Have scientists found SARS-CoV-2’s Achilles’ heel?

By Jocelyn Solis-Moreira Mar 29 2021

Have scientists uncovered the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Achilles’ heel? New preclinical research published in the journal Cell shows how neutralizing antibodies targeting the N-terminal domain of the coronavirus could be a potential target for antibodies to bind and effectively neutralize SARS-CoV-2.

Study: N-terminal domain antigenic mapping reveals a site of vulnerability for SARS-CoV-2. Image Credit: FURMANCHUK LARISA / Shutterstock

Most neutralizing antibodies from patient plasma and COVID-19 vaccines target the spike protein’s receptor binding domain to prevent the spike protein from binding to ACE2 receptors in human cells.

Researchers found 5%-20% of monoclonal antibodies created from memory B cells target the N-terminal domain. The new findings suggest areas outside the receptor-binding domain could also be effective and may help develop future treatments.

“Our cryo-EM structures of SARS-CoV-2 S bound to the ultrapotent RBD-specific mAb S2M11 and to S2X333, S2M28, or S2L28, together with evidence of additive neutralizing effect of RBD- and NTD-targeted mAbs, provide proof of concept for implementing countermeasures using mAb cocktails targeting both the NTD and the RBD and potentially also for vaccine design,” writes the team.

How they did it

The researchers tracked the antibody responses from the convalescent plasma of three patients who were positive for COVID-19 infection and 41 monoclonal antibodies targeting the N-terminal domain. Using cryoelectron microscopy, binding assays and antibody escape mutant analysis, the researchers designed an antigenic map of the N-terminal domain and classified supersites that would be recognized by neutralizing antibodies.

Antibodies targeting the N-terminal domain prevent SARS-CoV-2 infection

The antigenic map contained the heavily glycosylated SARS-CoV-2 N-terminal domain where the researchers identified six antigenic sites. Based on the three convalescent samples, all of the neutralizing antibodies, including monoclonal antibody 4A8, FC05, and CM25 — created from memory B cells target the same antigenic supersite labeled as site (i) using several germline V genes.

Syrian hamsters were challenged with SARS-CoV-2 to test the antibody’s ability to prevent an infection. The findings showed neutralizing antibodies targeting the N-terminal domain protected the hamsters from SARs-CoV-2. This suggests a potential benefit for therapeutic candidates.

N-terminal domain mutations evolved under selective pressure from humoral immunity

Single amino acid mutations to the spike protein receptor-binding domain such as N501Y and E484K have helped evade the immune system and decrease the effectiveness of neutralizing antibodies. The researchers sought to find if targeting the N-terminal domain would help reduce the likelihood of escape mutations.

They analyzed 508, 771 SARS-CoV-2 genome sequences available in the GISAID database as of February 12, 2021. They found the N-terminal domain harbored various mutations and deletions comparable to the spike protein’s mutations.

All neutralizing antibodies specific to the N-terminal domain were successful in binding to variants that carried a 69/70 deletion or the A222V substitution. Further analysis showed that having neutralizing antibodies targeting the N-terminal domain activated FcgRIIa and FcgRIIIa in vitro. However, this activation did not happen when antibody S2M24 targeted site vi of the N-terminal domain.

Based on the results, the researchers suggest the Fc-mediated effector could increase the activity of N-terminal domain-specific antibodies against SARS-CoV-2. It also provides evidence that antibodies targeting site i of the N-terminal domain would effectively neutralize the COVID variants.

However, variants of concerns — B.1.1.7, B.1.351, and P.1 — have several mutations, such as the L18F substitution and the deletion of Y144 residue in the N-terminal domain that could help it evade neutralizing antibodies. With this in mind, the researchers found other key residues for binding neutralizing antibodies, such as indirect binding of S12 and direct binding of R246 that could help with variants evasion.

Many circulating variants have mutations in the N-terminal domain, including in the antigenic site i, which may have been the result of selective pressure for the virus to evolve and escape neutralization.

“We anticipate that the protection efficacy of S2X333 (and related NTD mAbs) would be further enhanced in humans due to an optimal matching with human Fcg receptors. However, our results also indicate that potent NTD mAbs targeting the antigenic supersite (site i) impose a selection pressure driving viral evolution to escape neutralization,” write the research team.

The team suggests that having monoclonal antibody therapies that include antibodies targeting the N-terminal domain would provide broader protection and increasing the likelihood of neutralizing SARS-CoV-2 variants.