Enhancement of transgene expression by cotransfection of oriP plasmid withEBNA-1 expression vector

We have attempted to develop a system for specific enhancement of transgene expression, which has been one of the most important issues in human gene therapy,When an Epstein-Barr virus (EBV) nuclear antigen 1 (EBNA-1) express ion vector, pCMV-trEBNA-1, was cotransfected with an origin of latent viral DNA replication (oriP)-harboring plasmid, poriP-CMV-luciferase, luciferase gene expression was up to 20 times greater than in the absence of EBNA-1, This enhancement was regulated mainly at the transcriptional level and was dependent on the oriP sequence and the amount of EBNA-1. However, cointrodu ction of poriP-CMV-luciferase with purified recombinant EBNA-1 inhibited lu ciferase gene-expression whereas no inhibition was observed when pCMW-lucif erase was cointroduced with recombinant EBNA-1, We also introduced poriP-CM V-luciferase into mouse liver via the use of HVJ (hemagglutinating virus of Japan)-liposomes. By 10 days after transfer, luciferase gene expression wa s decreased to low levels. We then introduced pCMV-trEBNA-1 to mouse liver via HVJ-liposomes on day 10, Luciferase gene expression was reactivated, wh ereas no reactivation was detected by the injection of EBNA-1 expression pl asmid into liver injected with pCMV-luciferase lacking the oriP sequence, T hus, cotransfection of oriP-harboring expression vector with EBNA-1 express ion plasmid should be promising for human gene therapy, although the safety of the system must be investigated thoroughly.