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. 2021 Nov 24:12:777672.
doi: 10.3389/fimmu.2021.777672. eCollection 2021.

Monomeric IgA Antagonizes IgG-Mediated Enhancement of DENV Infection

Adam D Wegman  1 , Hengsheng Fang  1 , Alan L Rothman  2 , Stephen J Thomas  1   3 , Timothy P Endy  1 , Michael K McCracken  4 , Jeffrey R Currier  4 , Heather Friberg  4 , Gregory D Gromowski  4 , Adam T Waickman  1   3
Affiliations

Monomeric IgA Antagonizes IgG-Mediated Enhancement of DENV Infection

Adam D Wegman et al. Front Immunol. .

Abstract

Dengue virus (DENV) is a prevalent human pathogen, infecting approximately 400 million individuals per year and causing symptomatic disease in approximately 100 million. A distinct feature of dengue is the increased risk for severe disease in some individuals with preexisting DENV-specific immunity. One proposed mechanism for this phenomenon is antibody-dependent enhancement (ADE), in which poorly-neutralizing IgG antibodies from a prior infection opsonize DENV to increase infection of Fc gamma receptor-bearing cells. While IgM and IgG are the most commonly studied DENV-reactive antibody isotypes, our group and others have described the induction of DENV-specific serum IgA responses during dengue. We hypothesized that monomeric IgA would be able to neutralize DENV without the possibility of ADE. To test this, we synthesized IgG and IgA versions of two different DENV-reactive monoclonal antibodies. We demonstrate that isotype-switching does not affect the antigen binding and neutralization properties of the two mAbs. We show that DENV-reactive IgG, but not IgA, mediates ADE in Fc gamma receptor-positive K562 cells. Furthermore, we show that IgA potently antagonizes the ADE activity of IgG. These results suggest that levels of DENV-reactive IgA induced by DENV infection might regulate the overall IgG mediated ADE activity of DENV-immune plasma in vivo, and may serve as a predictor of disease risk.

Keywords: ADE; DENV; Dengue; IgA; antibody dependent enhancement.

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Conflict of interest statement

Authors ADW, MM, JC, HF, GG, and ATW are co-inventors on the provisional patent “IgA monoclonal antibodies as a prophylactic and therapeutic treatment for acute flavivirus infection”. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

Figure 1
Figure 1
Isotype conversion scheme, DENV binding, and DENV neutralization capacity of VDB33 and VDB50 mAbs. (A) Schematic of isotype conversion of VDB33 and VDB50 from respective parental isotypes, indicating conservation of antigen-binding domains and alteration of Fc domains. (B) DENV-3 binding capability of VDB33-IgG, VDB33-IgA, VDB50-IgG, and VDB50-IgA measured by DENV virus-capture ELISA. (C) DENV-3 neutralization capability of VDB33-IgG, VDB33-IgA, VDB50-IgG, and VDB50-IgA as assessed by FlowNT. Neutralization data are presented as a percent of the positive (no neutralizing mAb) control for each replicate. Error bars +/- SEM. Data are representative of at least 4 independent experiments.
Figure 2
Figure 2
ADE activity of DENV-reactive IgG and IgA isotype antibodies. (A) ADE activity of VDB33-IgG and VDB33-IgA against DENV-3 in K562 cells. (B) AUC values of 7 independent experimental replicates of DENV-3 ADE assay with VDB33-IgG and VDB33-IgA (C) ADE activity of VDB50-IgG and VDB50-IgA against DENV-3 in K562 cells. (D) AUC values of 7 independent replicates of DENV-3 ADE assay with VDB50-IgG and VDB50-IgA. Error bars +/- SEM. **p < 0.01, ****p < 0.0001, unpaired t test. Data are representative of at least 7 independent experiments.
Figure 3
Figure 3
Homotypic and heterotypic monoclonal IgA antagonizes IgG-mediated antibody-dependent enhancement. (A) DENV-3 ADE activity of VDB33-IgG when antagonized with VDB33-IgA. Total antibody concentration for each dilution point was held constant, with varying ratios of VDB33-IgG and VDB33-IgA as indicated. AUC of each ADE titration was calculated and normalized to that of the 100% IgG condition. (B) DENV-3 ADE activity of VDB33-IgG when antagonized with VDB50-IgA. The AUC of each ADE titration was calculated and normalized to that of the 100% VDB33-IgG condition. (C) DENV-3 ADE activity of VDB50-IgG when antagonized with VDB33-IgA. AUC of each ADE titration was calculated and normalized to that of the 100% VDB50-IgG condition. (D) DENV-3 ADE activity of VDB50-IgG when antagonized with VDB5o-IgA. AUC of each ADE titration was calculated and normalized to that of the 100% VDB33-IgG condition. Blue = 100% IgG/0% IgA. Green = 90% IgG/10% IgA. Orange = 50% IgG/50% IgA. Red = 0% IgG/100% IgA. "ns" denotes "non-significant" (p > 0.05). *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001 1-way ANOVA with Dunnett correction for multiple comparisons. Data are representative of at least 4 independent experiments.
Figure 4
Figure 4
Monoclonal IgA antagonizes ADE mediated by polyclonal DENV-immune plasma. (A) DENV immune plasma enhances DENV-3 infection of K562 cells. Each datapoint represents a unique plasma donor (n = 4). (B) VDB33-IgA antagonizes in vitro enhancement of DENV-3 infection mediated by polyclonal DENV-immune serum. Serum used at a 1:50 dilution for ADE assay, n = 4 unique plasma donors. The percentage of DENV-positive cells was normalized to that observed in the plasma-only condition. (C) VDB50-IgA antagonizes in vitro enhancement of DENV-3 infection mediated by polyclonal DENV-immune serum. Serum used at a 1:50 dilution for ADE assay, n = 4 unique plasma donors. The percentage of DENV-positive cells was normalized to that observed in the plasma-only condition. ***p < 0.001, ****p < 0.0001 1-way ANOVA with Dunnett correction for multiple comparisons. Data are representative of at least 4 independent experiments.
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