부산대학교 도서관

Retroviral sequence can silence transgene expression in vitro and in vivo. We report that this effect can be efficiently prevented by in vivo administration of the demethylating agent 5-azacytidine (aza-C). We engineered the U937 human cell line with a retroviral vector consisting of the thymidine kinase suicide gene (tk), which induces sensitivity to ganciclovir (gcv) and through an IRES sequence, the bacterial beta-galactosidase gene (lacZ) as a marker gene. About 90% of the U937 cells expressed the transgene. By injecting the transduced U937 cells in severe combined immunodeficient disease (SCID) mice, we generated a tumour which, during in vivo treatment with aza- C, maintained the high expression of lacZ and tk genes at the baseline values. LacZ-positive cells in the tumour masses after death was weak (1–2%) in the control group, while in mice treated with aza-C it was maintained at 90%. The delay in tumour onset was significanly longer when animals were treated with both aza-C and gcv (P < 0.0001) compared with animals treated with gcv or with aza-c alone. the prevention of silencing phenomena has important implications for gene therapy, because an efficient transduction associated with appropriate drug therapy, might be a powerful strategy for successful application of gene therapy protocols. [ABSTRACT FROM AUTHOR]