Listeriolysin O immunogenetic adjuvant enhanced potency of hepatitis C virus NS3 DNA vaccine

Abstract

Hepatitis C virus (HCV) is a major health problem all over the world. Among HCV proteins, nonstructural protein 3 (NS3) is one of the most promising target for anti-HCV therapy and a candidate for vaccine design. DNA vaccine is an efficient approach to stimulate antigen-specific immunity but the main problem with that is less immunogenic efficiency in comparison with traditional vaccines. Several approaches have been applied to enhance the immunogenicity of DNA. Recently, bacteria-derived substances are considered as one of the most attractive adjuvants for vaccines, which among them, Listeriolysin O (LLO) of Listeria monocytogenes is a toxin with an extremely immunogenic feature. We investigated detoxified form of LLO gene as genetic adjuvant to modulate NS3 DNA vaccine potency. Immunogenic truncated NS3 gene sequence of HCV (1095-1380aa) and detoxified LLO gene region (5-441aa) were amplified by PCR and cloned into the pcDNA3.1 plasmid separately. The expression of recombinant proteins (pc-NS3, pLLO) was confirmed in HEK293T cell line by western blotting. BALB/c mice models received three doses of different formula of plasmids in two-week intervals and two weeks after the final immunization, the immune responses were evaluated by specific total antibody level, lymphocyte proliferation, cytotoxicity, and cytokine levels assays. To evaluate in vivo cytotoxic activity, tumor challenge was performed. The recombinant plasmids were successfully expressed in mammalian cell line, and coadministration of pc-NS3 with pLLO induced the highest titer of total IgG against NS3 antigen compared with other controls. Determination of IgG subclasses confirmed the efficient increase in mixed responses with Th1 dominancy. Furthermore, significant levels of cytokines (p