Plasma and memory antibody responses to Gamma SARS-CoV-2 provide limited cross-protection to other variants

Abstract

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to be a global problem in part because of the emergence of variants of concern that evade neutralization by antibodies elicited by prior infection or vaccination. Here we report on human neutralizing antibody and memory responses to the Gamma variant in a cohort of hospitalized individuals. Plasma from infected individuals potently neutralized viruses pseudotyped with Gamma SARS-CoV-2 spike protein, but neutralizing activity against Wuhan-Hu-1-1, Beta, Delta, or Omicron was significantly lower. Monoclonal antibodies from memory B cells also neutralized Gamma and Beta pseudoviruses more effectively than Wuhan-Hu-1. 69% and 34% of Gamma-neutralizing antibodies failed to neutralize Delta or Wuhan-Hu-1. Although Class 1 and 2 antibodies dominate the response to Wuhan-Hu-1 or Beta, 54% of antibodies elicited by Gamma infection recognized Class 3 epitopes. The results have implications for variant-specific vaccines and infections, suggesting that exposure to variants generally provides more limited protection to other variants.

Figure 1.

Characterization of serological responses in Gamma-infected individuals. (A) Surface representation of structure of SARS-CoV-2 RBD. The positions of key Gamma mutations are highlighted in orange, and the ACE-2 epitope is indicated by purple dashed lines; inset shows the ACE-2 (shown as a ribbon diagram) and RBD interaction. (B) Binding to Wuhan-Hu-1 and variant RBDs by plasma IgG from Gamma-infected (black) and prepandemic (red) cohorts, summarized as area under the curve (AUC). (C) Neutralization activity against Wuhan-Hu-1 and variant pseudoviruses by plasma IgG from Gamma-infected (black) and Wuhan-Hu-1–infected (gray) cohorts, summarized as NT₅₀ values. B and C show averaged results from duplicate experiments. Numbers in red are mean geometric values; statistical differences determined by two-sided Kruskal-Wallis test with subsequent Dunn’s multiple comparisons.

Figure S1.

Identification of cross-reactive anti–Wuhan-Hu-1 and anti-417N/484K/501Y RBD antibodies from Gamma-infected individuals. (A) Gating strategy. Gating was performed on singlets that were CD20⁺ and CD3⁻ CD8⁻ CD14⁻ CD16⁻ Ova⁻. Sorted cells were Wuhan-Hu-1 RBD-PE⁺ and 417N/484K/501Y RBD-AF647⁺. (B) Representative flow cytometry plots showing PE-RBD– and AF-647-RBD–binding B cells for one control and three study donors. Gating strategy is shown in Fig. S2 A. (C) Distribution of antibody sequences obtained from three donors. The number in the inner circle indicates the number of sequences analyzed per individual. White indicates sequences isolated once, while gray slices are proportional to the number of clonally expanded sequences. The outer black arc denotes the frequency of clonal sequences per donor. Related to Fig. 2.

Figure 2.

Antibody V gene frequency and mutations. (A–C) Bar graphs show the frequency distributions of human V genes for heavy chain (A), kappa chain (B), and lambda chain (C) in antibodies from Gamma-infected donors (blue) and Sequence Read Archive accession SRP010970 (orange). Statistical significance determined by two-sided binomial test with unequal variance; significant differences are denoted by asterisks (*, P < 0.05; **, P < 0.01; ****, P < 0.0001). (D) Number of somatic nucleotide mutations in IGVH (P = 0.0253 for BRA14 versus BRA15), IGVK and IGVL combined (P = 0.0177 for BRA13 versus BRA15), and total IGV (P = 0.0162 for BRA13 versus BRA15; P = 0.0091 for BRA14 versus BRA15) as indicated per donor. Red horizontal bars indicate mean values; statistical differences determined by two-sided Kruskal-Wallis test with subsequent Dunn’s multiple comparisons.

Figure S2.

Antibody sequence hydrophobicity and CDR3 length. (A) Distribution of hydrophobicity GRAVY scores at the IGH CDR3 of antibodies from all donors combined and compared to human repertoire (Briney et al., 2019). (B) CDR3 lengths for all heavy and light chains of antibodies isolated in this study compared to human repertoire. For A and B, statistical significance is determined by two-sided binomial test with unequal variance and denoted by asterisks (*, P < 0.05; ****, P < 0.0001). Related to Fig. 2.

Figure 3.

Characterization of cross-reactive but not cross-neutralizing antibodies. (A) K_D for Wuhan-Hu-1 and Gamma RBDs of antibodies from Gamma-infected cohort. Red horizontal bars indicate geometric mean values; no significant difference by two-sided Wilcoxon matched-pairs signed rank test. BLI traces shown in Fig. S2, A and B; mean K_D calculated based on triplicate binding curves matching theoretical fit with R² value ≥ 0.8. (B) Neutralization of Wuhan-Hu-1 pseudovirus by monoclonal antibodies from Wuhan-Hu-1–infected (gray) and Gamma-infected (black) cohorts, summarized as IC₅₀ values. P = 0.0095 by two-sided Mann-Whitney test. (C) Neutralization of Wuhan-Hu-1 (R683G), Gamma (R683G), and Omicron (R683G) pseudovirus by monoclonal antibodies from Gamma-infected cohort, summarized as IC₅₀ values. Lines connect individual antibodies across variants. Dashed line indicates the limit of detection. P = 0.5338 for Wuhan-Hu-1 (R683G) versus Omicron BA.1 (R683G); P = 0.0158 for Wuhan-Hu-1 (R683G) versus Gamma (R683G); and P = 0.0001 for Gamma (R683G) versus Omicron BA.1 (R683G). Statistical significance determined by Friedman’s test followed by Dunn’s multiple comparisons. For A and B, red horizontal bars indicate geometric mean values. Average IC₅₀ values calculated based on duplicate experiments. (D) IC₅₀ values for n = 18 antibodies against indicated mutant SARS-CoV-2 pseudoviruses. Color gradient indicates IC₅₀ values ranging 0 (white) to 1,000 ng/ml (red). Average IC₅₀ values calculated based on duplicate experiments. (E) Schematic of BLI experiment. (F) Bar graph showing percentages of antibodies assigned to each binding class based on BLI epitope binning experiments for n = 28 antibodies each from Wuhan-Hu-1–infected (gray) and Gamma-infected (black) cohorts. Significance (P = 0.0005) determined using Fisher’s test for exact count data.

Figure S3.

Biolayer interferometry affinity measurements. (A and B) Graphs depict affinity measurements of anti-Gamma monoclonal antibodies for Wuhan-Hu-1 RBD (A) or N417/484K/501Y RBD (B); data are representative of experiments performed in triplicate. (C) Heat maps of two biolayer interferometry competition experiments showing relative inhibition of binding of monoclonal antibody (x axis) to preformed complexes of 417N/484K/501Y RBD with another monoclonal antibody (y axis). Gray indicates no binding; yellow indicates low binding; orange, intermediate binding; and red, high binding. Data are normalized by subtraction of autologous antibody control. Related to Fig. 3.