PRESENCE OF HUMAN PAPILLOMAVIRUS SEQUENCES IN TUMOR-DERIVED HUMAN ORAL KERATINOCYTES EXPRESSING MUTANT P53

A series of eight oral epithelial cell Lines derived from untreated hu man oral squamous cell carcinomas, which had arisen in patients with d ifferent tobacco histories, were examined for the presence of human pa pillomavirus (HPV) DNA, expression of stable p53 protein and p53 point mutation. Polymerase chain reaction (PCR)-based screening, but not So uthern blot analysis, showed HPV-16 early region sequences to be prese nt at low copy number(<1 copy per cell) in two cell lines at early pas sage (3-5) in vitro (H400, T45), implying that only subpopulations of cells harboured viral DNA. HPV sequences were undetectable in cells at later passage (12-15), suggesting that viral sequences had been lost during growth in vitro, or that negative selection of HPV-containing c ells had occurred. High levels of p53 were detected in the two HPV-pos itive cell lines and in three others (H103, H314, H357) by Western blo tting, suggesting expression of mutant (stable) p53 molecules. A sixth cell line (H157) expressed a truncated p53. Sequence analysis of exon s 2-11 of the p53 gene revealed missense mutations in six cell lines, one of which (H413) did not result in high levels of protein, and nons ense mutations in the remaining two cell lines (H157, H376). The resul ts suggest that p53 mutation is a frequent genetic event in oral cance r. In addition, the expression of mutant p53 in oral cancer cells does not preclude a papillomaviral aetiology for these tumours. Analysis o f p53 expression alone may result in underestimation of the frequency of p53 mutations in human cancers. In contrast to other studies, we de monstrate that positive staining of p53 in oral cancer does not necess arily reflect a tobacco aetiology.