The immunogenicity in mice of hepatitis C virus core protein delivered as DNA is modulated by its capacity to promote ROS production and activate Nrf2/ARE pathway

Hepatitis C virus (HCV) core protein is an attractive target for HCV vaccine, however little progress was achieved in trials. We aimed to delineate the factors restricting immunogenicity of HCV core, and to improve its performance as DNA immunogen in a mouse model. We investigated the plasmids encoding the full-length protein (1-191 aa), its C-terminally truncated mutants 1-173, 1-152, 1-98, 1-50, 1-36, N-terminally truncated 37-191 form, or harboring the codon-optimized gene (1-152s). They were assessed for the level of protein expression, route of degradation, ability to enhance ROS production and activate expression of heme oxygenase 1 (HO-1) and NAD(P)H:quinone oxidoreductase (Nqo1), regulated through Nrf2/ARE pathway. Cores truncated at the N-terminus and C-terminus beyond aa 152 accumulated at 10-100 times lower level than the full-length protein. Core(1-36) induced ROS production by Nox1 and Nox4, whereas Core(37-191) - by mitochondria, CYP2E1, and Ero1a, and both of them up-regulated the expression of HO-1 and Nqo1 at transcriptional level. The N-terminally truncated core protein upregulated production of ROS. HCV core genes were used to immunize BALB/c mice by intradermal injection with electroporation or C57Bl/6 mice by repeated intramuscular injections. The immunogenicity of HCV core gene in terms of IFN-γ response correlated with levels of ROS production and HO-1/Nqo1 expression. Anti-core immune response inversely correlated with Nqo1/HO-1 expression if recalculated per unit of HCV core protein in a cell. The strongest HCV-specific cellular response was induced by Core(1-152s) gene which supported a modest level of protein expression, moderate ROS production and stress response. Thus, cellular immunogenicity of HCV core is shaped by its ability to promote ROS production and to trigger the stress response.