Construction and efficacy testing of DNA vaccines containing HLA-A*02:01-restricted SARS-CoV-2 T-cell epitopes predicted by immunoinformatics

Abstract

Vaccines play essential roles in the fight against the COVID-19 pandemic. The development and assessment of COVID-19 vaccines have generally focused on the induction and boosting of neutralizing antibodies targeting the SARS-CoV-2 spike (S) protein. Due to rapid and continuous variation in the S protein, such vaccines need to be regularly updated to match newly emerged dominant variants. T-cell vaccines that target MHC I- or II-restricted epitopes in both structural and non-structural viral proteins have the potential to induce broadly cross-protective and long-lasting responses. In this work, the entire proteome encoded by SARS-CoV-2 (Wuhan-hu-1) is subjected to immunoinformatics-based prediction of HLA-A*02:01-restricted epitopes. The immunogenicity of the predicted epitopes is evaluated using peripheral blood mononuclear cells from convalescent Wuhan-hu-1-infected patients. Furthermore, predicted epitopes that are conserved across major SARS-CoV-2 lineages and variants are used to construct DNA vaccines expressing multi-epitope polypeptides. Most importantly, two DNA vaccine constructs induce epitope-specific CD8+ T-cell responses in a mouse model of HLA-A*02:01 restriction and protect immunized mice from challenge with Wuhan-hu-1 virus after hACE2 transduction. These data provide candidate T-cell epitopes useful for the development of T-cell vaccines against SARS-CoV-2 and demonstrate a strategy for quick T-cell vaccine candidate development applicable to other emerging pathogens.

1. Introduction

The global COVID-19 pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) first emerged at the end of 2019 and eventually transitioned from the emergency phase to the endemic phase after three years [1]. Vaccines produced by both classical vaccine platforms, such as inactivated virus, recombinant viral vector and recombinant subunit vaccines, and novel platforms, most notably synthetic mRNA-based vaccines, play vital roles in ameliorating the damage caused by viruses [2]. The development and evaluation of SARS-CoV-2 vaccine candidates have generally focused on their ability to induce and boost neutralizing antibodies against the viral spike protein (S), which is responsible for receptor binding and viral entry [3]. However, immune selection by infection- and vaccination-induced antibodies resulted in significantly quicker variation in S than in response to other structural and non-structural proteins [4]. Consequently, currently licensed vaccines require regular updates to provide sufficient protection against newly emerged dominant SARS-CoV-2 variants [5,6]. On the other hand, antibody titers have been shown to decrease rather rapidly in both vaccine recipients and infected patients, resulting in weakened protection against (re)infection that necessitates additional booster injections [7,8].

In addition to S-targeting neutralizing antibodies, cellular immune responses to SARS-CoV-2 are another important part of protective immunity [9,10]. Unlike neutralizing antibodies, cellular responses are not limited to the spike protein and can target epitopes located in all virally encoded structural and non-structural proteins. For this reason, T-cell immunity against epitopes that are conserved across SARS-CoV-2 lineages and variants could theoretically provide significant cross-protection, even against future variants. Although some of the currently licensed S-targeting vaccines have been shown to induce T-cell responses that last longer than antibody responses [11,12], vaccine platforms with a focus on inducing T-cell responses could be expected to perform better in this regard [13]. Indeed, multiple candidate SARS-CoV-2 T-cell vaccines have been studied and are currently in various stages of development [14,15].

Specific binding between T-cell receptors (TCRs) on the surface of T cells and short peptides (usually between 8 and 14 amino acid residues) bound to class I or II major histocompatibility complexes (MHC-I/II) on the surface of antigen-presenting cells is the key step in the T-cell response [16]. These peptides represent T-cell epitopes and are produced by proteolytic antigen processing of intracellularly expressed proteins in the ER (for presentation by MHC-I) or of phagocytosed extracellular proteins in endosomes/lysosomes (for presentation by MHC-II) [17]. Epitope presentation by MHC-I a