Transcriptomic analysis of the human immune response to Vesicular Stomatitis Virus-vectored vaccines

Transcriptomic analysis is becoming increasingly important in the study of gene expression and is widely applied to the study of immune response after vaccination. This approach allows the integration of classic immunological evaluations with gene expression correlating clinical and immunological data with specific gene signatures or biomarkers. Gene signatures can be useful to understand the activation of immune response, to monitor the response to vaccine and, in principle, to predict the onset of adverse event. In this thesis, transcriptomic analysis was applied in two studies on vaccines based on the recombinant Vesicular Stomatitis Virus (VSV) vector. The first one was rVSVΔG-ZEBOV-GP, a live VSV expressing Ebolavirus glycoprotein G, and the only Ebola vaccine with demonstrated clinical efficacy. RNA extracted from whole blood of vaccinated volunteers, 51 injected either with 107 or 5x107 pfu of rVSV-ZEBOV and 13 with placebo, collected at several times after vaccination, was used to perform targeted sequencing. A differential expression analysis (DEA) was performed at each time point using the edgeR software. Differentially expressed genes (DEGs), ranked by FDR, were analysed to find their biological function. The highest number of DEGs (5,469) was observed at day 1 after vaccination, this number decreased over time: at day 21 no gene was differentially expressed. Functional analysis showed the activation of 145 different modules. Pathways related to interferon and to innate immunity were markedly activated from day 1 to day 14. At days 2 and 3, neutrophils modules were downregulated while complement was activated, at days 7 and 14 T-cell and cell-cycle associated modules were upregulated. ZEBOV glycoprotein-specific antibody titres were used to perform a correlation analysis with module activation: six modules at day 14 resulted directly correlated, including one related to B cells. The second study involved a replication attenuated rVSV expressing gag from HIV. The clinical trial involved 60 healthy volunteers immunized with 5 different doses of vaccine. Total RNA was extracted and sequenced from PBMC before vaccination and after 7 days. Differential expression and functional analysis showed the activation of modules related to cellular proliferation at day 7, in particular to the activation and proliferation of CD4+ T cells. The highest vaccine dose also induced the activation of modules related to innate immunity, such as the dendritic cells via NF-kB pathway. Analysis of the expression level of IFI27 gene in both the studies was conducted to compare the ability to induce interferon/antiviral pathways. The expression of IFI27 was dose-dependent for rVSV-gag, but it was lower than that induced by a similar dose of rVSV-ZEBOV. Data of placebo samples from the two studies were used to identify differences in the expression profile between whole blood and PBMC. DEGs were related to globins, to cytokines and chemokine receptors genes, the former were more expressed in whole blood, while the latter in PBMC. Functional analysis showed 25 enriched modules, the most significant were related to heme biosynthesis, erythrocyte differentiation, but also to innate immunity.