Mannosidase-stabilized glycan immunogens for elicitation of high mannose patch antibodies

项目摘要/摘要 HIV广谱中和抗体(BNAbs)最常针对高甘露糖斑(HMP)，a HIV env富含甘露糖区，位于V3环和N332糖链附近。HMP bNAbs(例如， 2G12、PGT128、BG18)倾向于识别内的非还原末端Man1α-2man部分 低聚甘露糖；然而，在寡聚糖偶联物免疫中，抗体是 很少与Man1a-2man结合，而是与糖链的核心基序结合。我们假设在 体内甘露糖苷酶对man1a-2man末端的修剪是抗体不产生的主要原因 反对这一主题。在目标1中，我们将制备稳定的低聚甘露糖，其中一个硫键 阻止MAN-1α-2MAN的甘露糖苷酶切割，但在结构上模仿天然多糖 由bNAbs识别。在目标2中，我们将用含有CRM197的糖结合物免疫兔子 稳定或不稳定的葡聚糖，并比较产生的抗体的HIV Env结合和 中和活性。我们还将深入研究这些抗体结合Man1a的能力- 2使用低聚甘露聚糖阵列的MAN。

Project Summary/Abstract HIV broadly neutralizing antibodies (bnAbs) most commonly target the High Mannose Patch (HMP), a mannose-rich region of HIV Env in the vicinity of the V3 loop and the N332 glycan. HMP bnAbs (e.g., 2G12, PGT128, BG18) tend to recognize non-reducing-terminal Man1α-2Man moieties within oligomannose glycans; however, in immunizations with oligomannose glycoconjugates, antibodies are rarely raised against Man1a-2Man, and instead bind to core motifs of the glycan. We hypothesize that in vivo mannosidase trimming of Man1a-2Man termini is a major reason why antibodies do not develop against this motif. In Aim 1, we will prepare stabilized oligomannose glycans, in which a sulfur linkage prevents mannosidase cleavage of the Man1α-2Man, but structurally mimics the natural glycan and is recognized by bnAbs. In Aim 2, we will immunize rabbits with CRM197 glycoconjugates containing the stabilized or unstabilized glycans and compare the resulting antibodies’ HIV Env binding and neutralization activity. We will also intensively characterize the ability of these antibodies to bind Man1a- 2Man using an oligomannose-focused glycan array.