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. 2021 Jun 15;2(6):100296.
doi: 10.1016/j.xcrm.2021.100296. Epub 2021 May 9.

Decay of Fc-dependent antibody functions after mild to moderate COVID-19

Wen Shi Lee  1 , Kevin John Selva  1 , Samantha K Davis  1 , Bruce D Wines  2   3   4 , Arnold Reynaldi  5 , Robyn Esterbauer  1 , Hannah G Kelly  1   6 , Ebene R Haycroft  1 , Hyon-Xhi Tan  1 , Jennifer A Juno  1 , Adam K Wheatley  1   6 , P Mark Hogarth  2   3   4 , Deborah Cromer  5 , Miles P Davenport  5 , Amy W Chung  1 , Stephen J Kent  1   6   7
Affiliations

Decay of Fc-dependent antibody functions after mild to moderate COVID-19

Wen Shi Lee et al. Cell Rep Med. .

Abstract

The capacity of antibodies to engage with immune cells via the Fc region is important in preventing and controlling many infectious diseases. The evolution of such antibodies during convalescence from coronavirus disease 2019 (COVID-19) is largely unknown. We develop assays to measure Fc-dependent antibody functions against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike (S)-expressing cells in serial samples from subjects primarily with mild-moderate COVID-19 up to 149 days post-infection. We find that S-specific antibodies capable of engaging Fcγ receptors decay over time, with S-specific antibody-dependent cellular cytotoxicity (ADCC) and antibody-dependent phagocytosis (ADP) activity within plasma declining accordingly. Although there is significant decay in ADCC and ADP activity, they remain readily detectable in almost all subjects at the last time point studied (94%) in contrast with neutralization activity (70%). Although it remains unclear the degree to which Fc effector functions contribute to protection against SARS-CoV-2 re-infection, our results indicate that antibodies with Fc effector functions persist longer than neutralizing antibodies.

Keywords: ADCC; ADP; COVID-19; Fc effector functions; SARS-CoV-2; antibody; convalescence; decay; phagocytosis; trogocytosis.

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Conflict of interest statement

The authors declare no competing interests.

Figures

None
Graphical abstract
Figure 1
Figure 1
Dynamics of SARS-CoV-2 S and RBD-specific dimeric FcγR-binding antibodies in COVID-19 convalescent individuals (A) Timeline of sample collection for each COVID-19 convalescent subject (n = 53). Subjects with 2 samples at least 60 days apart were chosen for functional assay analysis (n = 36). (B and C) Kinetics of SARS-CoV-2 S and RBD-specific dimeric FcγRIIIa (V158) and dimeric FcγRIIa (H131) binding antibodies over time measured using the bead-based multiplex assay. The best-fit decay slopes (red lines) and estimated half-lives (t1/2) are indicated for COVID-19 convalescent individuals. Uninfected controls (n = 33) are shown in open circles, with the median and 90% percentile responses presented as thick and thin dashed lines, respectively. The limit of detection is shown as the shaded area. See also Figure S1.
Figure 2
Figure 2
ADCC responses in COVID-19 convalescent individuals over time (A) Schematic of the FcγRIIIa NF-κB activation assay. IIA1.6 cells expressing FcγRIIIa V158 and a NF-κB response-element-driven nanoluciferase reporter were co-incubated with A549 S-orange target cells and plasma from COVID-19 convalescent individuals or uninfected controls. The engagement of FcγRIIIa by S-specific antibodies activates downstream NF-κB signaling and nanoluciferase expression. (B) S-specific FcγRIIIa-activating plasma antibodies in COVID-19 convalescent individuals in the first (T1; filled) and last (T2; open) time points available. Blue lines indicate the median responses of COVID-19 convalescent individuals (N = 36), and dashed lines indicate median responses of uninfected controls (N = 8). (C) The best-fit decay slopes (red lines) and estimated half-life (t1/2) for FcγRIIIa-activating plasma antibodies in COVID-19 convalescent individuals. Uninfected controls are shown in open circles, with the median response presented as a dashed line. (D) Correlation of S-specific FcγRIIIa-activating antibodies to cell-associated S-specific IgG and S-specific dimeric FcγRIIIa-binding antibodies. (E) Schematic of the luciferase-based ADCC assay. Purified NK cells from healthy donors were co-incubated with Ramos S-luciferase target cells and plasma. ADCC is measured as the loss of cellular luciferase. (F) S-specific ADCC mediated by plasma antibodies from COVID-19 convalescent individuals in the first (T1; filled) and last (T2; open) time points available. Blue lines indicate the median responses of COVID-19 convalescent individuals (N = 36), and dashed lines indicate median responses of uninfected controls (N = 8). (G) The best-fit decay slopes (red lines) and estimated half-life (t1/2) for FcγRIIIa-activating plasma antibodies in COVID-19 convalescent individuals. Uninfected controls are shown in open circles, with the median response presented as a dashed line. (H) Correlation of S-specific ADCC to cell-associated S-specific IgG and S-specific dimeric FcγRIIIa-binding antibodies. Statistical analyses were performed with the Wilcoxon signed-rank test (∗∗∗∗p < 0.0001). Correlations were performed with the non-parametric Spearman test. See also Figures S2 and S3.
Figure 3
Figure 3
ADP responses in COVID-19 convalescent individuals over time (A) Schematic of the bead-based ADP assay. THP-1 cells were incubated with S-conjugated fluorescent beads and plasma from COVID-19 convalescent individuals or uninfected controls. The uptake of fluorescent beads was measured by flow cytometry. (B) ADP of S-conjugated beads mediated by plasma antibodies from COVID-19 convalescent individuals in the first (T1) and last (T2) time points available. Blue lines indicate the median responses of COVID-19 convalescent individuals (N = 36), and dashed lines indicate median responses of uninfected controls (N = 8). (C) The best-fit decay slopes (red lines) and estimated half-life (t1/2) for plasma ADP activity in COVID-19 convalescent individuals. Uninfected controls are shown in open circles, with the median response presented as a dashed line. (D) Correlation of ADP to cell-associated S-specific IgG and S-specific dimeric FcγRIIa-binding antibodies. (E) Schematic of the THP-1 FcγR-dependent cell association assay. Ramos S-orange cells were pre-incubated with plasma prior to co-incubation with THP-1 cells. The association of THP-1 cells with Ramos S-orange cells was measured by flow cytometry. (F) FcγR-dependent association of THP-1 cells with Ramos S-orange cells mediated by plasma antibodies from COVID-19 convalescent individuals in the first (T1) and last (T2) time points available. Blue lines indicate the median responses of COVID-19 convalescent individuals (N = 36), and dashed lines indicate median responses of uninfected controls (N = 8). (G) The best-fit decay slopes (red lines) and estimated half-life (t1/2) for THP-1 association in COVID-19 convalescent individuals. Uninfected controls are shown in open circles, with the median response presented as a dashed line. (H) Correlation of association of THP-1 cells with Ramos S-orange cells to cell-associated S-specific IgG and S-specific dimeric FcγRIIIa-binding antibodies. Red lines indicate the median responses of COVID-19 convalescent individuals (N = 36), and dashed lines indicate median responses of uninfected controls (N = 8). Statistical analyses were performed with the Wilcoxon signed-rank test (∗∗p < 0.01). Correlations were performed with the non-parametric Spearman test. See Figure S4 for gating strategies and Figure S5 for assay optimization.
Figure 4
Figure 4
Confocal microscopy visualization of Fc-mediated association and trogocytosis THP-1 monocytes were incubated with Ramos S-orange target cells in the presence of healthy control plasma or COVID-19 convalescent plasma. Cells were fixed on slides and visualized using a confocal microscope (60× objective). THP-1 cells were stained with CD32-AF647 (blue), and Ramos S-orange cells (expressing SARS-CoV-2 spike and mOrange2) were labeled with the membrane dye PKH-26 (red). Examples of trogocytosis in the COVID-19 convalescent plasma sample (white boxes) are enlarged and shown in the fourth and fifth rows. The white arrows indicate THP-1 cells (blue) that have received plasma membrane proteins from Ramos S-orange cells (red). No trogocytosis occurred in the healthy control sample (white box, enlarged in second row). Scale bars are 13 μM for the first and third rows and 18 μM for the enlarged images in the second, fourth, and fifth rows.
Figure 5
Figure 5
Dynamics of dimeric FcγR-binding antibodies against HCoV S antigens in COVID-19 convalescent individuals (A) Best-fit decay slopes of IgG and dimeric FcγR-binding antibodies against S from HCoV strains OC43, HKU1, 229E, and NL63. The responses at time point 1 for each parameter are set to 100%, and the %change over time is shown. (B–E) Kinetics of dimeric FcγRIIIa (V158) and FcγRIIa (H131) binding antibodies against S, S1, or S2 from HCoV strains (B) OC43, (C) HKU1, (D) 229E, and (E) NL63 over time in COVID-19 convalescent individuals (N = 53) measured using the bead-based multiplex assay. The best-fit decay slopes (red lines) and estimated half-lives (t1/2) are indicated for COVID-19 convalescent individuals. Uninfected controls (N = 33) are shown in open circles, with the median and 90% percentile responses presented as thick and thin dashed lines, respectively. The limit of detection is shown as the shaded area. See also Figure S6.
Figure 6
Figure 6
Decay kinetics of binding antibodies, neutralization, and Fc effector functions following SARS-CoV-2 infection (A) Best-fit decay slopes of various antibody parameters against SARS-CoV-2 S over time. The responses at time point 1 for each parameter are set to 100%, and the %change over time is shown. (B) The percentage of subjects having detectable responses above (red) and below (gray) background levels at the last visit are shown. Background levels for each assay were the median responses of uninfected controls.
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