Mutations of the tumor-suppressor gene TP53 and amplification of CCNDI gene have been reported to occur frequently in head and neck squamous cell carcinomas (HNSQCC). In experimental systems, TP53 mutations have been shown to lead to genomic instability, including an increased propensity for gene amplification. We have examined I6 HNSQCC cell lines for the association between TP53 over-expression/mutation and CCNDI amplification. p53 over-expression was detected in 50% of the cell lines by immunohistochemistry using the monoclonal antibody (MAb) PAbI80l. TP53 mutations were also detected in 50% of the cell lines by analysis of single-strand conformation polymorphism (SSCP) and DNA sequencing of exons 4 through 9. Six cell lines showed TP53 mutations and over-expression of the protein, 2 cell lines showed TP53 mutations but no p53 expression, and 2 cell lines showed over-expression of p53 protein but no TP53 gene mutations. CCNDI amplification was found in 38% of the cell lines by Southern blot analysis. Only I cell line showed both TP53 mutation and CCNDI amplification, whereas 7 of 8 cell lines with TP53 mutations had no CCNDI amplification. pRb expression was detected by Western blot analysis, and the level of pRb did not correlate with either CCNDI amplification or TP53 mutation. Our findings suggest that TP53 mutation and CCNDI amplification are common genetic alterations in HNSQCC and that the occurrence of either genetic event may be sufficient to abrogate normal cell cycle control. © 1994 Wiley-Liss, Inc.