Little is known regarding the molecular genetic events in head and neck carcinoma. Epidemiological evidence suggests that both alcohol and tobacco use are related to the development of these neoplasms, and viral infections have also been postulated to play a role in some tumors. Loss of p53 tumor suppressor gene function has been found in many malignancies and can occur through either gene mutation or by interaction with the E6 protein of oncogenic human papilloma viruses (HPV). Because the mucosal surfaces of the head and neck are exposed to mutagens and HPVs, we studied DNA derived from 30 stage I-IV squamous cell carcinomas of the head and neck (9 primary tumors and 21 early passage cell lines) for p53 gene mutations as well as for the presence of oncogenic HPV DNA. Exons 2 through 11 of the p53 gene were examined using single strand conformation polymorphism analysis followed by direct genomic sequencing of all variants. HPV detection was done using polymerase chain reaction amplification with HPV E6 region type specific primers as well as L1 region degenerate ("consensus") primers; HPV type was determined by restriction fragment length polymorphism analysis of the amplified fragment as well as by Southern blotting of genomic DNA. Sixteen of 30 tumors (53%) had p53 mutations and oncogenic HPV DNA was detected in 3 of 30 (10%) tumors, none of which had p53 mutations. The p53 mutational spectrum observed was characterized by equal frequencies of transversions (6 of 16), transitions (5 of 16), and deletions (5 of 16). This distribution of mutations differs from the spectrum of p53 mutation reported in esophageal (P = 0.05) and lung (P = 0.02) cancers, two other tobacco associated neoplasms. A previously undescribed clustering of 3 mutations at codon 205 was also observed. A trend toward a shorter time to tumor recurrence after treatment was noted for those patients with tumors exhibiting p53 gene mutations, and no relationship between p53 mutations and tumor stage or node status was noted. Alteration in p53 gene function appears common in head and neck cancer, and the mutational spectrum observed may reflect the role of different mutagens or mutagenic processes than those responsible for the p53 mutations in lung and esophageal neoplasms.