SRA

New universal prophylactic vaccine platforms are needed to prevent or reduce the impact of future respiratory viral infections. While the attribute of T memory cells is particularly attractive in the context of vaccine design for viral infection, the efficiency of T and B memory cell-mediated protection generated under the adenoviral vector was tested. We studied the mechanism using the platform/ immunological factor combination (AdV1+rRBD): adenovirus AdV1 (adjuvant) armed with inducible co-stimulator ICOS and CD40 ligand in combination with recombinant spike protein rRBD (antigen) to promote the differentiation of memory T and B cells. Additionally, adenovirus AdV2 (adjuvant) without ICOS and CD40 ligand in combination with rRBD was tested. Investigating ex vivo T cell responses to the platform-specific stimuli, we found higher frequencies of CD8+ than CD4+ platform-specific T cells vs mock. The cytometric analysis showed that T cells confer protection against SARS-CoV-2 via elevated T cell subsets: CD8+TEMRA (57.6% ± SD 3.3 vs mock 39.6% ± SD 4.83) and CD4+TNAÏVE (59.6% ± SD 0.4 vs mock 50.8% ± SD 1.9). We found that CD8+TEM, CD8+TSCM, and CD8+TCM are antigen-experienced subpopulations generated via the platform after 7 days. When stimulated with the immunological factors, B cells changed from the production of IgM and IgD to isotype IgG. The CD40 gene was significantly expressed (RT-qPCR) in VERO E6 after stimulation with the platform. RNA-seq profiling identified Th1, Th2, and Th17 cell differentiation gene expression patterns that effectively protect against different pathogens. RNA-seq analysis determined signaling pathways that control immunological mechanisms by which the platform induces protective immunity: MAPK cascade, adipocytokine, cAMP, TNF, and Toll-like receptor (TLR) signaling pathway. The formulation (AdV1+rRBD) increased the production of IL-6, IL-8, and TNF vs the mock (P<0.0001). The whole-genome sequencing of VERO E6 showed differences in the expression of apoptosis gene stimulated with the platforms vs mock. Moreover, apoptotic hallmarks were identified and quantified using flow cytometry. We proved that immunogenic factors compromised plasma membrane integrity (RNA-seq). To sum up, the proposed adenoviral platform generated innate and adaptive immunity for the complex and robust regulatory system to prevent SARS-CoV-2 infections. Overall design: Comparative gene expression profiling analysis of RNA-seq data for VERO E6 ATCC cells and its infected derivatives by AdV1 IC100 (AdV-D24-ICOSL-CD40L-100 VP/mL), AdV2 IC100 (AdV-D24-WT 100 VP/mL), Pseudo-SARS-CoV-2 IC100 (Pseudovirus SARS-CoV-2 100 VP/mL), AdV1+pseudo-SARS-CoV-2 IC100 (AdV-D24-ICOSL-CD40L + Pseudovirus SARS-CoV-2 100 VP/mL), AdV2+pseudo-SARS-CoV-2 IC100 (AdV-D24-WT + Pseudovirus SARS-CoV-2 100 VP/mL).