MOLECULAR EVENTS INCLUDING P53 AND K-RAS ALTERATIONS IN THE IN-VITRO PROGRESSION OF A HUMAN COLORECTAL ADENOMA CELL-LINE TO AN ADENOCARCINOMA

The aim of the current study was to identify genetic abnormalities in human colorectal adenoma and carcinoma derived cell lines, and to dete rmine whether the genetic changes which occur in vitro are relevant to the in vivo situation. Loss of 1p(33-35) region was shown to be the m ost common chromosome 1 abnormality and loss of heterozygosity (LOH) o f the DCC gene and/or adjacent sequences was detected in all adenoma d erived cells as well as the carcinoma cell lines. The level of p53 pro tein was also investigated as increased cellular p53 protein had previ ously been associated with mutation of the p53 gene. A further aim was to investigate genetic changes in our in vitro model of tumour progre ssion, where the adenoma derived PC/AA cell line has previously been c onverted in vitro to two distinct tumorigenic phenotypes, producing ei ther an adenocarcinoma or a mucinous carcinoma in athymic nude mice. P rogression to the adenocarcinoma phenotype was shown to involve a spec ific chromosome 1 rearrangement, loss of both normal copies of chromos ome 18 (although DCC gene sequences were retained), loss of the remain ing wild type allele of k-ras resulting in homozygosity for the k-ras codon 12 mutation and increased cellular p53 protein as detected by SD S-PAGE Western blotting. The increase in p53 protein was shown not to be due to the acquisition of a mutation in the p53 gene. Interestingly , progression of the adenoma derived PC/AA cell line to the mucinous m alignant phenotype did not involve any of these molecular rearrangemen ts, suggesting that different genetically distinct pathways are involv ed in colorectal carcinogenesis. These studies show that the genetic c hanges in our in vitro model of human colorectal tumour progression ar e similar to those observed in in vivo studies.