Many human cervical and oral carcinomas express RNA of human papillomaviruses, and the RNA transcript provides a potential target for gene therapy of these carcinomas. Three hammerhead ribozymes that were targeted to RNA of human papillomavirus type 18 (HPV-18) were cloned into a plasmid expression vector. Each plasmid was then transfected into the HPV-18-expressing cell line. HeLa, or the non-HPV-expressing oral cancer cell line, Tu167. None of the ribozymes had any effect on the phenotype of Tu167 cells. In contrast, each ribozyme affected the phenotype of HeLa cells, causing reduced growth rates, increased serum dependency, and reduced focus formation in soft agar. A molecule that had the same antisense sequences as a ribozyme but lacked the catalytic sequences affected the HeLa cell phenotype to a much lesser extent. The effects of two of the ribozymes could be attributed in part to an increased intracellular concentration of the tumor suppressor protein p53. The most effective ribozyme was targeted to nucleotide 309 in the HPV-18 transcript, but each of the three ribozymes appears to have potential for gene therapy of cancers that express HPV-18.