To investigate the effect of low dose irradiation on gene transfer efficiency and the effect of adenovi- ral-mediated exogenous P53 overexpression on radiosensitivity of radioresistant human melanoma cell line A375 with wild type p53

control vector

a replication deficient recombinant adenoviral vector containing a CMV promoter and green fluorescent protein (AdCMV-GFP)

was used to transfect the A375 cells preirradiated with or without 1 Gy X-ray radiation. The transduction efficiency of GFP gene was determined with fluorescence microscope directly. A375 cells radiated by 1 Gy X-ray were transfected with a replication deficient recombinant adenoviral vector carry- ing human wild p53 (AdCMV-P53)

and mRAN level was detected by RT-PCR. The cell cycle delay and the expres- sion of exogenous P53 were detected using flow cytometry (FCM) at different time after transfection. The radiosen- sivity of A375 cells after p53 transduction was assayed by clonoy formation. We found that 1Gy exposure increased the gene transfer efficiency of A375 cells. The expression of exogenous P53 was found to be 60% to 80% of trans- fected cells during the first three days after transduction and then declined continuously down to the control level on the day 10. The G1 cell cycle arrest was also observed after p53 gene transfer. A375 cells that were transfected with p53 showed higher sensitivity to X-ray –induced cell killing than those cells that either were transfected with the viral vector carrying a green fluorescent protein gene or were not transfected at all. Low dose ionizing radiation can im- prove gene transfer efficiency of A375 cells mediated by adenovirus vector. Althrough the overexpresion of exoge- nous P53 may not inhibit cell growth and induce apoptosis of melanoma cell line A375 in vitro

it made the tumor cells much sensitive to death by irradiation. The data suggested that p53 gene might be a potential gene for melanoma therapy and provide the experimental evidences to clinically using the combination of radiation with gene therapy on melanoma. Namely

there may be a reduction of side effects resulting from high dose radiation on radioresistant tu- mors or rAd-P53 administration alone in clinical applications.