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How the virus transforms itself to escape antibodies?

The spike (S) protein of SARS-CoV-2 is the principal target of vaccines but also of antibody treatments. It is organized in trimers (the association of 3 identical proteins) on the surface of the virus and is made up of the S1 domain, containing the RBD (Receptor Binding Domain) which interacts with the ACE2 cellular receptor, and the S2 domain, which governs the virus/cell fusion process. Neutralizing antibodies mainly target  the RBD and NTD (N-Terminal Domain) of the S1, and block the virus’ entry. Even though SARS-CoV-2 mutates less quickly than other RNA viruses, the appearance of variants shows that its antigenic drift, in other words, the gradual appearance of mutations, may cause problems for the current strategies of vaccination and antibody therapies.

Exactly how these surface proteins evolve is yet to be fully understood. Researchers at Harvard University (USA) have just updated the spike protein’s antigenic drift in an immunocompromised patient (with depleted B cells) who was chronically infected by SARS-CoV-2.

They first isolated, cloned and produced several monoclonal antibodies (each targeting one region of the spike) using samples from 4 convalescent patients (C1 to C4), of which 43 were isolated from the C1 patient. Using in vitro tests, they showed that all of these antibodies have affinities for the spike as well as variable neutralization capacities. They identified their respective fixation sites through crystallography and made predictions as to escape possibilities of current variants.

Secondly, they took several samples from the chronically ill patient, then sequenced the spike gene in order to track the emergence of  mutations. Several variants appeared without achieving dominance, until the 128th day following COVID-19 diagnosis (Day 0), when the Y489H mutation of the RBD became dominant. On day 145, the chronically ill patient received an emergency therapeutic cocktail REGN-COV2, made up of two monoclonal antibodies, which caused the emergence of 2 dominant variants on days 146 and 152. These variants had been present previously, but at a low level. They included 8 mutations in the RBD (with Y489H), of which 7 are present in the UK, South African and Brazilian variants.

This can be schematically represented as:

Finally, they reintroduced these mutations into the viral particles (“pseudotyped”) in order to evaluate in vitro their potential of being neutralized by the antibodies. To validate their structural predictions, they showed that the N501Y and Q493K mutations allow escape from therapeutic antibodies, or monoclonal and polyclonal (a mixture of antibodies targeting several areas of the spike)  antibodies taken from convalescent patients. These two spike mutations facilitate the interaction with the Y41 and K453 amino acids of the ACE2 by hydrophobic contacts. However, from day 152 the affinity of the pseudotype for the ACE2 diminished 10-fold because the F486I mutations suppresses some of the ACE2 hydrophobic contacts. Days 146 and 152 contain deletions (or losses) in the NTD (141-144) that are also present in the UK and South African variants. These deletions prevent certain antibodies from binding successfully, allowing the day 146 and day 152 mutations to evade their action. Patient C1 is a very particular example: his antibody response was so focused on the RBD that the single Q493K mutation was enough to enable escape. However, this mutation did not enable escape from the antibodies of the 3 other convalescent patients.

This can be schematically represented as:

So the results show that the virus develops solutions to adapt to the ACE2 while escaping neutralization by the antibodies, even if it means modulating its affinity for the receptor. Although this study was carried out on a single patient  and did not use real viruses for its in vitro tests, it shows how mutations that can be found in current variants associate, and how some of these associations allow the virus to escape a large number of antibodies. In vivo studies on the evolution of the spike protein are therefore essential so we can predict those mutations that will probably arise in new variants.

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