Abstract
Host immune response plays critical roles in virus-mediated gene therapy and vaccination as it may affect the transgene gene expression. Baculovirus has emerged as a promising vector for gene therapy and vaccination due to a number of distinct advantages. However, it remains unclear whether baculovirus administration elicits adaptive immune responses that in turn impairs the therapeutic effect. To address this question, a recombinant baculovirus expressing firefly luciferase was constructed and injected intramuscularly into the BALB/c mice at days 0 and 14. We found that the transgene expression was hindered after repeated injection, and the first baculovirus injection triggered the anti-baculovirus IgG, which neutralized the ability of baculovirus to transduce mammalian cells. The antibody could be antagonized by the baculoviral envelope protein gp64, suggesting that the neutralizing antibody blocked the virus transduction via gp64. Very importantly, baculovirus administration provoked the IFN- and IL-4 secretion as well as activated both CD4+ and CD8+ T cells. The stimulation of CD8+ cell responses was at least partly attributed to gp64 and the CD8+ cytotoxic T lymphocytes (CTL) epitope gp64457-465 was identified. In summary, baculovirus injection in vivo induces both neutralizing antibodies and gp64-specific CTL, and the humoral response provoked by the first injection hindered the transgene expression mediated by subsequent injection. Due to the transgene expression was hindered by host adoptive immunity. At the second part of the study, we aimed to develop a hybrid baculovirus for sustained transgene expression to avoid repeating injection. We first constructed plasmids featuring either EBNA-1/oriP, adeno-associated virus inverted terminal repeats and Rep recombinase (AAV ITRep) or Sleeping Beauty (SB) transposon and compared their efficacies in terms of persistent expression. In HEK293 cells, AAV ITR failed to prolong the expression while EBNA-1/oriP moderately extended the expression to 42 days. In contrast, the SB system led to stable expression beyond 77 days even without antibiotic selection. Given this finding, we constructed a hybrid SB-baculovirus expressing the SB transposase and harboring the transgene cassette flanked by IR/DR elements recognizable by SB. The hybrid SB-baculovirus efficiently transduced mammalian cells and mediated an expression duration longer than that by conventional baculoviruses, thanks to the transgene persistence and integration. Moreover, transgene expression mediated by i.m. injection of hybrid SB-baculovirus in vivo was extended for 1 year. The first part of the study underscores the significance of adaptive immune responses elicited by baculovirus and will benefit future rational design of virus administration schemes for gene therapy and vaccination. The second part of the study demonstrated the potential of the hybrid SB-baculovirus vector for prolonged transgene expression in vitro and in vivo. In summary, these data together will allow baculovirus vector to have better application in in vivo gene therapy.