T-cell-inducing mRNA COVID-19 vaccine improves effectiveness

By Pooja Toshniwal PahariaMay 25 2023Reviewed by Benedette Cuffari, M.Sc.

In a recent study published in Nature Communications, researchers develop a lipid nanoparticle (LNP)-formulated messenger ribonucleic acid (mRNA)-based T-lymphocyte-inducing antigen encoding three severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) peptides derived from SARS-CoV-2 non-structural proteins (NSP), thereby enriching class I human leukocyte antigen (HLA-I) epitopes (HLA-EPs).

Study: An mRNA-based T-cell-inducing antigen strengthens COVID-19 vaccine against SARS-CoV-2 variants. Image Credit: medienspot / Shutterstock.com

Background

Herd immunization attained through mass-scale vaccinations can effectively prevent the spread of infectious diseases. However, most of the coronavirus disease 2019 (COVID-19) vaccines that are currently authorized for use comprise SARS-CoV-2 Wuhan-Hu-1 strain spike (S) protein or S receptor-binding domain (RBD).

Many novel SARS-CoV-2 variants possess S protein mutations that facilitate evasion of humoral antibody-mediated immunity, thereby rendering these vaccines less effective. Furthermore, humoral immunity wanes with time; therefore, antigens that induce cell-mediated immunity, regulated by T lymphocytes, may be incorporated into COVID-19 vaccines to improve their efficacy and reduce the global burden of COVID-19.

About the study

In the present study, researchers investigate whether the induction of broad and potent cell-mediated immunity by the LNP-formulated mRNA-based vaccine could be a practical approach to strengthen COVID-19 vaccine effectiveness.

Serum samples were obtained from COVID-19 convalescents, from which peripheral blood mononuclear cells (PMBCs) were isolated and memory-type cluster of differentiation 8+ (CD8+) T-lymphocytes were expanded.

BALB/c mice, HLA-A*11:01/DR1 transgenic mice, HLA-A*02:01/DR1 transgenic mice, and rhesus macaques were used for SARS-CoV-2 challenge and vaccination experiments. Functional SARS-CoV-2 HLA-I epitopes were analyzed to identify epitope-enriched fragments with 50% inhibitory concentrations (IC50) below 10 nanomolar (nM).

The epitopes were predicted to correspond to the 78 most frequent HLA-I alleles. Further, human embryonic kidney 293T (HEK293T) cells were transfected with plasmids encoding the predicted fragments of SARS-CoV-2 open reading frames (ORFs). These cells were co-cultured with HLA genotype-matched memory T-lymphocytes from COVID-19 convalescents and sorted using flow cytometry.

The mRNAs were synthesized in vitro using T7 polymerase-mediated deoxyribonucleic acid (DNA)-dependent ribonucleic acid (RNA) transcription and naked mRNA expression in HEK193T cells was validated. Subsequently, LNP formulations were prepared.

To detect mRNA-LNP distribution in vivo, BALB/c mice were intramuscularly inoculated with luciferase mRNA-LNP. Further, HLA-A*02:01/DR1 and HLA-A*11:01/DR1 transgenic mice were randomly allotted to receive LNP-HLA-EPs, LNP comprising the SARS-CoV-2 Beta variant of concern (VOC), RBD (LNP-RBD_beta), or both. Rhesus macaques received similar injections.

Enzyme-linked immunosorbent assays (ELISAs) were performed to determine serological anti-RBD immunoglobulin G (IgG) titers. In addition, SARS-CoV-2 pseudoviruses were generated and neutralization assays were performed to assess SARS-CoV-2 VOC neutralization.

ELISpot assays of interferon-gamma (IFN-γ) and interleukin-4 (IL-4) were performed. Cells in the spleen and lymph nodes were profiled, and C-X-C chemokine receptor 5 (CXCR5) and C-X-C motif chemokine ligand 13 (CXCL13) levels in the lymph nodes were assessed.

HLA tetramer assays were performed. Tissues obtained following vaccination and SARS-CoV-2 challenge were subjected to histopathological examination. Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was performed for viral load determination.

Results

The recombinant mRNA transcript was coded for HLA-EPs representing the NSP-31443–1605, NSP-4232–444, and NSP-61-201 sites. The reporter cells presenting the epitopes activated cognate CD8+ T-lymphocytes, which secreted granzyme B (GzB) and induced apoptosis in the reporter cells.

LNP-HLA-EPs-vaccinated HLA-transgenic mice exhibited significant increases in the frequencies of CD8+ T-lymphocytes and helper T-cell type 1 (Th1)-based responses.

Vaccination significantly expanded the population of CD8+ CD44+ CD62L+ central memory T-cells (T_cm) and effector memory T-lymphocytes (T_em) in both HLA-transgenic murine models, thus indicating the activation of cell-based immunity by HLA-Eps. In addition, the fraction of epitope-specific IFN-γ- and tumour necrosis factor-alpha (TNF-α)-producing splenocytes enlarged following HLA-EP stimulation.

HLA-EPs induced potent cell-mediated responses against SARS-CoV-2 in HLA-transgenic mice. Notably, HLA-EP sequences were highly preserved in the SARS-CoV-2 variants.

Among the transgenic murine animals and macaques, dual vaccination with the HLA-EP-encoding LNP-based mRNAs and RBD_beta was more effective in preventing SARS-CoV-2 infection by Alpha, Beta, Gamma, and Delta VOCs, as well as the BA.1 sub-VOC of Omicron than LNP-RBD_beta vaccination only.

This was confirmed by the significantly increased CXCR5 and CXCL13 expression in the lymph nodes and minimal histopathological damage in dually immunized animals. HLA-EP immunogenicity was not affected by SARS-CoV-2 VOC mutations.

Conclusions

Based on the study findings, stimulating cell-mediated and humoral immune responses could improve COVID-19 vaccine effectiveness against the SARS-CoV-2 ancestral strain and novel VOCs. Next-generation vaccines must comprise cytotoxic lymphocyte-inducing S protein and non-S protein antigenic fragments that are highly conserved among SARS-CoV-2 VOCs for broad and potent action.