How delta and kappa variants of SARS-CoV-2 evade the immune system

Worldwide transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of the ongoing Coronavirus disease 2019 (COVID-19) pandemic, has ultimately led to the emergence of new strains of the virus due to mutations in the spike (S) protein of the virus. The S protein mediates entry into host cells, thus the main target of neutralizing antibodies and most vaccines.

The S glycoprotein comprises two functional subunits, S1 and S2. The S1 subunit contains the receptor-binding domain (RBD) that interacts with the angiotensin-converting enzyme 2 (ACE2) receptor and the N-terminal domain (NTD) that recognized different attachment factors. The S2 subunit comprises fusion machinery that mediated the fusion of the host and viral membrane.

The different variants that have arisen due to mutation include the alpha (α) variant, the beta (β) variant, and the delta (δ) variant. Currently, the latter is globally prevailing and causing a surge in infections among unvaccinated individuals. Most of the mutations involved in creating these variants are localized in the RBD and NTD, targets for therapeutics and vaccines. This raises concerns as to how effective currently available treatments and vaccines are against variants.

A new study, available on the bioRxiv* preprint server, describes the neutralizing ability of the monoclonal antibodies against the variants and the difference of the ACE2 binding affinity found in the variants.

Study: Molecular basis of immune evasion by the delta and kappa SARS-CoV-2 variants

What did the study involve?

The researchers collected blood samples from individuals who had received double doses of Pfizer’s BNT162b2 vaccine, Moderna’s mRNA-1273 vaccine, or J&J/Janssen Ad26.COV2.S vaccine. Following this, recombinant expression of mAb in ExpiCHO cell lines, recombinant RBD glycoprotein production, pseudovirus production, and neutralization of pseudovirus was completed.

Western blot analysis was carried out using the undiluted pseudovirus, and ELISA was carried out using the NTD-targeted mAb. Subsequently, cryo-electron microscopy and ACE2 binding measurements experiments were conducted.

What did the study find?

Comparison of the serum neutralizing activity against the ancestral  S protein and those of the three new variants showed a reduction in neutralizing ability of all three vaccines investigated. Although the decrease in neutralizing ability differed as vaccine type and variant did. Despite the delta variant becoming dominant worldwide by June 2021, the kappa and delta plus variants demonstrated a higher ability to evade vaccine neutralization. The higher incidence of the delta variant owing to enhanced transmissibility, replication kinetics, and viral loads in oropharyngeal and nose-throat swabs of infected individuals relative to the ancestral virus and other variants.

CryoEM structures of the SARS-CoV-2 B.1.617.1 and B.1.617.2 S ectodomain trimers and analysis of ACE2 binding. (A-B) Structure of the B.1.617.1 (A) and B.1.617.2 (B) S trimer (surface rendering) bound to the S2L20 and S309 (A) or S2M11 (B) Fabs (ribbons). SARS-CoV-2 S protomers are colored pink, cyan, and gold, whereas the S2L20 Fab heavy and light chains are colored dark and light green, respectively. The S309 Fab heavy and light chains are colored dark and light orange, respectively (A). The S2M11 Fab heavy and light chains are colored dark and light gray, respectively (B). Only the Fab variable domains are resolved and therefore modeled in the map. N-linked glycans are rendered as dark blue spheres. (C) Zoomed-in view of the S309-bound B.1.617.1 RBD with L452R and E484Q shown as red spheres. (D) Zoomed-in view of the S2M11-bound B.1.617.2 RBD with L452R and T478K shown as red spheres. (E) Superimposition of the LY-CoV555–bound SARS-CoV-2 RBD structure (PDB 7KMG) on the SARS-CoV-2 B.1.617.1 S cryoEM structure show that L452R would clash with the mAb and E484Q would disrupt electrostatic interactions. (F) Superimposition of the CT-P59–bound SARS-CoV-2 RBD structure (PDB 7CM4) on the SARS-CoV-2 B.1.617.2 S cryoEM structure shows that L452R would sterically clash with the mAb. (G) Enzyme-linked immunosorbent assay (ELISA) binding analysis of the SARS-CoV-2 wildtype, B.1.1.7 (α), B.1.617.1 (κ), B.1.617.2 (δ), and B.1.617.2+ (δ+) RBDs to immobilized human ACE2 ectodomain (residues 1-615) shown as 50% effective concentrations (EC50). Data from two biological replicates are shown with 2-4 technical replicates each. (H) Surface plasmon resonance (SPR) binding affinity analysis of the human ACE2 ectodomain (residues 1-615) for immobilized biotinylated wildtype, B.1.1.7, B.1.617.1, B.1.617.2, and B.1.617.2+ RBDs. Data from two biological replicates are shown with 2-6 technical replicates each. (I) Biolayer Interferometry (BLI) binding analysis of the human ACE2 ectodomain (residues 1-615) to immobilized biotinylated SARS-CoV-2 wildtype, B.1.1.7, B.1.617.1, B.1.617.2, and B.1.617.2+ RBDs. Data from two biological replicates are shown with 1-2 technical replicates each. (J-K) Superimposition of the ACE2-bound SARS-CoV-2 RBD structure (PDB 6VW1) on the SARS-CoV-2 B.1.617.1 (J) and B.1.617.2 (K) S cryoEM structures show that L452R and T478K point away from the interface with ACE2, while K417 contacts D30 from ACE2.

Research also showed that mutation in the L452R side chain of the RBD in the S protein of the kappa and delta variants decreased neutralization by antibodies. Mutations in RBD also alters its ability to bind to the ACE2 receptor. Mutations within the antigenic sites of NTD of these variants also resulted in escape from NTD specific monoclonal antibodies.

What did the authors conclude?

The researchers concluded that mutations found in the kappa and delta variants mediate immune evasion by eroding infection- and vaccine-elicited serum neutralizing Ab titers due to structural alteration present in major antigenic sites within the RBD and NTD.

Research is being carried out against multiple additional conserved antigenic sites that can be recognized by RBD-specific monoclonal antibodies and can cope with the emergence of different variants of SARS-CoV-2 for the development of future vaccines.

*Important notice

bioRxiv publishes preliminary scientific reports that are not peer-reviewed and, therefore, should not be regarded as conclusive, guide clinical practice/health-related behavior, or treated as established information.

Written by

Suchandrima Bhowmik

Suchandrima has a Bachelor of Science (B.Sc.) degree in Microbiology and a Master of Science (M.Sc.) degree in Microbiology from the University of Calcutta, India. The study of health and diseases was always very important to her. In addition to Microbiology, she also gained extensive knowledge in Biochemistry, Immunology, Medical Microbiology, Metabolism, and Biotechnology as part of her master's degree.