A pair of noncompeting neutralizing human monoclonal antibodies protecting from disease in a SARS‐CoV‐2 infection model

Abstract

TRIANNI mice carry an entire set of human immunoglobulin V region gene segments and are a powerful tool to rapidly isolate human monoclonal antibodies. After immunizing these mice with DNA encoding the spike protein of SARS‐CoV‐2 and boosting with spike protein, we identified 29 hybridoma antibodies that reacted with the SARS‐CoV‐2 spike protein. Nine antibodies neutralize SARS‐CoV‐2 infection at IC50 values in the subnanomolar range. ELISA‐binding studies and DNA sequence analyses revealed one cluster of three clonally related neutralizing antibodies that target the receptor‐binding domain and compete with the cellular receptor hACE2. A second cluster of six clonally related neutralizing antibodies bind to the N‐terminal domain of the spike protein without competing with the binding of hACE2 or cluster 1 antibodies. SARS‐CoV‐2 mutants selected for resistance to an antibody from one cluster are still neutralized by an antibody from the other cluster. Antibodies from both clusters markedly reduced viral spread in mice transgenic for human ACE2 and protected the animals from SARS‐CoV‐2‐induced weight loss. The two clusters of potent non‐competing SARS‐CoV‐2 neutralizing antibodies provide potential candidates for therapy and prophylaxis of COVID‐19. The study further supports transgenic animals with a human immunoglobulin gene repertoire as a powerful platform in pandemic preparedness initiatives. This article is protected by copyright. All rights reserved

A surrogate cell‐based SARS‐CoV‐2 spike blocking assay

“…5C ). These results are consistent with virus neutralization assays [ 35 ], thereby, confirming the specificity of SUBA for the use of SARS‐CoV‐2 spike mutations.…”

“…S1 ), we preferred RSp cells because they grew faster and did not require trypsinization. To test whether anti‐SARS‐CoV‐2 RBD antibodies can indeed block the binding of Rsp cells to hACE2, we incubated RSp cells with increasing concentrations of a fully human recombinant anti‐RBD neutralizing antibody (TRES224) that was established in our lab [ 34 , 35 ]. In contrast, neither TRES328, a recombinant antibody recognizing the N‐terminal domain (NTD) of the SARS‐CoV‐2 spike protein, nor the isotype‐matched control IgG1 control antibody (TRES 567.II) showed a specific blocking activity (Fig.…”

“…To directly show that the SUBA assay can distinguish between two antibodies that differ in their RBD‐blocking strength, we titrated the monoclonal anti‐RBD antibody TRES224 [ 34 , 35 ] and the clinically tested anti‐RBD antibody R10933 [ 19 ] (Fig. 3E ) either diluted in PBS or serum from an SARS‐CoV‐2‐negative serum donor.…”

“…While also antibodies recognizing the NTD of SARS‐CoV‐2 can neutralize the virus in vivo [ 35 ], SUBA can only specifically measure the disruption of the hACE2‐RBD interaction. Nevertheless, in summary, we propose that SUBA is a fast, inexpensive, reproducible, and valid method to monitor the COVID‐19 vaccination efficacy and the recovery from COVID‐19.…”

Increased risk of chronic fatigue and hair loss following COVID-19 in individuals with hypohidrotic ectodermal dysplasia

“…Stained cell populations were analyzed using a Gallios flow cytometer (Beckman-Coulter). The antibody TRES 224 [15] recognizing the SARS-CoV-2 spike protein served as positive control.…”