Vaccination of SARS-CoV-2-infected individuals expands a broad range of clonally diverse affinity-matured B cell lineages

Abstract

Vaccination of SARS-CoV-2 convalescent individuals generates broad and potent antibody responses. Here, we isolate 459 spike-specific monoclonal antibodies (mAbs) from two individuals who were infected with the index variant of SARS-CoV-2 and later boosted with mRNA-1273. We characterize mAb genetic features by sequence assignments to the donors' personal immunoglobulin genotypes and assess antibody neutralizing activities against index SARS-CoV-2, Beta, Delta, and Omicron variants. The mAbs used a broad range of immunoglobulin heavy chain (IGH) V genes in the response to all sub-determinants of the spike examined, with similar characteristics observed in both donors. IGH repertoire sequencing and B cell lineage tracing at longitudinal time points reveals extensive evolution of SARS-CoV-2 spike-binding antibodies from acute infection until vaccination five months later. These results demonstrate that highly polyclonal repertoires of affinity-matured memory B cells are efficiently recalled by vaccination, providing a basis for the potent antibody responses observed in convalescent persons following vaccination.

Fig. 1. Study design, serum neutralizing activity, and properties of spike-binding monoclonal antibodies (mAbs).

a Schematic of study design with timepoints for infection and vaccination, mAb isolation, and samples for Rep-seq. b IML3694 and IML3695’s serum neutralizing antibody values against index SARS-CoV-2 (index) and variants of concern (VOCs). c Pie charts of mAb subdomain specificities in the two donors. d mAb IGHV allele frequencies colored by donor. e mAb IGHV allele frequencies colored by subdomain specificity.

Fig. 2. Properties of SARS-CoV-2 neutralizing mAbs.

An IC₅₀ threshold of 0.4 μg/ml against any of the SARS-CoV-2 variants qualified mAbs as neutralizing. The table includes heavy and light chain VDJ assignments as well as SHM values and IC₅₀s against 6 SARS-CoV-2 variants.

Fig. 3. Isolation timepoints and evolution of HCoV-HKU1 S-cross-reactive mAbs.

a Pie chart of timepoints at which HCoV-HKU1 S-binding mAbs were isolated. b Dot plot of % nucleotide IGHV SHM for the HCoV-HKU1 S-cross-reactive and SARS-CoV-2 S-specific mAbs isolated at the acute timepoint. A two-sided Mann–Whitney U test was used for comparison; *** indicates a P value < 0.001, with the exact value being P value = 1.261e-5. c Maximum-likelihood phylogenetic trees of HCoV-HKU1 S-binding traced lineage ADI-66175. The germline sequence was obtained from the IgBLAST-generated “germline_alignment” column of the sequence with the minimum IGHV SHM in the lineage.

Fig. 4. Evolution of SARS-CoV-2 S-specific antibody lineages over the sampling timepoints.

a Maximum-likelihood phylogenetic tree of traced lineages containing IgM Rep-seq sequences: ADI-66196, ADI-67860, and ADI-67983. Germlines sequences were obtained from the IgBLAST-generated “germline_alignment” column of the sequence with the smallest IGHV SHM in the lineage. b Dot plot of % nucleotide IGHV SHM for the ADI-66196, ADI-67860, and ADI-67983 lineage sequences. Purple crossbars represent mAb values, while the blue crossbars represent average values of traced variants for each timepoint.

Fig. 5. IgM and IgG repertoire sequencing and lineage tracing of SARS-CoV-2 S-specific antibody lineages at the longitudinal timepoints.

a Bubble plots of SARS-CoV-2 S-specific mAb lineages found before and after vaccination in IML3694 and IML3695. The bubble sizes correspond to post-FAD and chimera-cleaned data. b Line plots of median % nucleotide IGHV SHM for lineages found in acute IgM/IgG and post-vax IgG Rep-seq data. c Boxplot of median % nucleotide IGHV SHM for lineages containing pre-vax timepoint mAbs and post-vax IgG Rep-seq. The box represents the median ± interquartile range and the whiskers represent the maximum and minimum values. A two-sided Wilcoxon signed-rank test was used for comparison; P value = 0.3107.