Computationally validated SARS-CoV-2 CTL and HTL Multi-Patch vaccines, designed by reverse epitomics approach, show potential to cover large ethnically distributed human population worldwide

Abstract

The SARS-CoV-2 (Severe Acute Respiratory Syndrome Coronavirus 2) is responsible for the COVID-19 outbreak. The highly contagious COVID-19 disease has spread to 216 countries in less than six months. Though several vaccine candidates are being claimed, an effective vaccine is yet to come. A novel reverse epitomics approach, 'overlapping-epitope-clusters-to-patches' method is utilized to identify the antigenic regions from the SARS-CoV-2 proteome. These antigenic regions are named as 'Ag-Patch or Ag-Patches', for Antigenic Patch or Patches. The identification of Ag-Patches is based on the clusters of overlapping epitopes rising from SARS-CoV-2 proteins. Further, we have utilized the identified Ag-Patches to design Multi-Patch Vaccines (MPVs), proposing a novel method for the vaccine design. The designed MPVs were analyzed for immunologically crucial parameters, physiochemical properties and cDNA constructs. We identified 73 CTL (Cytotoxic T-Lymphocyte) and 49 HTL (Helper T-Lymphocyte) novel Ag-Patches from the proteome of SARS-CoV-2. The identified Ag-Patches utilized to design MPVs cover 768 overlapping epitopes targeting 55 different HLA alleles leading to 99.98% of world human population coverage. The MPVs and Toll-Like Receptor ectodomain complex shows stable complex formation tendency. Further, the cDNA analysis favors high expression of the MPVs constructs in a human cell line. We identified highly immunogenic novel Ag-Patches from the entire proteome of SARS CoV-2 by a novel reverse epitomics approach and utilized them to design MPVs. We conclude that the novel MPVs could be a highly potential novel approach to combat SARS-CoV-2, with greater effectiveness, high specificity and large human population coverage worldwide. Communicated by Ramaswamy H. Sarma.

Keywords: Ag-Patch (antigenic patch); COVID-19; Coronavirus; Multi-Epitope Vaccine; Multi-Patch Vaccine; Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2); Toll-Like Receptor (TLR); epitope; overlapping-epitope-clusters-to-patches; reverse epitomics.

Graphical abstract

Figure 1.

Schematic presentation for revers epitomics approach, ‘overlapping-epitope-clusters-to-patches’ method. (A) Schematic presentation of the process of epitope presentation by the antigen-presenting cells facilitated by TAP transporter and HLA classes I & II alleles; (B) presentation of ‘overlapping-epitope-clusters-to-patches’ method (Steps 1 & 2) to identify antigenic patches (Ag-Patches) from SARS-CoV-2/pathogen. The cluster of overlapping epitopes leads us to identify antigenic patches from the source protein, a reverse epitomics approach. The identified antigenic patches could be used to design Multi-Patch Vaccine as shown in Step 3.

Figure 2.

Immunogenic Ag-Patches (antigenic patches) identified from overlapping CTL epitope clusters of SARS-CoV-2 proteins. Immunogenic Ag-Patches are identified (red amino acid sequences) on the basis of the overlapping epitope clusters by the revers epitomics approach, ‘Overlapping-epitope-clusters-to patches’ method.

Figure 3.

Immunogenic Ag-Patches (antigenic patches) identified from overlapping HTL epitope clusters of SARS-CoV-2 proteins. Immunogenic Ag-Patches identified (blue amino acid sequences) on the basis of the overlapping epitope clusters by the revers epitomics approach, ‘Overlapping-epitope-clusters-to patches’ method.

Figure 4.

The identified immunogenic Ag-Patches (antigenic patches) shown in the tertiary structure models of the SARS-CoV-2 proteins. The tertiary structure models of all the SARS-CoV-2 proteins have been retrieved from I-TASSER homology modeling server (https://zhanglab.ccmb.med.umich.edu/COVID-19/). The CTL Ag-Patches are shown in red, and the HTL Ag-Patches are shown in blue color. The identification of novel Ag-Patches has been done by the reverse epitomics approach, ‘overlapping-epitope-clusters-to-patches’ method. Most of the Ag-Patches identified in the SARS-CoV-2 proteins are observed to be on the exposed surface of the proteins.

Figure 5.

The CTL and HTL Ag-Patches were utilized to design Multi-Patch Vaccine. Short peptide linkers GGGGS and EAAK were used to fuze the Ag-Patches (antigenic patches) and the adjuvants, respectively. The CTL MPV constructs includes (A) CTL-MPV-1, (B) CTL-MPV-2 and (C) CTL-MPV-3. The HTL MPV construct includes (D) HTL-MPV-1 and (E) HTL-MPV-2 (Supplementary Table S6). CTL and HTL Patches are shown in red and blue, respectively; adjuvants, linkers and 6xHis tags are shown in gray, green and magenta, respectively.

Figure 6.

Tertiary structure modelling of CTL and HTL Multi-Patch Vaccines. Tertiary structural models of CTL and HTL MPVs have been shown in both cartoon and surface presentations. (A) CTL-MPV-1, (B) CTL-MPV-2, (C) CTL-MPV-3, (D) HTL-MPV-1 and (E) HTL-MPV-2. The Ag-Patches are shown in Red (CTL Ag-Patches) and blue (HTL Ag-Patches). Linkers are in green, adjuvants are in gray and 6xHis tag is shown in magenta.

Figure 7.

Molecular docking study of CTL and HTL MPVs with TLR3-ECD complexes. (A, B, C, D, E): Complex formation by the MPVs (CTL-MPV-1, CTL-MPV-2, CTL-MPV-3, HTL-MPV-1, HTL-MPV-2) and TLR3-ECD, respectively. (A-I, B-IV, C-VII, D-X, E-XIII): B-factor analysis presentation for the docked MPVs to the TLR3-ECD. The presentation is in VIBGYOR color, with blue showing low B-factor and red show high B-factor. Here, most of the MPV regions are in blue showing low B-factor and hence suggesting a stable complex formation with TLR3-ECD. (A-II, B-V, C-VIII, D-XI, E-XIV): Binding site formed by the MPVs and TLR3-ECD at the molecular interaction interface, represented in magenta-binding patches. All the CTL and HTL MPVs have shown to form prominent and close binding site region with TLR3-ECD. (A-III, B-VI, C-IX, D-XII, E-XV): Representation of hydrogen bond formation is shown. The residues involved in hydrogen bond formation from MPVs are shown in cyan, and the residues from TLR3-ECD are shown in magenta. The hydrogen bonds are shown by yellow dots.