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Development of Methods about Safe Application of Viral Strains with DNA-Genome as a Material for Design of New Molecular Vaccines against SARS-CoV-2/COVID-19

Iskra Sainova1*, Vera Kolyovska 1, Tzvetanka Markova2, Dimitrina Dimitrova-Dikanarova3, Desislava Drenska4, Dimitar Maslarov4

Taking in consideration the risk of thrombs by SARS-CoV-2 Spike (S) protein and of amyloid brain plaques by its Nucleocapsid (N) protein, design of molecular (DNA, RNA and/or protein) vaccines against viral Envelope (E) protein or viral Membrane (M) protein, and boosting by siRNAs against virus genes, coding proteins S and N. Laboratory-incubated mammalian cells were transfected by recombinant gene construct. This vector and separate sub-populations of transfected cells, contained additional gene copy, demonstrated safety and capability for generation of adequate immune response in vitro and in vivo. Other mammalian in vitro-cellular cultures were inoculated with low infectious titers (high viral suspensions dilutions) of vaccine avipoxviral strains (Poxviridae family), freezed in the presence of cryo-protector Dimethylsulfoxide (DMSO), thawed and reincubated. As a source of extra-cellular virus served the cultural fluids, and of intra-cellular-scraped cellular monolayers. The titers of the intra-cellular forms were significantly higher that these of the extra-cellular. Transfer of nucleotide (DNA and/or RNA) fragments from virus to cellular genome, and in opposite direction due to activated fusion on the influence of DMSO plus drastic temperature changes were suggested. A possibility about application of viruses with DNA and/or RNA-genome for production of molecular vaccines and of appropriate siRNAs was proposed. In vitro-incubated cultures of non-malignant mouse embryonic cells; of containing endogenous retrovirus mouse malignant myeloma cells, and of mixed cultures of both were lysed and subjected on ELISA for determination the antibodies presence and titers. A possibility for production of membrane receptor glycoproteins and antibodies/immunoglobulin’s by non-myeloid and nonlymphoid cells in appropriate conditions was suggested. Because the produced antibodies are out of the germinative centers in the lymphoid tissues and organs, for escape of chronic inflammation, often leading to malignant transformation or degenerative changes, they should be controlled by small ions or molecules.