Capsid-incorporation of HIV antigens as a novel adenovirus HIV vaccine approach

描述(申请人提供)：尽管Ad载体在疫苗应用中具有许多潜在的优势，但当前的Ad疫苗的充分利用可能会受到宿主抗载体免疫反应的限制。具体地说，抗Ad体液免疫取消了后续给药的有效性，混淆了编码的转基因的表达，从而实际上限制了可能通过Booster效应获得的收益。为了利用Ad载体固有的抗原性，我们开发了一种基于免疫抗原表位直接掺入Ad衣壳的疫苗接种方法。这一新的范例是基于Ad将抗原呈现为衣壳的一部分，而不是编码的转基因。将免疫原肽结合到Ad衣壳中具有潜在的优势。最值得注意的是，通过外源途径处理衣壳结合的抗原应该会产生类似于天然Ad衣壳蛋白引起的强烈体液反应。此外，由于抗Ad衣壳蛋白应答通过重复给药而增强，针对作为Ad衣壳蛋白一部分的抗原表位的免疫应答也应该通过反复给药来增强，从而允许加强免疫。这些考虑表明，这种新的衣壳结合抗原方法可能为实现基于腺病毒的疫苗策略提供令人兴奋的潜力，从而绕过与腺病毒载体相关的主要限制。 实现这一方法的关键是确定腺病毒衣壳中抗原的最佳构型。为此，我们建立了几项关键技术，使我们能够实现我们的目标。特别是，我们已经开发出将异源多肽表位整合到主要的Ad衣壳蛋白六邻体蛋白的表面暴露区域中的方法。我们已经开始确定预测这些结构域在六角体中的功能效用的大小和结构因素。此外，我们还开发了应用低温电子显微镜(CryoEM)单粒子重建方法的方法，使我们能够以前所未有的亚纳米分辨率探索衣壳结合多肽的定位。基于这些技术，我们将能够建立衣壳内抗原位置/可获得性与疫苗效力之间的关键关联。 在这些已建立的可行性的基础上，我们假设可以用新型衣壳结合的抗原创建Ad载体，在动物模型中用作抗HIV的疫苗制剂。将应用低温EM引导的衣壳设计来开发具有最佳抗HIV免疫的优化载体。我们预计，我们提议的结构研究将为体外分析和生物学读数提供补充信息，从而使我们能够了解所结合的HIV表位的功能决定因素。该项目将设计新的创新方法来创造艾滋病毒疫苗，希望防止艾滋病毒疾病的传播。

DESCRIPTION (provided by applicant): Despite the many potential advantages of Ad vectors for vaccine application, full utility of current Ad vaccines may be limited by the host anti-vector immune response. Specifically, the anti-Ad humoral immunity abrogates the effectiveness of subsequent administrations of the Ad vector, confounding expression of the encoded transgene, and thus practically restricting the gains that might be accrued via booster effect. In order to exploit the inherent antigenicity of the Ad vector we have developed a vaccination approach based on incorporation of the immunizing antigen epitope directly into the Ad capsid. This novel paradigm is based upon Ad presenting the antigen as a component of the capsid rather than an encoded transgene. Incorporation of immunogenic peptides into the Ad capsid offers potential advantages. Most noteworthy, the processing of the capsid incorporated antigen via the exogenous pathway should result in a strong humoral response akin to the response provoked by native Ad capsid proteins. In addition, since anti-Ad capsid responses are augmented by repeated vector administration, immune responses against antigenic epitopes that are part of the Ad capsid should be augmented by repeated administration as well, thus allowing boosting. These considerations suggest that this novel capsid-incorporated antigen approach may offer exciting potentials to realize Ad-based vaccine strategies that circumvent the major limitations associated with Ad vectors. Critical to the realization of this approach is to define the optimal configuration of antigen in the adenoviral capsid context. To this end, we have established several key technologies that will enable us to reach our goal. In particular, we have developed the means to incorporate heterologous peptide epitopes within the surface-exposed domains of the major Ad capsid protein hexon. We have begun to determine the size and structural factors that predicate functional utility of these domains in the hexon. In addition, we have developed the means to apply cryoelectron microscopy (cryoEM) single particle reconstruction methods to allow us to explore the capsid-incorporated peptide localization with unprecedented, subnanometer resolution. Based on these technologies, we will be able to establish the critical correlates between antigen locale/accessibility within the capsid context and vaccine efficacy. On the basis of these established feasibilities, we hypothesize that Ad vectors can be created with novel capsid-incorporated antigens that can serve as vaccine agents against HIV in animal models. CryoEM-guided capsid design will be applied to develop an optimized vector with optimal anti-HIV immunization. We envision that our proposed structural studies will provide complementary information to in vitro assays and biological readouts and thereby will enable us to understand the functional determinants of incorporated HIV epitopes. This project will design new and innovative methodologies to create HIV vaccines, in hopes of preventing the spread of HIV disease.