Adjuvant, mimicry and booster requirements for shepherding the development of neutralizing antibodies to the high-mannose patch on HIV-1

项目摘要 HIV-1包膜刺突（Env）带有一簇寡甘露糖型聚糖，其是广泛的免疫调节剂的靶标。 中和抗体（bnAb）。然而，当nAb到达这个被称为高甘露糖斑块的簇时， (HMP)已知至少在一些HIV感染者中发生，过去试图引起类似的 通过免疫接种产生抗体在很大程度上不成功。以前的大多数方法都涉及 在载体蛋白的表面上呈现天然或合成的高甘露糖聚糖簇。的 通过免疫诱导高甘露糖靶向nAb的困难被认为，至少部分地， 靶向聚糖的“自身”性质;携带具有所需精细特异性的IG受体的幼稚B细胞 或亲和力可能是无反应性的或由于耐受机制而处于相对低的频率。 我们所追求的方法是基于这样的科学前提，即抗原模拟 哺乳动物宿主结构可以刺激交叉反应性抗体， 真正的“外国”环境我们的首要假设是，在免疫后， 哺乳动物寡甘露糖比天然或合成的寡甘露糖更容易引发结合HMP的抗体 寡甘露糖在我们的进展报告中，我们表明我们的前导寡聚甘露糖的CRM 197-缀合物 模拟物，其设计灵感来自于与哺乳动物相似的细菌多糖 寡甘露糖以高亲合力与各种HMP特异性bnAb以及它们的种系结合 前体此外，用这种新糖缀合物免疫的人抗体转基因小鼠产生 结合重组HIV-1 SOSIP三聚体的抗体，尽管仅当缀合物被配制在 刺激TLR 4的Th 1佐剂GLA-SE;当配制在Th 2佐剂或混合的Th 1/2佐剂中时，则不是。 根据我们到目前为止的调查结果，我们在这次续期申请中建议扩大我们的调查范围。第一 第一步（目标1）将通过评估免疫原性，进一步探索Th 1刺激的相关性。 当与其他Th 1导向佐剂一起配制时，我们的先导糖缀合物。这些额外的佐剂 不会激动TLR 4，因此有助于揭示刺激TLR 4是否相关。我们将评估 还（目的2）抗原模拟对于触发高甘露糖特异性抗体的意义，通过 比较我们的先导模拟物与合成寡甘露糖的免疫原性。我们也将评估 是否将革兰氏阴性细菌LPS-Kdo和脂质A-的典型组分掺入 我们的先导苷对免疫有益我们的第三步（目标3）是调查 在特定时间点用选择的SOSIP三聚体加强注射糖缀合物引发的转基因小鼠 nAb滴度。我们希望我们的研究结果有助于加强我们的战略，并为未来的追求提供重要信息。 临床前研究。这项研究的结果也可以为其他艾滋病毒疫苗设计策略提供信息。

PROJECT SUMMARY The HIV-1 envelope spike (Env) bears a cluster of oligomannose-type glycans that is a target for broadly neutralizing antibodies (bnAbs). However, while nAbs to this cluster, dubbed the high-mannose patch (HMP), are known to develop in at least some HIV-infected individuals, past attempts to elicit similar antibodies by immunization have been largely unsuccessful. Most previous approaches have involved presenting clusters of natural or synthetic high-mannose glycans on the surface of carrier proteins. The difficulty in eliciting high-mannose-targeting nAbs by immunization is believed to relate, at least in part, to the ‘self’ nature of the targeted glycans; naïve B cells bearing Ig receptors of the desired fine specificity or affinity might be anergic or at relatively low frequency due to tolerance mechanisms. The approach that we are pursuing is based on the scientific premise that antigenic mimicry of mammalian host structures can stimulate cross-reactive antibodies if such mimics are presented in the proper ‘foreign’ milieu. Our overarching hypothesis is that, upon immunization, an antigenic mimic of mammalian oligomannose will more readily elicit antibodies that bind the HMP than native or synthetic oligomannose. In our progress report, we show that a CRM197-conjugate of our lead oligomannose mimetic, the design of which is inspired by a bacterial polysaccharide with resemblance to mammalian oligomannose, is bound with high avidity by various HMP-specific bnAbs as well as their germline precursors. Furthermore, human antibody transgenic mice immunized with this neoglycoconjugate yield antibodies that bind recombinant HIV-1 SOSIP trimers, albeit only when the conjugate is formulated in the TLR4-stimulating Th1-adjuvant GLA-SE; not when formulated in Th2- or mixed Th1/2-adjuvants. Based on our findings so far, we propose in this renewal application to extend our investigations. A first step (Aim 1) will be to probe the relevance of Th1 stimulation further, by assessing immunogenicity of our lead glycoconjugate when formulated with other Th1-directing adjuvants. These additional adjuvants will not be agonistic for TLR4, thus helping to reveal whether stimulating TLR4 is relevant. We will assess also (Aim 2) the significance of antigenic mimicry for triggering high mannose-specific antibodies, by comparing the immunogenicity of our lead mimetic to synthetic oligomannose. We will evaluate also whether incorporating an archetypical component of gram-negative bacterial LPS –Kdo and lipid A—into our lead glycoside is immunologically beneficial. Our third step (Aim 3) will be to investigate whether injecting glycoconjugate-primed transgenic mice at specific time points with select SOSIP trimers boosts nAb titers. We expect our findings to help sharpen our strategy and critically inform the pursuit of future preclinical studies. Results from this research could inform other HIV vaccine design strategies also.