To evaluate the effects of adenovirus (Ad)-mediated transfer of p53 and p16 on human bladder cancer cells EJ, EJ were transfected with Ad-p53 and Ad-p16. Cell growth, morphological change, cell cycle, apoptosis were measured using MTT assay, flow cytometry, cloning formation, immunocytochemical assays. Ad-p16 or Ad-p53 alone could inhibit the proliferating activity of EJ cells in vitro. Ad-p53 could induce apoptosis of partial EJ cells. G1 arrest was observed 72 h after infection with Ad-p16, but apoptosis was not obvious. The transfer of Ad-p16 and Ad-p53 could significantly inhibit the growth of EJ cells, decrease the cloning formation rate and induce apoptosis of large number of EJ cells. The occurrence time of subcutaneous tumor was delayed and the tumor volume in 4 weeks was diminished by using Ad-p53 combined with Ad-p16 and the difference was significant compared with using Ad-p53 or Ad-p16 alone. It was suggested that the transfer of wild-type p53 and p16 could significantly inhibit the growth of human bladder cancer in vitro and in vivo.