Cytotoxicity of DOXª²GA3 to ¦Âª²glucuronidase cDNA transfected human renal carcinoma GRCª²1 cells in vitro

    ZHANG Shiª²Rui, WANG He, QIN Weiª²Jun, LIU Heª²Liang, WEN Weiª²Hong

    1Department of Urology, Xijing Hospital, 2Department of Immunology, School of Basic Medicine, Fourth Military Medical University, Xi¬ðan 710033, China

    ¡¾Absract¡¿ AIM: To establish a renal carcinoma model with high expression of ¦Âª²glucuronidase (¦ÂG) and to observe the killing effect of ¦ÂGª²prodrug Nª²[4ª²Doxorubicinª²Nª²carbonyl(oxymethyl)phehyl]ª²Oª²betaª²glucuronyl carbamate (DOXª²GA3) on ¦ÂG cDNA transfected human renal carcinoma GRCª²1 cells in vitro. METHODS: A eukaryotic expression vector pcDNA3.1(+)ª²¦ÂG was transferred into renal carcinoma GRCª²1 cells with lipofection reagent and the ¦ÂG vectorª²producer cell line GRCª²1/¦ÂG was isolated in 1000 mg/L neomycin medium. It was confirmed by genomic PCR, immunofluorescency and immunohistochemistry. The cytotoxicity and bystander effect were estimated by MTT and trypan blue exclusion test. RESULTS: ¦ÂG gene was stably integrated into the genomic DNA of the GRCª²1 cell and got a high expression. The DOXª²GA3, with the concentration of 100 ¦Ìmol/L for 3 d, had little effect on parental cells, with a survival rate of 82%. However, it killed all the transfected cells. When IC50 GRCª²1/¦ÂG was 1.24 ¦Ìmol/L, IC50 GRCª²1 was more than more than 100 ¦Ìmol/L, and the GRCª²1/¦ÂG cells were mixed with parental cells with various proportions, the GRCª²1/ ¦ÂG cells accounted for 20%, 80% of the mixed cells were killed. CONCLUSION: We have successfully established the renal carcinoma model with high expression of ¦ÂG gene. DOXª²GA3 has obvious cell killing effect on the GRCª²1/¦ÂG, which is strengthened with the rising concentration and the prolonging time, and potent bystander effect. ......