MUTATION OF THE P53 GENE PRECEDES ANEUPLOID CLONAL DIVERGENCE IN COLORECTAL-CARCINOMA

To establish whether p53 mutation precedes or follows clonal divergenc e in human colorectal carcinomas, 17 tumours were analysed at multiple sites (2-5 each) for single-strand conformation polymorphisms (SSCP) within exons 5-8 of the p53 gene. A previous study had demonstrated su bclones of differing DNA ploidy in these tumours, but all showed immun ocytochemical evidence for p53 stabilisation, using the monoclonal ant ibody PAb 1801. Mutations within exons 5-8 of p53 were identified by t he presence of an abnormally migrating band in 10 of the 17 carcinomas : five in exon 5, four in exon 7 and one in exon 8. In each of these p ositive cases, samples from different parts of the carcinoma showed id entical gel migration patterns in SSCP analysis. Similarly, the remain ing seven rumours were concordant for absence of band shift across all samples of each tumour. Six SSCP-positive cases contained multiple po pulations differing in DNA ploidy, while four were homogeneously diplo id or aneuploid throughout. Very similar proportions were observed in the SSCP-negative cases. In four positive tumours the mutation was con firmed by sequencing or through alteration of nucleotide-specific rest riction enzyme cleavage. Identical mutations appeared in every sample from the same tumour. The results provide unequivocal evidence that th e same mutant allele of p53 is present throughout each tumour bearing a mutation, regardless of the clonal variation identified by analysis of DNA ploidy. We conclude that in colorectal tumorigenesis mutation o f p53 occurs as a single event which precedes and may facilitate the a neuploid clonal divergence of carcinomas.