Structure and immunogenicity of novel trimeric HIV-1 Env immunogens

需要一种疫苗来阻止艾滋病毒-1/艾滋病的流行，然而，保护性疫苗的开发 这仍然是一个巨大的挑战，需要新的战略。多措并举稳定全局 HIV-1包膜的胞外区(Env)已导致疫苗设计的极大改进，例如 开发SOSIP三聚体，它除了提供了对SOSIP三聚体的精细结构理解外，还 Env三聚体被认为是一种很有前途的免疫原，因为它包含了所有已知的易受攻击的部位 BNAbs靶向的环境三聚体。然而，基于SOSIP的免疫原仅取得了有限的成功 由于不想要的分散抗原表位的位置，将抗体反应转移到菌株特异性的。与整个环境不同 细胞外域方法，我们一直在使用分而治之的策略，从个人开始 以开发具有免疫优势的区域特异性免疫原。 将抗体反应集中于选定的包膜结构域和表位并避免将抗体诱导为不需要的抗体 表位区域。我们从V1V2域开始，不仅因为它的域结构在空间上位于环境中 在RV144试验中，APEX及其抗体与感染风险呈负相关，已经开发出一组 通过将gp120的V1V2支架在三聚体非HIV蛋白上而形成的三聚体V1V2结构域免疫原，以及 证明它们可以诱导V1V2集中的抗体反应。此外，进一步的设计努力也导致了 在单链串联的V1V2三聚体中，显示出与三聚体特异性bNAbs的抗原反应性。我们假设 这种V1V2结构域的免疫原可以通过进一步的改进来呈现天然的Env顶点配置 结构表征和工程。为了从Env顶端扩展，我们随后设计了一种“重新取芯” 通过将灌流三聚体结构中的gp120内部结构域和gp41替换为 稳定的三聚体非HIV支架蛋白；抗原性测试表明该分子含有所有关键的bNab 三聚体Env顶端的结合部位和EM可视化显示它具有结构良好的三聚体 配置。这种新的结构为开发携带密钥的三聚体免疫原提供了一个起点 Env三聚体的易受攻击的部位，可能成为SOSIP三聚体的有吸引力的替代品，提供 更多地关注最有前景的bNab表位。这个R01项目的目标是在结构上描述 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，它们可以作为针对天然三聚体构型的有效免疫原。我们 有三个目标。1)表征和提纯模仿本地的三聚体V1V2-支架结构 Env三聚体顶端的预融合构象。2)研制重组Env三聚体免疫原包埋键 BNab站点。3)精制免疫原在动物体内的免疫原性试验。在完成这项工作后 项目中，我们的新型三聚体免疫原将得到充分的表征，并选择类似天然的免疫原可以 然后在疫苗开发和NHP挑战研究方面取得进展。

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.