Utilization of the adjuvant effect of CRM197 protein to develop a trivalent protein-vaccine against Streptococcus pneumoniae infections

摘要 肺炎链球菌（Streptococcuspneumoniae，Spn）是一种主要的呼吸道病原体，可引起一系列非侵袭性（耳炎） 中膜、鼻窦炎、肺炎）以及侵袭性（败血症、脑膜炎）疾病。尽管总体上有所下降， 现有疫苗（Spn/肺炎球菌结合疫苗：PCV）引起的Spn疾病负担， spn疾病继续发生，仍然是一个重大的医疗问题。由于新的、 替代血清型（ST），旨在预防Spn疾病的Spn蛋白疫苗， 独立于类型的方式被认为是最好的解决方案。然而，迫切需要 开发能够增强针对Spn的抗体应答及其生物学功能的新型佐剂 蛋白质抗原，因为这些疫苗的目标人群是儿童、老年人和免疫功能低下的人。 个体 我们开发了一种三价蛋白疫苗，涉及两种众所周知的Spn疫苗候选物，N-末端 肺炎球菌表面蛋白A（PspA）和肺炎球菌溶血素结构域4（PlyD4），与白喉基因融合 选择N-PspA和PlyD4的目的是基于 它们在具有可证明的抗Spn定殖效力的疫苗接种动物模型中的先前表征 和败血症因此，CRM197与经证实的Spn候选物的基因融合将有助于比较CRM197驱动的Spn基因。 利用已发表的工作增强免疫原性和保护效力，并扩大平台， 包括其他有希望候选疫苗。 脱毒抗原CRM197（CRM197-N-PspA-PlyD4）。 我们的初步数据显示， CRM197显著增强（数倍）针对N-末端PspA的抗体滴度。另夕h 基因融合（三价疫苗）导致抗血清与Spn细菌的结合显著增加，这是一种测量方法。 抗体的功能。此外，用三价抗血清孵育Spn细菌显著降低了 Spn粘附于A549细胞。 基于我们的初步数据，我们假设"CRM197的基因融合增强了记忆性T细胞 对Spn抗原的应答，导致增强的抗体功能和针对Spn定殖的保护 疾病”。这一假设将在两个结构化的目标，将建立CRM 197的作用， 增强Spn抗原、N-PspA和PlyD4的免疫原性和保护效力。的调查结果 拟议的工作可能会导致更广泛地利用CRM 197的佐剂作用，以增强免疫力。 对Spn以及非Spn蛋白抗原的应答，导致更稳健的T细胞依赖性免疫 反应和保护作用。

Abstract Streptococcus pneumoniae (Spn) is a major respiratory pathogen that causes a spectrum of non-invasive (otitis media, sinusitis, pneumonia) as well as invasive (sepsis, meningitis) diseases. Despite a decline in the overall Spn disease burden caused by currently available vaccines (Spn/pneumococcal conjugate vaccines: PCVs), Spn diseases continue to occur and remain a significant medical problem. Due to the emergence of new, replacement serotypes (STs), Spn protein-based vaccines that aim to protect against Spn diseases in a serotype-independent manner are considered to be the best solution. However, there is a pressing need to develop novel adjuvants that can enhance antibody response and its biological function directed against Spn proteins antigens, given the target populations of these vaccines are children, elderly, and immunocompromised individuals. We developed a trivalent protein vaccine involving two well-known Spn vaccine candidates, N-terminal pneumococcal surface protein A (PspA) and domain 4 of pneumolysin (PlyD4), genetically fused with diphtheria The purpose of choosing N-PspA and PlyD4 is based on their prior characterization in animal models of vaccination with demonstratable efficacy against Spn colonization and sepsis. Therefore, genetic fusion of CRM197 to proven Spn candidates will help compare the CRM197 driven enhancement of immunogenicity and protective efficacy with published work and to expand the platform to include other promising vaccine candidates subsequently. detoxified antigen CRM197 (CRM197-N-PspA-PlyD4). Our preliminary data shows that the genetic fusion of CRM197 significantly enhanced (several folds) the antibody titers against N-terminal PspA. Additionally, the genetic fusion (trivalent vaccine) led to a significant increase in the binding of antisera to Spn bacteria, a measure of antibody functionality. Furthermore, the incubation of Spn bacteria with trivalent antisera significantly reduced Spn adherence to A549 cells. Based on our preliminary data, we hypothesize that “Genetic fusion of CRM197 enhances the memory T-cell response to Spn antigens, leading to enhanced antibody functionality and protection against Spn colonization and disease.” This hypothesis will be tested in two structured aims that will establish the role of CRM197 in enhancing the immunogenic and protective efficacy of Spn antigens, N-PspA and PlyD4. The findings of the proposed work could lead to a broader utilization of the adjuvant effect of CRM197 in enhancing the immune response to Spn as well as non-Spn protein antigens, leading to a more robust T-cell dependent immune response and protective effect.