Key Considerations for the Development of Safe and Effective SARS-CoV-2 Subunit Vaccine: A Peptide-Based Vaccine Alternative

Abstract

COVID-19 is disastrous to global health and the economy. SARS-CoV-2 infection exhibits similar clinical symptoms and immunopathological sequelae to SARS-CoV infection. Therefore, much of the developmental progress on SARS-CoV vaccines can be utilized for the development of SARS-CoV-2 vaccines. Careful antigen selection during development is always of utmost importance for the production of effective vaccines that do not compromise recipient safety. This holds especially true for SARS-CoV vaccines, as several immunopathological disorders are associated with the activity of structural and nonstructural proteins encoded in the virus's genetic material. Whole viral protein and RNA-encoding full-length proteins contain both protective and "dangerous" sequences, unless pathological fragments are deleted. In light of recent advances, peptide vaccines may present a very safe and effective alternative. Peptide vaccines can avoid immunopathological pro-inflammatory sequences, focus immune responses on neutralizing immunogenic epitopes, avoid off-target antigen loss, combine antigens with different protective roles or mechanisms, even from different viral proteins, and avoid mutant escape by employing highly conserved cryptic epitopes. In this review, an attempt is made to exploit the similarities between SARS-CoV and SARS-CoV-2 in vaccine antigen screening, with particular attention to the pathological and immunogenic properties of SARS proteins.

Keywords: SARS-CoV-2; angiotensin-converting enzyme 2; antibody-dependent enhancement; critical binding residues; neutralizing antibodies; receptor binding domain; spike protein; type-I interferons.

Figure 1

SARS‐2 genome, spike protein, and receptor binding domain (RBD). A) Viral RNA‐encoding structural (S, E, M, and N) and nonstructural proteins (NSPs). B) S‐protein subdomains. C) Sequence and structural conformation of RBD. In the RBD sequence: the residues highlighted in red are critical ACE2‐receptor binding residues; grey highlighted residues are the binding motif (RBM); the blue arrows indicate strand structure; the red cylinders indicate helical structure; and the black bonds between cysteine residues are native disulfide bonds.

Figure 2

Schematic representation of A) binding, B) priming, C) S‐protein conformational changes, and the D) fusion process of SARS‐2 or SARS‐1 to a host cell bearing ACE2 receptor. A) SARS‐2 virus binds to the ACE2 receptor of a host cell via S‐protein RBD. B) Host proteases prime the S‐protein intra‐, or extracellularly. C) S‐protein adopts a hairpin coiled coils conformation and exposes the fusion peptide. D) Primed S‐protein with exposed fusion peptide drives virus fusion to the host cell, and viral RNA is injected for intracellular translation and transcription, and replication.

Figure 3

Comparison of SARS S‐proteins. A) Graphically aligned SARS‐2‐RBD (purple, from protein database, PDB: 6M0J) and SARS‐1‐RBD (blue, from PDB: 2AJF). SARS‐2‐RBM is marked in green. B) Graphically aligned RBDs of both viruses with a surface mesh electrostatic map of S‐protein RBM (grey) in contact with ACE2, produced using Schrodinger Bioluminate software. The close similarity between these two viral RBDs suggests the presence of common neutralizing epitopes.

Figure 4

Schematic representation depicting IFN‐I production and signaling pathways. Pathways are depicted using black arrows between involved cofactors, adaptors, signal proteins and enzymes. Two pathways are involved in IFN‐I production: A) TLR‐TRAF3, and B) RIG‐1/MDA5‐MAVS, which are blocked by structural and nonstructural SARS‐1 proteins (blue lines and red x marks). C) A signaling pathway for IFN‐I production: IFNARs‐STAT1/2, which is blocked by nonstructural proteins ORF‐3a, ORF‐6, and NSP‐1. Simultaneously, inflammatory cytokine production pathways are stimulated through the TLR‐MAPK pathway (blue arrows) driving the translocation of NF‐κB or AP‐1 via S‐protein, N‐protein, and ORF‐3b.

Figure 5

Alignment of SARS‐2, SARS‐1, and RaTG13 structural S‐protein sequences, as generated by ClustalW. The black and blue boxes represent T‐cell epitopes that bind in humans and mice, respectively. The thin and bold boxes represent experimentally immunogenic cytotoxic T‐cell/MHC‐I epitopes and CD4⁺ T‐cell/MHC‐II epitopes, respectively. The orange boxes show experimentally immunogenic or neutralizing S‐RBD B‐cell epitopes, while the green boxes show experimental immunopathological sequences.

Figure 6

Alignment of SARS‐2, SARS‐1, and RaTG13 structural E‐, M‐, and N‐proteins, as generated by ClustalW. The black and blue boxes represent T‐cell epitopes that bind in humans and mice, respectively. The thin and bold boxes represent experimentally immunogenic cytotoxic T‐cell/MHC‐I epitopes and CD4⁺T‐cell/MHC‐II epitopes, respectively. The orange boxes show experimentally immunogenic B‐cell epitopes, and the green boxes are experimental immunopathological sequences.

Figure 7

Dose response curves for A) mean IgG neutralizing Ab (nAb) titers and B) mean total anti‐S‐protein IgG titers expressed as the mean reciprocal serum dilution (n = 6–7 mice per group). Both are plotted against mouse survival, following the administration of SARS‐1‐S‐protein as antigen with TLR‐3‐ligand (poly I:C adjuvant) or without adjuvant.^{[ 154 ]} Log₁₀ of the total anti‐S‐protein titers of 3.5 offers 100% protection in (B), which was equivalent in protective efficacy to neutralizing Ab (nAb) log₁₀ titers of 2 in (A), as both plots are from the same serum samples, (C) relationship between human immune sera against SARS‐2 and neutralization, fitting log IgG concentration and log SARS‐2 neutralization titer 50%, gave  LogVNT₅₀ =   − 1.53 + 0.94 · Log IgG_{Anti − RBD}, n = 59, R² = 0.86.^{[ 149}, ^{150 ]}

Figure 8

Schematic representation of subunit vaccines’ evolution from A) whole‐pathogen (SARS‐2), to B) subunit protein (S‐protein), to C) immunogenic protein fragment (RBD), and finally D) peptide‐based vaccine antigens (red, pep1; blue, pep2).

Figure 9

Binding scores using the i‐tasser SPRING server for RBD and several RBD‐fragments to ACE2.^{[ 188 ]}