菌抗原。NPS将被开发和优化，以实现Mtb表位和5‘’的双重传递 胞质模式识别受体(PRR)激动剂三磷酸化RNA(5‘PPP-RNA) 酸诱导基因1(RIG-I)，具有作为疫苗佐剂的巨大潜力。具体来说，我们 建议利用NP疫苗的独特递送能力来增强肺Th1和CD8+T细胞 对已建立的和临床相关的Mtb MHC I类和II类限制性表位以及 我们团队最近发现的自然加工的I类限制性表位。我们集结了一支强大的 拥有药物输送、疫苗设计和开发、RNA工程、 表位发现和结核分枝杆菌感染的小鼠模型。因此，我们处于理想的位置来完成我们的 目的通过以下具体目标：1)研制一种诱导肺内CD8+的纳米颗粒疫苗 和Th1记忆T细胞对MTB亚单位抗原的反应；2)评价其免疫原性和保护性 天然处理的结核分枝杆菌表位的潜力。在目标1中，我们将研究鼻腔内NP的作用。 免疫对肺抗原/佐剂递送和细胞内摄取的幅度和动力学的影响 阐明5‘PPP-RNA的免疫刺激佐剂作用，并严格表征肺和 系统的CD4+和CD8+T细胞对结核分枝杆菌抗原的反应。在目标2中，我们将使用NPs来交付新的MTB I类限制性表位，单独或与已建立的免疫优势II类Mtb结合 表位，表征局部和系统的T细胞反应，并评估疫苗预防疾病的能力 肺结核分枝杆菌挑战。该项目将产生一种新的疫苗技术，以增强粘膜细胞 对亚单位疫苗的免疫，将扩大疫苗佐剂的武器库，并将提供一种工具， 使免疫原性和保护性Mtb表位的发现成为可能。通过结合表位发现努力 凭借合理设计的交付系统，成功完成拟议的研究将建立一个 合理设计T细胞靶向疫苗的新的和潜在的变革性范例。

PROJECT SUMMARY Mycobacterium tuberculosis (Mtb) is a bacterial pathogen that causes tuberculosis (TB), a disease that afflicts 2 billion people and results in 1.5 million deaths each year. The global burden of TB is enormous and growing, and an effective vaccine would provide the best long-term solution to this problem. However, such a vaccine still does not exist. Major barriers that have hindered the development of an effective TB vaccine include an incomplete understanding of which antigenic determinants confer protective immunity and a dearth of vaccination strategies capable of eliciting memory CD8+ and CD4+ T cell responses that can rapidly respond to and clear infection. The objective of this project is to design and evaluate an innovative nanoparticle (NP) vaccine engineered to elicit robust and durable pulmonary CD8+ and CD4+ Th1 memory T cell responses against Mtb antigens. NPs will be developed and optimized for dual-delivery of Mtb epitopes and 5' triphosphorylated RNA (5'ppp-RNA), an agonist of the cytosolic pattern recognition receptor (PRR) retinoic acid-inducible gene 1 (RIG-I) that has immense untapped potential as a vaccine adjuvant. Specifically, we propose to harness the unique delivery capabilities of NP vaccines to enhance pulmonary Th1 and CD8+ T cell responses to established and clinically relevant Mtb MHC class I- and class II-restricted epitopes as well as naturally processed class I-restricted epitopes recently discovered by our team. We have assembled a strong multidisciplinary team with expertise in drug delivery, vaccine design and development, RNA engineering, epitope discovery, and mouse models of Mtb infection. Therefore, we are ideally positioned to accomplish our objective through the following Specific Aims: 1) Develop a nanoparticle vaccine that elicits lung-resident CD8+ and Th1 memory T cell responses to Mtb subunit antigens; 2) Evaluate the immunogenicity and protective potential of naturally processed Mtb epitopes. In Aim 1, we will investigate the effect of intranasal NP immunization on the magnitude and kinetics of pulmonary antigen/adjuvant delivery and intracellular uptake, elucidate the immunostimulatory adjuvant effects of 5'ppp-RNA, and rigorously characterize pulmonary and systemic CD4+ and CD8+ T cell responses to Mtb antigens. In Aim 2, we will use NPs to deliver novel Mtb class I-restricted epitopes, both alone and in combination with an established immunodominant class II Mtb epitope, characterize local and systemic T cell responses, and assess the ability of vaccines to protect against pulmonary Mtb challenge. This project will result in a new vaccine technology for enhancing mucosal cellular immunity to subunit vaccines, will expand the armamentarium of vaccine adjuvants, and will provide a tool that empowers the discovery of immunogenic and protective Mtb epitopes. By combining epitope discovery efforts with rationally engineered delivery systems, successful completion of the proposed research will establish a new and potentially transformative paradigm for the rational design of T cell-targeted vaccines.