High expression of transgenes mediated by hybrid retroviral vectors in hepatocytes: comparison of promoters from murine retroviruses in vitro and in vivo.

To achieve high transgene expression in the liver, we have compared the reporter gene expression among various murine retroviral long terminal repeats (LTRs) or leader sequences in vitro. Transient reporter gene expression assays revealed the highest gene expression by the polycythemic strain of spleen focus-forming virus (SFFVp) LTR in differentiated hepatocellular carcinoma cell lines, HuH-7 and PLC/PRF/5. However, remarkable difference was not observed among LTRs in other types of human liver tumor cell lines. Essentially the same results were obtained by infecting these cells with a series of retroviral vectors. Repression of transgene expression was observed by the leader sequences from Moloney murine leukemia virus (MoMLV), but not from mouse embryonic stem cell virus (MESV). Strengths of the promoters were further compared in murine hepatocytes in vivo. Although the proportions of genomic integration were almost the same, higher gene expression was observed by the FMEV-type vector, which contained the SFFVp LTR and the MESV leader, in comparison with that by the MoMLV-based vector. Thus, FMEV-type vectors may represent a novel type of vectors for human gene therapy with hepatocytes.