Key Points The receptor binding site (CD4bs) of HIV Env is a desirable vaccine target. Envs lacking CD4bs-flanking glycans and bearing CD4i Ab moieties were generated. CD4bs-directed Ab responses have been elicited by these modified Envs. Elicitation of broadly neutralizing Ab (bNAb) responses toward the conserved HIV-1 envelope (Env) CD4 binding site (CD4bs) by vaccination is an important goal for vaccine development and yet to be achieved. The outcome of previous immunogenicity studies suggests that the limited accessibility of the CD4bs and the presence of predominant nonneutralizing determinants (nND) on Env may impede the elicitation of bNAbs and their precursors by vaccination. In this study, we designed a panel of novel immunogens that 1) preferentially expose the CD4bs by selective elimination of glycosylation sites flanking the CD4bs, and 2) minimize the nND immune response by engineering fusion proteins consisting of gp120 Core and one or two CD4-induced (CD4i) mAbs for masking nND epitopes, referred to as gp120–CD4i fusion proteins. As expected, the fusion proteins possess improved antigenicity with retained affinity for VRC01-class, CD4bs-directed bNAbs and dampened affinity for nonneutralizing Abs. We immunized C57BL/6 mice with these fusion proteins and found that overall the fusion proteins elicit more focused CD4bs Ab response than prototypical gp120 Core by serological analysis. Consistently, we found that mice immunized with selected gp120–CD4i fusion proteins have higher frequencies of germinal center–activated B cells and CD4bs-directed memory B cells than those inoculated with parental immunogens. We isolated three mAbs from mice immunized with selected gp120–CD4i fusion proteins and found that their footprints on Env are similar to VRC01-class bNAbs. Thus, using gp120–CD4i fusion proteins with selective glycan deletion as immunogens could focus Ab response toward CD4bs epitope.

中文翻译：

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HIV-1 gp120–CD4 诱导的抗体复合物在小鼠中引发 CD4 结合位点特异性抗体反应

要点 HIV Env 的受体结合位点 (CD4bs) 是理想的疫苗靶点。生成了缺少 CD4bs 侧翼聚糖和带有 CD4i Ab 部分的 Env。CD4bs 导向的 Ab 反应是由这些修饰的 Envs 引起的。通过接种疫苗引起对保守的 HIV-1 包膜 (Env) CD4 结合位点 (CD4bs) 的广泛中和抗体 (bNAb) 反应是疫苗开发的重要目标，但尚未实现。先前免疫原性研究的结果表明，CD4b 的有限可及性和 Env 上主要非中和决定簇 (nND) 的存在可能会阻碍 bNAb 及其前体通过疫苗接种的诱导。在这项研究中，我们设计了一组新型免疫原，1) 通过选择性消除 CD4bs 侧翼的糖基化位点来优先暴露 CD4bs，和 2) 通过工程融合蛋白由 gp120 Core 和一种或两种 CD4 诱导 (CD4i) 单克隆抗体组成，用于掩蔽 nND 表位（称为 gp120–CD4i 融合蛋白）来最小化 nND 免疫反应。正如预期的那样，融合蛋白具有改进的抗原性，保留了对 VRC01 类、CD4bs 导向的 bNAb 的亲和力，并减弱了对非中和性 Ab 的亲和力。我们用这些融合蛋白对 C57BL/6 小鼠进行免疫接种，发现总体上通过血清学分析，融合蛋白比原型 gp120 Core 引发更集中的 CD4bs Ab 反应。一致地，我们发现用选定的 gp120-CD4i 融合蛋白免疫的小鼠比接种亲本免疫原的小鼠具有更高频率的生发中心激活 B 细胞和 CD4bs 定向记忆 B 细胞。我们从用选定的 gp120-CD4i 融合蛋白免疫的小鼠中分离出三种 mAb，发现它们在 Env 上的足迹与 VRC01 类 bNAb 相似。因此，使用具有选择性聚糖缺失的 gp120-CD4i 融合蛋白作为免疫原可以将抗体反应集中在 CD4bs 表位上。