SARS-CoV-2 spike protein antibody: from structure to neutralizing potential

​Over the past few months, we have launched several new coronavirus antibodies and proteins, including the CR3022 clone that was used to identify the SARS-CoV-2 protein crystal structures. Here we review the coronavirus research performed with this antibody.

The SARS-CoV-2 pandemic continues to affect lives around the globe. With no treatments currently available for COVID-19, vaccine and drug development are a priority. Many researchers are pursuing neutralizing antibodies to SARS-CoV-2 that would function to prevent the binding of the virus to its target cell receptors, leading to aggregation of virus particles and subsequent degradation of the virus through antibody-mediated immunomodulatory effects or complement activation.

Overview

• Solving structures
• SARS-CoV-2 antibody epitopes
• Diagnostic or therapeutic potential?

Solving structures

Two recent articles published in Science have reported on the crystal structure of the receptor binding domain (RBD) of the SARS-CoV-2 spike protein ^1,2.

In the first publication, Wrapp et al. found that the binding affinity of the SARS-CoV-2 RBD to ACE2 is 10–20 times greater than the binding affinity of the earlier identified coronavirus, SARS-CoV. However, when neutralizing antibodies for the SARS-CoV RBD (clones S230, m396 and 80R) were tested against the new SARS-CoV-2 RBD, they showed no interaction.

In the second Science publication, Yuan et al. identified the crystal structure of the SARS-CoV-2 RBD ². In this study, they used a combination of biolayer interferometry (BLI)and ELISA to show that a different SARS-CoV neutralizing antibody, the CR3022 clone, binds to the SARS-CoV-2 RBD, though with reduced affinity compared with the SARS-CoV RBD (K_D 115±3 nM and < 0.1 nM for the SARS-CoV-2 and SARS-CoV RBD respectively) ². This interaction of the CR3022 antibody with the SARS-CoV-2 spike protein was confirmed in a further report that stated that the RBD domain of the SARS-CoV-2 spike protein shows a higher affinity for the CR3022 clone than for the host receptor human ACE2 (K_D=6.3 nM for CR3022 vs 15.2 nM for ACE2) ³. Other SARS-CoV neutralizing antibodies tested in this study (m396 and CR3014) showed limited affinity for the RBD domain of SARS-CoV-2.

SARS-CoV-2 antibody epitopes

Both the SARS-CoV-2 spike protein crystal structure and BLI studies suggest that the epitope of the CR3022 SARS-CoV-2 spike protein antibody does not overlap with the RBD-ACE2 binding site ^1-3.

The RBD of the homotrimeric SARS-CoV-2 spike protein switches between ‘up’ and ‘down’ conformations like a hinge. ACE2 can only interact with the RBD when it is in the ‘up’ conformation. Similarly, the CR3022 antibody can only bind to the SARS-CoV-2 RBD when it is present in the ‘up’ conformation and requires that at least two of the spike protein subunits are in this conformation and slightly rotated to overcome steric hindrance at the antibody epitope ². This conformational change in the SARS-CoV-2 spike protein may be physiologically significant as the CR3022 antibody can neutralize SARS-CoV, but did not neutralize SARS-CoV-2 in vitro at the highest tested dose in this study (400 µg/mL).

Diagnostic or therapeutic potential?

Recent analysis of the RBD of the spike protein in SARS-CoV-2 identified it as immunodominant and unique among known human and animal coronaviruses ⁴. Consequently, antibodies to this domain are likely to be highly specific to SARS-CoV-2, and could offer the basis for new COVID-19 antibody tests, revealing if an individual has been exposed to the virus.

Neutralizing therapies could be given to those exposed to the virus, such as a health and social-care workers, to prevent infection, as well as to patients already infected by the virus, to help treat and prevent disease progression. Although the study from Wrapp et al. reported that the CR3022 antibody did not neutralize the SARS-CoV-2 virus in vitro ¹, it cannot be ruled out that CR3022 may protect against SARS-CoV-2 in vivo. Previous studies have seen comparable results for influzena A virus in which the viral HA protein shows a similar change in 'up' and 'down' conformations as the SARS-CoV-2 spike protein ^5-7. Antibodies targeting influenza A virus HA on the surface of the trimer show no neutralizing effect in vitro, but are able to protect individuals against the attack of influenza A virus in vivo ⁸. Similarly, antibodies that do not neutralize in vitro but have protective effects in vivo have been observed for the herpes simplex virus ⁹, cytomegalovirus ¹⁰, alphavirus ¹¹, and dengue virus ¹². Further verification of the neutralizing capacity of the CR3022 SARS-CoV-2 spike protein antibody in animal models is required to fully understand the potential of this antibody in COVID-19.

Some have reported on the therapeutic potential of the CR3022 antibody to function in combination with a second antibody for the SARS-CoV-2 RBD, CR3014, which targets a different epitope on the RBD and overlaps with ACE2 binding ^{3, 13}. Earlier research showed that in combination these antibodies could effectively control the immune escape caused by SARS-CoV to increase the host's resistance to attack from a range of mutant viral strains ¹³, suggesting such effects may also be seen with SARS-CoV-2.

Despite its unique binding mechanism to the SARS-CoV-2 spike protein and lack of overlap with the ACE-RBD, the CR3022 antibody for the SARS-CoV-2 spike protein may still effectively neutralize SARS-CoV-2 in vivo, or could be used as part of a neutralizing antibody combination for the treatment of COVID-19.

Abcam’s products are intended for research use only, not for use in diagnostic, therapeutic or any other purposes.

References

1. Wrapp, D. et al. Cryo-EM structure of the 2019-nCoV spike in the prefusion conformation. Science 367, 1260-1263 (2020).
2. Yuan, M. et al. A highly conserved cryptic epitope in the receptor binding domains of SARS-CoV-2 and SARS-CoV. Science 368, 630-633 (2020).
3. Tian, X. et al. Potent binding of 2019 novel coronavirus spike protein by a SARS coronavirus-specific human monoclonal antibody. Emerging microbes & infections 9, 382-385 (2020).
4. Premkumar, L. et al. The receptor binding domain of the viral spike protein is an immunodominant and highly specific target of antibodies in SARS-CoV-2 patients. Science Immunol 5(48), eabc8413 (2020).
5. Bangaru, S. et al. A Site of Vulnerability on the Influenza Virus Hemagglutinin Head Domain Trimer Interface. Cell 177, 1136-1152 e1118 (2019).
6. Watanabe, A. et al. Antibodies to a Conserved Influenza Head Interface Epitope Protect by an IgG Subtype-Dependent Mechanism. Cell 177, 1124-1135 e1116 (2019).
7. Bajic, G. et al. Influenza Antigen Engineering Focuses Immune Responses to a Subdominant but Broadly Protective Viral Epitope. Cell host & microbe 25, 827-835 e826 (2019).
8. Lee, J. et al. Molecular-level analysis of the serum antibody repertoire in young adults before and after seasonal influenza vaccination. Nature medicine 22, 1456-1464 (2016).
9. Petro, C. et al. Herpes simplex type 2 virus deleted in glycoprotein D protects against vaginal, skin and neural disease. eLife 4 eLife.06054 (2015).
10. Bootz, A. et al. Protective capacity of neutralizing and non-neutralizing antibodies against glycoprotein B of cytomegalovirus. PLoS pathogens 13, e1006601 (2017).
11. Burke, C. W. et al. Human-Like Neutralizing Antibodies Protect Mice from Aerosol Exposure with Western Equine Encephalitis Virus. Viruses 10 (4), 147 (2018).
12. Henchal, E. A., Henchal, L. S. & Schlesinger, J. J. Synergistic interactions of anti-NS1 monoclonal antibodies protect passively immunized mice from lethal challenge with dengue 2 virus. The Journal of general virology 69 ( Pt 8), 2101-2107 (1988).
13. ter Meulen, J. et al. Human monoclonal antibody combination against SARS coronavirus: synergy and coverage of escape mutants. PLoS medicine 3, e237 (2006).