Identification of the cellular and humoral immune responses elicited against COVID-19 through the transfer of the SARS-COV-2 spike gene via a human adenoviral vector

Aim: The urgency of developing a safe and effective vaccine against COVID-19, caused by the SARS-CoV-2 virus, has led to the development of various vaccine platforms. Among these, viral vector-based vaccines, particularly those utilizing adenoviruses, offer significant advantages. Adenovirus-based vectors are notable for their broad tissue tropism, well-characterized vector genomes, high transgene capacity, natural adjuvant effect, and their ability to elicit robust transgene-specific T cell and B cell responses. This study aims to develop vaccine vectors encoding the SARS-CoV-2 Spike protein (Ad5Spike) using human adenoviral vectors. Method: A human adenoviral expression plasmid encoding the SARS-CoV-2 Spike protein (pAd5Spike) was constructed using Gateway® Cloning Technology. Ad5Spike was produced by transient transfection of pAd5Spike into 293A cells via the CaCO4-mediated method, then purified and concentrated by CsCl density gradient ultracentrifugation. For immunization, the vector was intraperitoneally injected at different concentrations (108, 1010, and 1012 viral particules) into 6-8-week-old female BALB/c mice in 10 mM Tris-Cl. Both cellular and humoral immune responses were evaluated at 30- and 90-days post-immunization using ELISA, ELISpot, and neutralization assays. Results: Elevated levels of anti-spike IgG were observed in the serum of vaccinated mice. Increased secretion of TNF-α, IFN-γ, and IL-2 cytokines from T cells of vaccinated mice was observed. Neutralization tests conducted using a third-generation lentiviral-based pseudovirus demonstrated the presence of neutralizing antibodies against SARS-CoV-2, particularly in the high-dose group. Conclusion: This study illustrates the generation of the Ad5Spike vector with the potential to activate both humoral and cellular immune systems. Thus, we can conclude that this vaccine may provide protection against COVID-19.