The 5-year survival rate of patients with squamous cell carcinoma of the head and neck (HNSCC) has remained poor despite innovative surgery and new radiation and chemotherapeutic strategies. In such patients, gene therapy relying on the modification of tumor cells by gene transfer may have great potential as a new treatment modality in the therapy of HNSCC. In the present study we developed an in vitro model to show the efficacy and technical feasibility of cationic liposome-mediated gene transfer into HNSCC. Five adherent squamous cell carcinoma cell lines were transfected with SV40- or CMV-promoter-driven CAT (chloramphenicol-acetyl-transferase)-expression plasmids using DOTAP as the liposome carrier. The level of CAT expression was shown to correlate directly with the amount of transfected DNA and could be measured by a CAT-enzyme-linked immunosorbent assay. The results of gene transfer by liposome-DNA complexes obtained for all cell lines showed a dose-dependent efficacy correlating to the amount of DOTAP employed. The data demonstrate the successful in vitro transfection of epithelial cell lines with DNA, suggesting its usefulness as a new tool for head and neck cancer therapy in vivo.