Enhancement of protective efficacy of Coccidioides vaccines by adjuvants

 描述(由申请人提供)：这项提案的重点是在人源化的球孢子菌病小鼠模型中创建和临床前评估一种新型疫苗，该疫苗包含重组、嵌合多肽抗原(RCPA)和免疫原性最强的类型的葡聚糖颗粒(GP)作为佐剂和递送平台。初步数据显示，疫苗对球虫感染的免疫依赖于MyD88和CARD9介导的Th17和Th1反应。此外，Th17免疫是不可或缺的。GP是一种中空的、高纯度的酵母细胞壁，主要由1，3-葡聚糖组成，已被证明能刺激Th17和Th1免疫。初步研究表明，用“负载”抗原的GP免疫小鼠后，T细胞具有强大和持久的适应性反应。核心假设是，当一个多价抗原(RCPA)整合到最具免疫原性的GP形式，当与一个配体(S)结合时，引起Toll样受体(TLR)或C型凝集素受体(CLR)介导的免疫反应，将通过激发对球虫感染的强大和持久的Th17和/或Th1免疫来增强疫苗效力。目的1.评价RCPA的免疫原性，优化抗原设计。含有多个抗原的多价球虫疫苗在刺激保护性免疫方面比单一抗原更有效。融合了球虫4种保护性抗原的RCPA已成功表达和纯化。这些单独的抗原和RCPA将使用表达人类MHC II的HLA-DR4转基因小鼠和从皮试阳性和阴性志愿者中分离的外周血单核细胞对临床批准的球虫抗原的免疫原性进行评估。RCPA的设计将得到优化，只包含4种选定抗原中能够刺激所有疫苗免疫的免疫原性多肽，包括激活产生IL-17A和干扰素-的T细胞和产生抗体。目的2.寻找一种有效的佐剂配体(S)与改良的GPS结合，以增强RCPA疫苗对球虫感染的免疫力。优化后的RCPA将被加载到4种不同类型的葡聚糖/甘露聚糖/甲壳素颗粒(GxP)中，并在人源化的HLA-DR4转基因小鼠中单独测试保护效果。葡聚糖颗粒是一种正在进行临床试验的实验性疫苗佐剂。最具保护性的GP将与选定的模式识别受体配体(S)共同负载，以增强T细胞免疫和保护效果。将分析赋予对球孢子虫感染的抵抗力的疫苗(S)，以剖析保护的相关性，包括T细胞和抗原提呈细胞在免疫反应的传出和传入肢体中的细胞因子表达，该疫苗经鼻腔攻击具有潜在致死量的球孢子虫孢子。该项目完成后，将产生一种新的领先多价疫苗候选疫苗，用于进入临床试验，以评估其对球孢子菌病的安全性和保护效果。

 DESCRIPTION (provided by applicant): This proposal is focused on the creation and preclinical evaluation of a novel vaccine containing a recombinant, chimeric polypeptide antigen (rCPA) combined with the most immunogenic type of glucan particles (GPs) as an adjuvant and delivery platform in a humanized murine model of coccidioidomycosis. Preliminary data have revealed that vaccine immunity to Coccidioides infection is dependent on both MyD88- and Card9-mediated Th17 and Th1 responses. Furthermore, Th17 immunity is indispensable. GPs are hollow, highly purified yeast cell walls composed predominantly of ß-1,3-glucan that has been shown to stimulate Th17 and Th1 immunity. Preliminary studies demonstrate robust and long-lasting adaptive T-cell responses following immunization of mice with GPs "loaded" with antigens. The central hypothesis is that a multivalent antigen (rCPA) incorporated into the most immunogenic form of GPs, when combined with a ligand(s) that elicits Toll- like receptor (TLR)- or C-type lectin receptor (CLR)-mediated immune responses will augment vaccine efficacy by stimulating robust and durable Th17 and/or Th1 immunity to Coccidioides infection. The two Specific Aims are: Aim 1. To evaluate immunogenicity of rCPA and to optimize antigen design. Multivalent Coccidioides vaccines containing multiple antigens have been shown to be more effective in stimulation of protective immunity than individual antigens. The proposed rCPA that incorporates 4 protective antigens of Coccidioides has been successfully expressed and purified. The immunogenicity of these individual antigens and rCPA will be evaluated using human MHC II-expressing HLA-DR4 transgenic mice and peripheral blood monocytic cells isolated from skin test-positive and -negative volunteers to a clinically-approved antigen of Coccidioides. The design of the rCPA will be optimized to contain only the immunogenic polypeptides among the 4 selected antigens that can stimulate all arms of vaccine immunity including activation of IL-17A- and IFN--producing T cells and antibody production. Aim 2. To identify an effective adjuvant ligand(s) combined with modified GPs to enhance rCPA vaccine immunity to Coccidioides infection. The optimized rCPA will be loaded into 4 different types of glucan/mannan/chitin particles (GxP) and tested individually for protective efficacy in humanized HLA-DR4 transgenic mice. Glucan particles are an experimental vaccine adjuvant under clinical trials. The most protective type of GPs will be co-loaded with a selected ligand(s) for pattern recognition receptors to augment T-cell immunity and protective efficacy. The vaccine(s) that confer resistance to Coccidioides infection will be analyzed to dissect correlates of protection including cytokine expression by T cells and antigen-presenting cells during the efferent and afferent limbs of immune responses after an intranasal challenge with a potentially lethal dose of Coccidioides spores. Upon completion of this project, a novel lead multivalent vaccine candidate will be generated for advancement into clinical trials for assessment of its safety and protective efficacy against coccidioidomycosis.