Oral squamous cell carcinoma (OSCC) is associated with heavy smoking and drinking, but the molecular pathway of tumorigenesis is not understood. Inactivation of the p53 tumor suppressor gene is likely to play an important role since p53 mutation is frequently found. The p14ARF tumor suppressor gene is functionally linked to p53, because it is activated by oncogenes and causes p53-dependent growth arrest and apoptosis. The relationship between p14ARF and p53 inactivation has not been described for OSCC. We studied 25 cases of OSCC to determine if there is an inverse correlation between p53 mutation and p14ARF inactivation by homozygous deletion or mutation. p53 mutation was found in 16 of 25 cases (64%), including nine missense and seven truncating mutations. While all cases with missense mutations showed abnormal accumulation of p53 protein, there were also five carcinomas which showed increased p53 staining in the absence of mutation. p14ARF deletion or mutation was found in eight cases (32%), six of which also demonstrated p53 mutation. Our findings indicate that OSCC often involves loss of both p14ARF and p53 function and suggest that inactivation of these two tumor suppressor genes are not functionally equivalent during tumorigenesis.