THE IMMUNE RESPONSES OF THE PRIME-BOOST REGIMEN WITH rBCG-E12 AND rDIs-E12 CANDIDATE VACCINE

摘要

In the previous study, a research group in NIID, Japan demonstrated that the V3 sequence of 12 amino acids of HIV-1 CRF01_AE (E12 epitope) fused with mycobacterial alpha-antigen was secreted from BCG cells (rBCG-E12) and could induce NT-Ab against CRF01 AE primary isolates. However, the NT-Ab liter in guinea pig was not enough to obtain protective efficacy. So, we attempted to boost the NT-Ab by rDIs expressing E12 epitope-alpha-antigen fusion protein (rDIs-E12). We have started the second generationAIDS vaccine research project and successfully constructed rDIs-E12 which expressed E12 epitope under control of vaccinia p7.5 promoter in infected chicken embryo fibroblast cell. The rBCG-E12 clone that could secrete El2 epitope under control of alpha-antigen promoter was provided us by NIID, Japan group. The expression of alpha-antigen-E12 fusion protein in the both rDIs-E12 and rBCG-E12 were checked by Western blot analysis using HIV~+ human serum and the size of the fusion protein was approximately32 kDa. To test immunogenicity, we primed Balb/c mice with rBCG-E12 and boosted with rDIs-E12 to analyze effect of prime-boost regimen for NT-Ab production. The ELISPOT was performed for the quantitation of antigen-specific CD8 T cells responses. We found that the E12 itself could not stimulate CTL response, it might not be CTL epitope in Balb/c mice. The rBCG and rDIs vectors could stimulate CTL activity, this means CTL activity was induced with alpha antigen and PPD. The CTL responses increase aftersecond boosting with rDIs. The ELISA was also performed for check the antibody titer against E12 and alpha antigen. There are antibody liter against alpha antigen but no antibody tiler against E12 peptide. From the experimenl, we conclude lhal priming wilh rBCG-E12 and boosting with rDIs-E12 could stimulate CTL responses and have boosting effect but could nol stimulale antibody tiler. Further study will develop the regimen of immunization and using new epilope for candidate vaccine.