We reconstructed the recombinant p53-expressing adenovirus and examined its infections and effects in head and neck squamous cell carcinoma cell lines. Eight human head and neck squamous cell carcinoma cell lines were infected by the recombinant adenovirus harboring the lacZ gene (AxCAiLacZ) or the wild-type p53 gene (AxCAip53), and the effects were investigated. The eight cell lines were successfully infected by AxCAiLacZ at a level of more than 50%. The survival of all 8 squamous cell lines were inhibited in the range from 8 to 26.7% by only one treatment of the AxCAip53 infection. This result suggested that p53 gene therapy might become a useful tool in head and neck squamous cell carcinoma treatment.