SARS-CoV-2 Neutralizing Antibody
The COVID-19 pandemic has spread over the world, and effective therapeutic and prophylactic interventions are urgently needed. Human-sourced monoclonal antibodies generated by convalescent patients’ B cells are promising therapeutic candidates. However, due to VDJ recombination and somatic hypermutation, B cells exhibit diverse B-cell repertoires, and this necessitates the analysis of one B cell at a time to obtain paired immunoglobulin heavy-light chain RNA sequences for monoclonal antibodies production. Using high-throughput single-cell RNA and VDJ sequencing, we rapidly identified multiple SARS-CoV-2 neutralizing antibodies from antigen-binding B cells from convalescent COVID-19 patients (Figure 1).![](/research/single_cell_genomics/neutralizing_antibody/neu1.png)
Figure 1. Efficient neutralizing antibody identification through antigen-enriched high-throughput single-cell RNA sequencing. Schematic overview of the neutralizing antibody identification process. The sequence of the mAbs could be obtained within two days using 10X Genomics 5’ VDJ sequencing.
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Figure 2. Affinity specificity and neutralizing abilities of the potent neutralizing mAbs. A) Neutralization potency measured by using a pseudotyped virus neutralization assay. B) Neutralization potency measured by an authentic SARS-CoV-2 plaque reduction neutralizing test (PRNT) assay. C) Characteristics of the neutralizing mAbs. IC50 and IC80 were calculated by using a four-parameter logistic curve-fitting. Kd targeting RBD was measured by using SPR with a 1:1 binding model.
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Figure 3. BD-368-2 showed high therapeutic and prophylactic efficacy in SARS-CoV-2-infected hACE2 transgenic mice. A) Experimental design for therapeutic and prophylactic testing of BD-368-2 in hACE2 transgenic mice. BD-368-2 or unrelated antibody HG1K (20 mg/kg of body weight) was intraperitoneally injected into the transgenic mice before or after SAR-CoV-2 infection. B) Body weight change (%) of the hACE2 transgenic mice recorded over 5 days (one-sided permutation test, *p<0.05). Each group contains 3 mice. C) Virus titers of lung tissue at 5 dpi. The viral loads of the lung were determined by qRT-PCR (one-tailed t-test, ***p<0.001).
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Figure 4. Cryo-EM structure of BD23-Fab in complex with the Spike trimer. A) Cryo-EM structure of the S trimer in complex with BD23-Fab reconstructed at 3.8 Å resolution. The three protomers in the S trimer are depicted in cyan, green, and yellow, respectively. BD23-Fab is depicted in magenta (heavy chain) and blue (light chain). B) N165 glycan in the NTD of protomer C facilitates the interaction between BD23-Fab and the RBD of protomer B. C) The crystal structure of the RBD/ACE2 complex is overlaid onto the RBD/BD23-Fab structure. BD23-Fab would collide with ACE2 and therefore block the interaction between RBD and ACE2. RBD is shown in green and white, whereas ACE2 in orange.
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Figure 5. Characteristics of the neutralizing mAbs identified based on CDR3H structural similarity to SARS-CoV neutralizing mAbs. A) The CDR3 sequence comparison between SARS-CoV neutralizing mAb m396 and the SARS-CoV-2 neutralizing mAbs identified based on CDR3H structure similarity. B) Neutralization potency measured by using a pseudotyped virus neutralization assay. C) Characteristics of the neutralizing mAbs identified based on structure similarity. The CDR3H structure prediction was performed using FREAD. D) The crystal structure of m396-Fab/SARS-CoV-RBD.