Identification of HLA-A*02:01-Restricted Candidate Epitopes Derived from the Nonstructural Polyprotein la of SARS-CoV-2 That May Be Natural Targets of CD8+ T Cell Recognition In Vivo

COVID-19 vaccines are being rapidly developed and human trials are under way. Almost all of these vaccines have been designed to induce antibodies targeting the spike protein of SARS-CoV-2 in expectation of neutralizing activities. However, nonneutralizing antibodies are at risk of causing antibody-dependent enhancement. Further, the longevity of SARS-CoV-2-specific antibodies is very short. Therefore, in addition to antibody-inducing vaccines, novel vaccines developed on the basis of SARS-CoV-2-specific cytotoxic T lymphocytes (CTLs) should be considered. Here, we attempted to identify HLA-A*02:01-restricted CTL epitopes derived from the nonstructural polyprotein 1a of SARS-CoV-2. Eighty-two peptides were first predicted as epitope candidates based on bioinformatics. Fifty-four of the 82 peptides showed high or medium binding affinities to HLA-A*02:01. HLA-A*02:01 transgenic mice were then immunized with each of the 54 peptides encapsulated into liposomes. The intracellular cytokine staining assay revealed that 18 out of 54 peptides were active as CTL epitopes because of the induction of gamma interferon (IFN-gamma)-producing CD8 T cells. Of the 18 peptides, 10 peptides were chosen for the following analyses because of their high responses. To identify dominant CTL epitopes, mice were immunized with liposomes containing the mixture of the 10 peptides. Some peptides were shown to be statistically predominant over the other peptides. Surprisingly, all mice immunized with the liposomal 10-peptide mixture did not show the same reaction pattern to the 10 peptides. There were three response patterns, suggesting the existence of an immunodominance hierarchy following peptide vaccination, which may provide more variations in the epitope selection for designing CTL-based COVID-19 vaccines. IMPORTANCE For the development of vaccines based on SARS-CoV-2-specific cytotoxic T lymphocytes (CTLs), we attempted to identify FILA-A*02:01-restricted CTL epitopes derived from the nonstructural polyprotein 1a of SARS-CoV-2. Out of 82 peptides predicted by bioinformatics, 54 peptides showed good binding affinities to HLA-A*02:01. Using HLA-A*02:01 transgenic mice, 18 in 54 peptides were found to be CTL epitopes in the intracellular cytokine staining assay. Out of 18 peptides, 10 peptides were chosen for the following analyses because of their high responses. To identify dominant epitopes, mice were immunized with liposomes containing the mixture of the 10 peptides. Some peptides were shown to be statistically predominant. Surprisingly, all immunized mice did not show the same reaction pattern to the 10 peptides. There were three reaction patterns, suggesting the existence of an immunodominance hierarchy following peptide vaccination, which may provide us more variations in the epitope selection for designing CTL-based COVID-19 vaccines.

Automated summary

In the development of vaccines based on SARS-CoV-2-specific cytotoxic T lymphocytes (CTLs), 55 CTL epitope candidates were identified, with 18 of them validated for their effectiveness in HLA-A*02:01 transgenic mice. 10 peptides with high responses were selected for further analysis to reveal an immunodominance hierarchy following peptide vaccination.