IMMUNIZATION WITH PLASMID DNA ENCODING HEPATITIS-C VIRUS ENVELOPE E2 ANTIGENIC DOMAINS INDUCES ANTIBODIES WHOSE IMMUNE REACTIVITY IS LINKEDTO THE INJECTION MODE

Plasmids expressing different domains of the hepatitis C virus (HCV) e nvelope E2 glycoprotein from a genotype la isolate were constructed to compare the immunogenic potential of E2 in nucleic acid-based immuniz ations. One plasmid, pCIE2t, expressed a C-terminally truncated form o f E2, while others, pS2.SE2A to pS2.SE2E, encoded the adjacent 60-amin o-acid (aa) sequences of E2 (inserts A to E) expressed as a fusion wit h the hepatitis B virus surface antigen. BALB/c mice were given inject ions of the plasmids intramuscularly (i.m.) or intraepidermally (i.e.) via a gene gun (biolistic introduction), and induced humoral immune r esponses were evaluated. The i.e. injections resulted in higher seroco nversion rates and antibody titers, up to 100-fold, than did the i.m. injections (P = 0.01 to 0.04). Three restricted immunogenic domains, E 2A (aa 384 to 443), E2C (aa 504 to 555), and E2E (aa 609 to 674), that yielded antibody titers ranging from 1:59 to >1:43,700 could be ident ified. Subtype 1a-and 1b-derived E2 antigens and synthetic peptides we re used in Western blot and enzyme-linked immunosorbent assay analyses , which revealed that the cross-reactivity of the plasmid-induced anti bodies was linked both to the type of antigen expressed and to the inj ection mode. Induced anti-E2 antibodies could immunoprecipitate noncov alent E1E2 complexes believed to exist on the surface of HCV virions. This study allowed us to identify restricted immunogenic domains withi n E2 and demonstrated that different routes of injection of HCV E2 pla smids can result in quantitatively and qualitatively different humoral immune responses.