Class switch towards non-inflammatory, spike-specific IgG4 antibodies after repeated SARS-CoV-2 mRNA vaccination [CITE-Seq]

RNA vaccines are efficient preventive measures to combat the SARS-CoV-2 pandemic. High levels of neutralizing SARS-CoV-2-antibodies are an important component of vaccine-induced immunity. Shortly after the initial two mRNA vaccine doses, the IgG response mainly consists of the pro-inflammatory subclasses IgG1 and IgG3. Here, we report that several months after the second vaccination, SARS-CoV-2-specific antibodies were increasingly composed of non-inflammatory IgG4, which were further boosted by a third mRNA vaccination and/or SARS-CoV-2 variant breakthrough infections. IgG4 antibodies among all spike-specific IgG antibodies rose on average from 0.04% shortly after the second vaccination to 19.27% late after the third vaccination. This induction of IgG4 antibodies was not observed after homologous or heterologous SARS-CoV-2 vaccination with adenoviral vectors. Single-cell sequencing and flow cytometry revealed substantial frequencies of IgG4-switched B cells within the spike-binding memory B-cell population (median 14.4%; interquartile range (ICR) 6.7-18.1%) compared to the overall memory B-cell repertoire (median 1.3%; ICR 0.9-2.2%) after three immunizations. Importantly, this class switch was associated with a reduced capacity of the spike-specific antibodies to mediate antibody-dependent cellular phagocytosis and complement deposition. Since Fc-mediated effector functions are critical for antiviral immunity, these findings may have consequences for the choice and timing of vaccination regimens using mRNA vaccines, including future booster immunizations against SARS-CoV-2. We performed single-cell RNA sequencing (scRNA seq) of spike-specific B cells from four selected donors 210 days after the second or 10 days after the third Comirnaty dose. We combined this technology to CITE-sequencing to obtain measurements of surface proteins and the transcriptome and to enable HTO HASH-sequencing in order to demultiplex data sets.

RNA疫苗是对抗SARS-CoV-2大流行的高效预防手段。高滴度中和性SARS-CoV-2抗体是疫苗诱导免疫的重要组成部分。在完成初始两剂mRNA疫苗接种后不久，IgG应答主要由促炎亚类IgG1与IgG3构成。本研究报道，在第二次接种数月后，SARS-CoV-2特异性抗体中非炎症性IgG4的占比逐步升高，而第三剂mRNA疫苗接种或SARS-CoV-2变体突破感染可进一步提升该比例。在所有刺突蛋白（spike）特异性IgG抗体中，IgG4的占比从第二次接种后不久的平均0.04%，升至第三次接种后晚期的19.27%。采用腺病毒载体进行同源或异源SARS-CoV-2疫苗接种后，并未观察到此类IgG4抗体的诱导现象。单细胞测序与流式细胞术结果显示，在完成三次免疫接种后，与整体记忆B细胞库（中位数1.3%；四分位距（interquartile range，IQR）0.9-2.2%）相比，刺突蛋白结合性记忆B细胞群体中发生IgG4类别转换的B细胞占比可观（中位数14.4%；四分位距（interquartile range，IQR）6.7-18.1%）。值得注意的是，这种类别转换与刺突蛋白特异性抗体介导抗体依赖细胞吞噬作用及补体沉积的能力下降相关。由于Fc段介导的效应功能对抗病毒免疫至关重要，本研究结果或对mRNA疫苗接种方案的选择与时机安排产生影响，包括未来针对SARS-CoV-2的加强免疫接种。我们对4名选定受试者在第二次接种后210天或第三次复必泰（Comirnaty）接种后10天的刺突蛋白特异性B细胞开展了单细胞RNA测序（single-cell RNA sequencing，scRNA-seq）。本研究将该技术与CITE测序（CITE-sequencing）相结合，以获取表面蛋白与转录组的检测数据，并采用HTO HASH测序（HTO HASH-sequencing）实现数据集的多重复用拆分。