[Enhancement of HBV gene-induced specific cell-mediated immunoresponse by C3d-P28]

To investigate whether P28 derived from complement C3d can enhance the cell-mediated immunoresponse to HBV-preS2/S induced by direct injection of naked plasmid DNA containing four tandem repeats of C3d-P28 gene and HBV-preS2/S gene existed in fusion form.

Four copies of gene coding for C3d-P28, amplified by PCR and cut by restriction endonucleases digestion, were subcloned into a eukaryotic expression vector pVAON33 to construct pVAON33-P28.4. HBV-preS2/S gene was then introduced into the pVAON33 and pVAON33-P28.4 respectively to form pVAON33-S2/S and pVAON33-S2/S-P28.4. The recombinant plasmids were identified by PCR and restriction endonucleases digestion as well as DNA sequencing. BALB/c mice were immunized i.m. three times at 3 weeks' intervals with 100 microg of pVAON33-S2/S DNA, pVAON33-S2/S-P28.4 and mock DNA, respectively. Splenocytes from immunized mice were stimulated by HBsAg and then harvested to analyze the specific lymphocytic proliferative response and CTL cytotoxic activity by (3)H-TdR incorporation assay and isotopic release analysis, respectively.

Specific lymphocytic proliferation and CTL cytotoxic activity against HBV-preS2/S were observed in mice immunized by both pVAON33-S2/S and pVAON33-S2/S-P28.4 in dose-dependent form. Specific lymphocytic proliferation and CTL response in mice immunized by pVAON33-S2/S-P28.4 were markedly stronger than those in mice immunized by pVAON33-S2/S.

A C3d-P28 can enhance the cell-mediated immunoresponse induced by HBV-preS2/S gene immunization.