Structure and immunogenicity of novel trimeric HIV-1 Env immunogens

需要一种疫苗来阻止HIV-1/艾滋病的流行，然而， 仍然是一个巨大的挑战，需要新的战略。多种方式稳定整体 HIV-1包膜的细胞外结构域（Env）已经导致大大改进的疫苗设计，例如，的 开发的SOSIP三聚体，其中，除了提供了一个完善的结构理解， Env三聚体被认为是一种有前途的免疫原，因为它含有所有已知的脆弱位点， bnAb靶向的Env三聚体。然而，基于SOSIP的免疫原只取得了有限的成功 这是由于不需要的分散表位位点将Ab应答转移到菌株特异性应答。与整个环境不同 细胞外结构域的方法，我们一直在使用分而治之的策略，从个人开始， Env三聚体的结构域，以开发具有免疫- 将Ab应答集中于选定的Env结构域和表位，并避免将Ab诱导至不需要的 表位区。我们从V1 V2结构域开始，不仅因为其结构域结构空间上位于Env 在RV 144试验中，apex及其Abs与感染风险呈负相关，已经开发了一组 - 通过在三聚体非HIV蛋白上支架化gp 120的V1 V2的三聚体V1 V2结构域免疫原，和 证明它们可以诱导V1 V2聚焦的Ab应答。此外，进一步的设计工作也导致了 在单链串联V1 V2三聚体中显示与三聚体特异性bnAb的抗原反应性。我们假设 这样的V1 V2-结构域免疫原可以被改进以呈现天然Env顶点构型， 结构表征和工程。为了从Env顶点扩展，我们设计了一个“重新核心” Env三聚体免疫原，通过用一个融合蛋白替代融合前三聚体结构中的gp 120内部结构域和gp 41， 稳定的三聚体非HIV支架蛋白;抗原性试验证明该分子含有所有关键bnAb 三聚体Env顶端的结合位点和EM可视化显示其具有良好形成的三聚体 配置.这种新的构建体提供了一个起点，以开发一种三聚体免疫原， Env三聚体的脆弱位点，其可以成为SOSIP三聚体的有吸引力的替代物， 更多地关注最有前途的bnAb表位。本R 01项目的目标是在结构上表征 这些合理设计的免疫原，并进一步应用我们的基于结构的平台来开发新的 因此，它们可以作为靶向天然三聚体构型的有效免疫原。我们 有三个目标。1)表征和改进模拟天然V1 V2-支架的三聚体V1 V2-支架构建体， Env三聚体顶端的融合前构象。2)开发具有密钥的重新核心的Env三聚体免疫原 bnAb位点。3)在动物中测试这些精制免疫原的免疫原性。在完成这项工作后， 项目，我们的新的三聚体免疫原将充分表征和选择的天然样免疫原可以 然后在疫苗开发和NHP挑战研究的管道中向前推进。

A vaccine is needed to stop the HIV-1/AIDS pandemic, however, the development of a protective vaccine remains a great challenge and requires novel strategies. Various approaches to stabilize the whole extracellular domain of the HIV-1 envelope (Env) has led to greatly improved vaccine designs, e.g., the development of the SOSIP trimer, which, in addition to having provided a refined structural understanding of the Env trimer, is considered as a promising immunogen because it harbors all the known vulnerable sites on the Env trimer targeted by bnAbs. However, SOSIP-based immunogens have achieved only limited success due to unwanted distracting epitope sites that divert Ab responses to strain-specific ones. Unlike whole Env extracellular domain approaches, we have been using a divide-and-conquer strategy by starting with individual domains of the Env trimer to develop domain-specific immunogens which have the advantage of immune- focusing Ab responses to selected Env domains and epitopes and of avoiding induction of Abs to unwanted epitope regions. We started with the V1V2 domain, not only for its domain structure spatially located at the Env apex but also its Abs inversely correlate with the risk of infection in the RV144 trial, have developed a panel of trimeric V1V2 domain immunogens by scaffolding V1V2 of gp120 on trimeric non-HIV proteins, and demonstrated that they can induce V1V2-focused Ab responses. Moreover, further design efforts have resulted in a single chain tandem V1V2 trimer showing antigenic reactivity with trimer-specific bnAbs. We hypothesize that such V1V2-domain immunogens can be improved to present the native Env apex configuration by further structural characterization and engineering. To expand from the Env apex, we have then designed a ‘re-cored’ Env trimer immunogen by replacing the gp120 inner domain and gp41 in the prefusion trimer structure with a stable trimeric non-HIV scaffold protein; antigenicity tests demonstrated that this molecule harbors all key bnAb binding sites of the trimeric Env apex and EM visualization showed it to have a well-formed trimeric configuration. This novel construct provides a starting point to develop a trimeric immunogen that carries key vulnerable sites of the Env trimer which could become an attractive alternative to the SOSIP trimers, offering greater focus on the most promising bnAb epitopes. The goal of this R01 project is to structurally characterize these rationally-designed immunogens and further apply our structure-based platform to develop new generations of them so that they serve as effective immunogens targeting native trimeric configurations. We have three Aims. 1) Characterize and refine the trimeric V1V2-scaffold constructs mimicking the native prefusion conformation of the Env trimer apex. 2) Develop the re-cored Env trimer immunogen harboring key bnAb sites. 3) Test the immunogenicity of these refined immunogens in animals. At the completion of this project, our novel trimeric immunogens will be fully characterized and selected native-like immunogens can then be move forward in the pipeline for vaccine development and for NHP challenge studies.