Super-antibody UT28K shows broad-spectrum activity against SARS-CoV-2 variants

Even as the Omicron variant of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to circulate, causing thousands of new cases and breakthrough infections due to its immune evasion characteristics, a new research paper describes a virus-neutralizing antibody that is active against Omicron as well as several other variants.

Study: Novel super-neutralizing antibody UT28K is able to protect against infection from a wide variety of SARS-CoV-2 variants. Image credit: Leonid Altman / Shutterstock

Introduction

The coronavirus disease 2019 (COVID-19) pandemic is far from over, although the number of reported hospitalizations and deaths has declined significantly in some parts of the world. SARS-CoV-2 has homotrimeric spike protein antigens on its surface, with each spike monomer comprising S1 and S2 subunits. The first of them has a receptor binding domain (RBD) and an N-terminal domain (NTD).

The RBD mediates attachment of the host cell by the virus as the first step in the infection process. The attachment is to the host angiotensin converting enzyme receptor 2 (ACE2) which is expressed on a variety of human cells. Neutralizing antibodies often work by inhibiting the interaction between ACE2 and RBD, thereby preventing SARS-CoV-2 infection.

A number of monoclonal antibodies (mAbs) have been isolated from convalescent serum, with activity against RBD. The mAb cocktail casirivimab (REGN10933) and imdevimab (REGN10987) have been approved for the treatment of early COVID-19, as has the mAb sotrovimab (VIR-7831).

New variants of concern (VOCs) such as Alpha, Beta, Gamma, and Delta were susceptible to these mAbs, although to varying degrees. However, Omicron showed immune evasion, resistant to neutralization by casirivimab and imdevimab, likely due to mutations at the ACE2 binding site of the RBD, an antigenic supersite. On the other hand, sotrovimab continues to show neutralizing activity, although at 3-fold lower levels.

The current study, published online in the journal AcMdescribes a novel mAb, UT28K, which has promising broad neutralizing activity against the virus.

What did the study show?

In this study, researchers isolated neutralizing mAbs from peripheral blood mononuclear cells of a COVID-19 patient who recovered from severe illness assuming strongly neutralizing antibodies would be detected. Neutralization assays showed activity against pseudoviruses expressing the D614G spike, as well as several other COV spike variants.

This serum was tested for single anti-RBD antibodies using the immunospot array on a chip (ISAAC) assay. The result was the discovery of UT28K, one of five clones that inhibits spike-ACE2 binding.

UT28K showed picomolar inhibitory concentrations against SARS-CoV-2 VOCs, but 10-fold lower neutralizing activity against Omicron. In other words, while the half-maximal inhibitory concentration (IC50) against previous COVs ranged from 40 to 120 pM, it rose to between 500 and 5000 pM against Omicron compared to wild-type virus while retaining its neutralizing capacity.

Structures of SARS-CoV-2 S-protein-bound UT28K antibody and their interactions.  (a) Cryo-EM structure of Fab UT28K bound to SARS-CoV-2 protein S trimer.  The heavy and light chains of Fab UT28K are shown in pink and navy blue respectively.  The S1 and S2 subunits are shown in gray and black, respectively.  N-linked glycans are shown in cyan.  (b) The crystal structure of Fab UT28K bound to the SARS-CoV-2 S RBD protein.  Fab UT28K colors are the same as shown in A. SARS-CoV-2 S RBD is shown in green.  (c) A comparison of the binding modes of the UT28K and 253XL55 antibodies (VH; yellow and VL; orange) bound to the SARS-CoV-2 S RBD protein.  (dg) Interactions of key residues between Fab UT28K and the SARS-CoV-2 S RBD.

Structures of SARS-CoV-2 S-protein-bound UT28K antibody and their interactions. (a) Cryo-EM structure of Fab UT28K bound to SARS-CoV-2 protein S trimer. The heavy and light chains of Fab UT28K are shown in pink and navy blue respectively. The S1 and S2 subunits are shown in gray and black, respectively. N-linked glycans are shown in cyan. (b) The crystal structure of Fab UT28K bound to the SARS-CoV-2 S RBD protein. Fab UT28K colors are the same as shown in A. SARS-CoV-2 S RBD is shown in green. (c) A comparison of the binding modes of the UT28K and 253XL55 antibodies (VH; yellow and VL; orange) bound to the SARS-CoV-2 S RBD protein. (dg) Interactions of key residues between Fab UT28K and the SARS-CoV-2 S RBD.

In a Syrian hamster model, intraperitoneal use of UT28K protected against infection by SARS-CoV-2 mutants or pseudoviruses when the latter was introduced into the trachea. The trachea and lungs were almost completely free of virus after 24 hours. The protective dose was 0.3 mg/kg for Alpha, Gamma, Delta and Kappa, but ten times higher for Beta and Omicron. This shows a strong protective effect against all variants, although weaker for the last two.

Further analysis showed that unlike other neutralizing antibodies first identified here, UT28K uses a different pair of antibody genes (IGHV1-58/IGKV3-20) that produce highly avid RBD binding and neutralizing powerful of the virus. Structural studies using cryo-electron microscopy (cryo-EM) of the spike protein bound to the antibody-binding fragment (Fab) of UT28K showed the same mode of binding as with earlier mAbs that use these genes. X-ray crystallography confirmed these findings, with the RBD F486 residue interacting with the hydrophobic pocket of the antibody.

Side chain linkage was also present with RBD residue Q493 and the Fab. In addition, some unique interactions were observed, with the side chain of RBD N487 and the main chain residues S477 and T478, forming hydrogen bonds at various points, with the oxygen and nitrogen atoms of the main and side chains Fab.

UT28K did not react with RBD mutants carrying F486S or N487R, but other mutations did not affect UT28K binding, showing that the above two are essential for binding to this mAb. This again supports its use of IGHV1-58/IGKV3-20. Finally, when grown in an environment containing UT28K, the emergence of the Y489H mutation is favored.

What are the implications?

The identification of public antibodies that use this gene pair was reported earlier during research on mAbs that neutralize SARS-CoV-2 and other similar viruses. RBD mutants bound by these mAbs must have F486, of necessity. In this study, UT28K is a public mAb from this group, with a sequence that closely resembles others in this group.

However, unlike the others, its Fab distinctly interacts with the main chain of the RBD spike, a feature that may explain the broad neutralization spectrum of this mAb. It is similar in this way to a broad neutralizing anti-HIV1 mAb that also has interactions with the virus mediated through main chain linkages.

The main chain interactions of UT28K involved A475V, G476S, S477N, but it does not bind with side chain mutants F486S and N487R. RBD residues N487 and Y489 appear to help correctly orient residue F486 for binding to UT28K at the hydrophobic pocket, and mutations at these sites allow antibody leakage. The infamous E484 residue of the RBD which is a known immune escape mutation site, especially in beta and Omicron COVs, is outside the UT28K binding site

These two elements seem to explain the largely neutralizing activity of UT28K, although the Q493R mutation may be the cause of the weakened neutralization against Omicron. This could be due to the elimination of hydrogen bonds or because of steric hindrance.

Mutations at F486, N487, and Y489 could cause UT28K breakout, but since the first two appear to be essential for RBD-ACE2 interactions, and since mutations at these sites are extremely rare so far, it appears that they are not likely to be a publish.

The F486, N487 and Y489 mutations are likely losing their competitive advantage over circulating SARS-CoV-2. As such, the emergence of a SARS-CoV-2 variant resistant to UT28K knockdown is unlikely.. These data suggest that UT28K is a new viable mAb. UT28K will likely confer potent protection against SARS-CoV-2 mutants, including new emerging variants, as well as vaccines.”

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