Evaluation of human adenovirus-based vectors for immunization in consideration of seroprevalence, immunogenicity and effects of pre-existing immunity

腺病毒载体是实验性疫苗开发的常用工具；然而，如果载体是基于高流行的人腺病毒类型(如Ad5)，那么它们在人类中的应用可能会有问题。因此，人们已经做出了许多努力来开发替代载体。在初步实验中，我们可以证明对腺病毒载体的预先存在的免疫可以削弱转基因特异性CD8+T细胞反应，但它也可以对转基因特异性抗体反应的诱导产生积极的影响。在初步实验中，我们已经测试了一个学生队列的245份人血清对30种人腺病毒类型的抗体反应性，并与Ad5进行了直接比较。我们能够识别出与Ad5相比明显不那么流行的一些腺病毒类型。在拟议的项目中，我们希望评估我们根据学生队列中较低的血清阳性率选择的10种人腺病毒类型，以确定它们作为疫苗载体的潜力。在第一个项目部分，我们将在小鼠实验中测定针对人腺病毒类型的抗体和CD8+T细胞反应，包括交叉反应免疫反应的分析。在进一步的实验中，我们将构建编码Friend病毒Gag的载体，并在体外测试它们刺激CD8+T细胞增殖和激活抗原提呈细胞的效果。最好的载体将在体内用于CD8+T细胞反应的评估和对Friend病毒攻击感染的保护。此外，我们将在实验中解决基因组刺激性TLR基序内容或衣壳蛋白正在决定腺病毒类型的免疫原性的假设。在项目的最后部分，我们将分析免疫前介导的抗体应答增强的机制，并建立队列样腺病毒免疫，以评估使用新载体在现有免疫的现实条件下优化免疫方案的有效性。我们期待该项目的结果将为开发新的有效的基于腺病毒的疫苗开发提供重要的新见解。

Adenovirus-based vectors are popular tools for experimental vaccine development; however, their application in humans can be problematic if vectors are based, as they historically were, on high-prevalent human adenovirus types such as Ad5. Therefore, many efforts have been made to develop alternative vectors. In preliminary experiments, we could show that pre-existing immunity against the adenovirus-based vector can impair the transgene-specific CD8+ T cell response, but it can also have a positive impact on the induction of transgene-specific antibody responses. In preliminary experiments, we have already tested the antibody reactivity of 245 human sera of a student cohort against 30 human adenovirus types in direct comparison to Ad5. We were able to identify a number of adenovirus types that were significantly less prevalent in comparison to Ad5. In the proposed project, we would like to evaluate 10 human adenovirus types that we selected based on their low seroprevalence in the student cohort for their potential as vaccine vectors. In the first project part, we will determine antibody and CD8+ T cell responses against the human adenovirus types in mouse experiments, including an analysis of cross-reactive immune responses. In further experiments, we will construct vectors encoding Friend virus Gag and test them in vitro for their efficacy in stimulating CD8+ T cell proliferation and activating antigen-presenting cells. The best vectors will then be used in vivo for the evaluation of CD8+ T cell responses and protection from Friend virus challenge infection. Furthermore, we will experimentally address the hypotheses that the genomic stimulatory TLR motif content or capsid proteins are determining an adenovirus type’s immunogenicity. In the last part of the proposed project, we will analyse the mechanisms underlying the pre-immunity-mediated enhancement of antibody responses, and we will establish a cohort-like adenovirus immunity to evaluate the effectiveness of an optimized immunization protocol using the new vectors under realistic conditions of pre-existing immunity.We expect that the results of this proposed project will yield important new insights for the development of new effective adenovirus-based vectors for vaccine development.