Immunogenic Properties of the DNA Construct Encoding the Receptor-Binding Domain of the SARS-CoV-2 Spike Protein

Abstract

— The development of preventive vaccines became the first order task in the COVID-19 pandemic caused by SARS-CoV-2. This paper reports the construction of the pVAX-RBD plasmid containing the Receptor-Binding Domain (RBD) of the S protein and a unique signal sequence 176 which promotes target protein secretion into the extracellular space thereby increasing the efficiency of humoral immune response activation. A polyglucine-spermidine conjugate (PGS) was used to deliver pVAX-RBD into the cells. The comparative immunogenicity study of the naked pVAX-RBD and pVAX-RBD enclosed in the PGS envelope showed that the latter was more efficient in inducing an immune response in the immunized mice. In particular, RBD-specific antibody titers were shown in ELISA to be no higher than 1 : 1000 in the animals from the pVAX-RBD group and 1 : 42 000, in the pVAX-RBD-PGS group. The pVAX-RBD‒PGS construct effectively induced cellular immune response. Using ELISpot, it has been demonstrated that splenocytes obtained from the immunized animals effectively produced INF-γ in response to stimulation with the S protein-derived peptide pool. The results suggest that the polyglucine-spermidine conjugate-enveloped pVAX-RBD construct may be considered as a promising DNA vaccine against COVID-19.

Self-Assembled Particles Combining SARS-CoV-2 RBD Protein and RBD DNA Vaccine Induce Synergistic Enhancement of the Humoral Response in Mice

“…To create virus-like particles, we encapsulated DNA vaccines in a PGS and RBD protein conjugate (Figure 2B). We have previously used the PGS conjugate to deliver candidate DNA vaccines against HIV-1 and Ebola [33,42,43]. We used the PGS-TBI protein conjugate to create a vaccine against HIV-1, namely CombiHIVvac, which has successfully passed the first phase of clinical trials [44].…”

“…Plasmid isolation and purification were carried out as described earlier [43]. Transfection of HEK293T cells with plasmid pVAXrbd was performed by using Lipofectamine 3000 (Invitrogen, Waltham, MA, USA) according to the manufacturer's instructions.…”

“…The neutralizing properties of antibodies of blood serum were determined by inhibition of the cytopathic effect (CPE) of the virus in cell culture in vitro, as described previously [43]. Briefly, the SARS-CoV-2 nCoV/Victoria/1/2020 strain (100 TCID 50 ), incubated with serial dilutions of the studied sera, was plated on a 100% confluent monolayer of Vero E6 cells.…”

Immunogenicity of the DNA/Protein Combined Vaccine against COVID-19

“…This sequence was optimized for expression in mammalian cells, contained a tissue plasminogen activator signal peptide, Tpa (MDAMKRGLCCVLLLCGAVFVSA), was synthesized as part of the pVAX plasmid vector in DNK-Sintez Company, and was named pVAXs. Plasmid production and isolation were performed as described in [6]. The expression of protein S gene was studied using HEK293 cells transfected with pVAXs plasmid.…”

Cellular immune response to DNA vaccine encoding receptor-binding domain of SARS-CoV-2 S protein: dependence on the packing mode

“…Конструирование плазмиды pVAXrbd было описано ранее [2]. Последовательность гена, кодирующего белок RBD, была клонирована в составе эукариотического плазмидного вектора pVAX.…”