Production and characterization of a chimeric antigen, based on nucleocapsid of SARS-CoV-2 fused to the extracellular domain of human CD154 in HEK-293 cells as a vaccine candidate against COVID-19.

Despite that more than one hundred vaccines against SARS-CoV-2 have been developed and that some of them were evaluated in clinical trials, the latest results revealed that these vaccines still face great challenges. Among the components of the virus, the N-protein constitutes an attractive target for a subunit vaccine because it is the most abundant, highly conserved and immunogenic protein. In the present work, a chimeric protein (N-CD protein) was constructed by the fusion of the N-protein to the extracellular domain of human CD154 as the molecular adjuvant. HEK-293 cells were transduced with lentiviral vector bearing the N-CD gene and polyclonal cell populations were obtained. The N-CD protein was purified from cell culture supernatant and further characterized by several techniques. Immunogenicity studies in mice and non-human primates showed the N-CD protein induced high IgG titers in both models after two doses. Moreover, overall health monitoring of non-human primates demonstrated that animals were healthy during 228 days after first immunization. Data obtained support further investigation in order to develop this chimeric protein as vaccine candidate against COVID-19 and other coronavirus diseases.

Chimeric Antigen by the Fusion of SARS-CoV-2 Receptor Binding Domain with the Extracellular Domain of Human CD154: A Promising Improved Vaccine Candidate.

COVID-19 is a respiratory viral disease caused by a new coronavirus called SARS-CoV-2. This disease has spread rapidly worldwide with a high rate of morbidity and mortality. The receptor-binding domain (RBD) of protein spike (S) mediates the attachment of the virus to the host's cellular receptor. The RBD domain constitutes a very attractive target for subunit vaccine development due to its ability to induce a neutralizing antibody response against the virus. With the aim of boosting the immunogenicity of RBD, it was fused to the extracellular domain of CD154, an immune system modulator molecule. To obtain the chimeric protein, stable transduction of HEK-293 was carried out with recombinant lentivirus and polyclonal populations and cell clones were obtained. RBD-CD was purified from culture supernatant and further characterized by several techniques. RBD-CD immunogenicity evaluated in mice and non-human primates (NHP) indicated that recombinant protein was able to induce a specific and high IgG response after two doses. NHP sera also neutralize SARS-CoV-2 infection of Vero E6 cells. RBD-CD could improve the current vaccines against COVID-19, based in the enhancement of the host humoral and cellular response. Further experiments are necessary to confirm the utility of RBD-CD as a prophylactic vaccine and/or booster purpose.