OBJECTIVE:To investigate the different killing effects on human hilar cholangiocarinoma cells FRH with cytosine deaminase(CD) and herpes simplex virus thymidine kinase(HSV tk)double suicide genes coexpression compared with single gene mediated by retrovirus.To find a more efficient and low toxicity suicide gene therapy for hilar cholangiocarinoma.
METHODS:CD and HSV tk double suicide genes were transfected into PA317 cells using lipofectamine.The positive clones were picked out and cultured after G418 selected.The viral supernatant was collected.The FRH cells were infected with the virus containing the double suicide genes.After G418 selection,RT PCR was resorted to demonstrated the successful transcription of CD and HSV tk genes.The FRH/CD+tk and FRH cells in culture were respectively treated with 5 Fc and /or GCV.The cytoxicity efficacy was evaluated by microculture tetrajolium test (MTT) method.
RESULTS:The virus containing double suicide gene was produced in PA317 cells.Double suicide genes were stably expressed in RFH cells after being infected with the virus.The killing effect of combination 5 Fc with GCV on FRH/CD+tk cells is more effective than that of using 5 Fc or GCV alone.
CONCLUSION:The CD+TK/5 Fc+GCV co expression system is more effective for killing effects on FRH cells than that of CD/5 Fc or tk/GCV system alone.