In a recent study published in Virology, researchers evaluated the immunogenicity of DNA-based candidate vaccines against coronavirus 2 (SARS-CoV-2) of severe acute respiratory syndrome in mice.
Study: Immunogenicity of SARS-CoV-2 candidate DNA vaccines based on ear protein. Image credit: CROCOTHERY / Shutterstock
Fund
Infection with SARS-CoV-2, the causative agent of the 2019 coronavirus disease pandemic (COVID-19), causes respiratory symptoms, usually accompanied by fatigue, fever, cough, dyspnea, acute pneumonia, and anorexia. The virus uses its spike protein (S) for host cell entry by binding to the angiotensin 2 converting enzyme (ACE2) in cells.
Protein S includes many neutralizing epitopes that could be exploited for vaccine design / development. Several COVID-19 vaccines have been authorized for human immunization, and many more are still in different developmental / clinical stages.
About the study
In the present study, the researchers evaluated the potential of the SARS-CoV-2 DNA vaccine and determined the appropriate antigen and dosing regimen. The first human isolate of SARS-CoV-2 in South Korea was passed and assessed as plaque-forming units (PFUs) in Vero cells. The sequence of the S gene was optimized using the Optimum gene algorithm to improve its expression.
Four vaccine candidates were constructed that expressed 1) full-length S protein, truncated S protein without 2) cytoplasmic domains (SΔCD) or 3) transmembrane (SΔTM), and 4) the S1 subunit alone. The tissue plasminogen activator (tPA) leader sequence bound to the N-terminus to enhance in vivo expression. The optimized sequence was digested with endonucleases and cloned into pVax1, a mammalian expression vector.
C57BL / 6 mice were inoculated three times with candidate DNA vaccines at a three-week interval. The vaccine was administered to the anterior tibialis muscle of mice. Shortly afterwards, two-needle matrix electrodes were inserted into the muscle for in vivo electroporation. Blood samples were collected four to seven weeks after the first vaccination.
They performed a virus neutralization assay and defined the neutralizing antibody titer (nAb) as the dilution factor corresponding to a 50% reduction in plaque formation relative to the controls. Briefly, 60 PFUs of SARS-CoV-2 were mixed with an equal volume of a twice-diluted sample of mouse serum, which was used to infect Vero cells. Cytokine expression analysis was performed using an enzyme-linked immunosorbent mouse interferon (IFN) -γ assay (ELISpot).
Discoveries
The authors noted that total S1 and S2-specific immunoglobulin G (IgG) antibodies were high two weeks after vaccination in mice receiving full-length S, SΔCD, and SΔTM vaccines. In addition, IgG levels increased with additional doses. The full-length S-DNA vaccine resulted in higher nAb titers than other candidate vaccines. Similarly, cytokine levels were higher in full-length S-DNA vaccine receptors.
This meant that removal of the cytoplasmic or transmembrane domain of the S protein or S2 subunit would result in poor humoral responses. Because the full-length S vaccine was the most effective antigen, the authors performed additional experiments to identify the appropriate dose of this vaccine. Mice were stratified into three groups to receive 5, 20, or 50 μg of the full-length S vaccine. Animals that received the 50 μg dose of the vaccine showed the highest levels of antibodies.
In addition, virus neutralization assays revealed higher nAb titers in animals of the 50 μg regimen than in others. NAb titers were significantly higher after the third dose than after the second vaccination. Although all mice showed a significant increase in cytokine levels regardless of dose relative to controls, vaccines with higher doses (50 or 20 μg) had higher IFN-γ levels than those administered with 5 μg of the same antigen.
Conclusions
The authors designed four candidates for DNA-based vaccines using the S protein from the first Korean isolate of SARS-CoV-2. The gene sequence was optimized and a leading tPA sequence at the N (S) end was linked for translation efficiency. The study revealed that total IgG antibodies were higher in mice vaccinated with full-length S and truncated S without transmembrane or cytoplasmic domains than in those immunized with S1.
The full-length S vaccine induced significantly higher nAb titers than other candidate vaccines, except the SΔCD vaccine. They observed that any dose (5, 20, or 50 μg) of full-length S obtained adequate IgG and nAb titers. However, 5 or 20 μg of the vaccine resulted in similar titers, and the highest tested dose of the vaccine induced maximum antibody titers.
In conclusion, the study showed that the SARS-CoV-2 DNA vaccine induced cellular and humoral immune responses. Further research is required to investigate the protection afforded by this vaccine against SARS-CoV-2 infection.