Key Points Immunization with COBRA P1 elicits broadly HA-binding ASC. P1 HA-elicited mAbs have different binding and functional activities. The P1-elicited 1F8 mAb exhibits broad H1N1 binding and functional activity. Visual Abstract Computationally optimized broadly reactive Ags (COBRA) targeting H1 elicit a broad cross-reactive and cross-neutralizing Ab response against multiple H1N1 viral strains. To assess B cell breadth, Mus musculus (BALB/c) Ab-secreting cells elicited by a candidate COBRA hemagglutinin (HA) (termed P1) were compared with Ab-secreting cells elicited by historical H1N1 vaccine strains. In addition, to evaluate the Ab response elicited by P1 HA at increased resolution, a panel of P1 HA-specific B cell hybridomas was generated following immunization of mice with COBRA P1 and the corresponding purified mAbs were characterized for Ag specificity and neutralization activity. Both head- and stem-directed mAbs were elicited by the P1 HA Ag, with some mAbs endowed with Ab-dependent cell-mediated cytotoxicity activity. P1 HA-elicited mAbs exhibited a wide breadth of HA recognition, ranging from narrowly reactive to broadly reactive mAbs. Interestingly, we identified a P1 HA-elicited mAb (1F8) exhibiting broad hemagglutination inhibition activity against both seasonal and pandemic H1N1 influenza strains. Furthermore, mAb 1F8 recognized an overlapping, but distinct, epitope compared with other narrowly hemagglutination inhibition–positive mAbs elicited by the P1 or wild-type HA Ags. Finally, P1 HA-elicited mAbs were encoded by distinct H chain variable and L chain variable gene segment rearrangements and possessed unique CDR3 sequences. Collectively, the functional characterization of P1 HA-elicited mAbs sheds further insights into the underlying mechanism(s) of expanded Ab breadth elicited by a COBRA HA-based immunogen and advances efforts toward design and implementation of a more broadly protective influenza vaccine.

中文翻译：

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一种计算优化的广泛反应抗原亚型特异性流感疫苗策略引发独特的强效广泛中和抗血凝素抗体

使用 COBRA P1 进行免疫接种会引发广泛的 HA 结合 ASC。P1 HA 引发的 mAb 具有不同的结合和功能活性。P1 引发的 1F8 mAb 表现出广泛的 H1N1 结合和功能活性。视觉摘要 针对 H1 的计算优化的广泛反应性抗原 (COBRA) 引发了针对多种 H1N1 病毒株的广泛交叉反应和交叉中和抗体反应。为了评估 B 细胞宽度，将由候选 COBRA 血凝素 (HA)（称为 P1）诱发的小家鼠 (BALB/c) Ab 分泌细胞与历史 H1N1 疫苗株诱发的 Ab 分泌细胞进行比较。此外，为了评估 P1 HA 在提高分辨率下引起的 Ab 反应，用 COBRA P1 对小鼠进行免疫接种后产生了一组 P1 HA 特异性 B 细胞杂交瘤，并表征了相应纯化的 mAb 的 Ag 特异性和中和活性。P1 HA Ag 可诱导头部和干细胞定向的 mAb，其中一些 mAb 具有依赖于 Ab 的细胞介导的细胞毒性活性。P1 HA 引发的 mAb 表现出广泛的 HA 识别，范围从窄反应性到广泛反应性 mAb。有趣的是，我们鉴定了一种 P1 HA 引发的 mAb (1F8)，它对季节性和大流行的 H1N1 流感毒株表现出广泛的血凝抑制活性。此外，与由 P1 或野生型 HA Ag 引起的其他狭义血凝抑制阳性 mAb 相比，mAb 1F8 识别出重叠但不同的表位。最后，P1 HA 引发的 mAb 由不同的 H 链可变和 L 链可变基因片段重排编码，并拥有独特的 CDR3 序列。总的来说，P1 HA 引发的 mAb 的功能特征进一步深入了解了基于 COBRA HA 的免疫原引发的扩大 Ab 广度的潜在机制，并推动了设计和实施更广泛的保护性流感疫苗的努力。