Leigh, Sarah Elizabeth Anne; (1993) Alterations in genomic organization and gene expression in colorectal carcinogenesis. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

This study has examined some of the features that characterize the changing pattern of genomic organization and gene expression in colorectal carcinogenesis. The transition from normal colonic mucosa, to adenoma, to carcinoma is accompanied by the progressive accumulation of genetic defects. DNAs from a panel of premalignant adenomas, predominantly from familial adenomatous polyposis (FAP) patients, and carcinomas mostly from non-FAP patients, were screened for the presence of somatic mutations. RNA was extracted from a similar panel of samples for use in gene expression studies. Allele losses were detected in tumour DNA samples with polymorphic markers from chromosomes 1, 5q, 7 and 11p. Such loss of genetic material may indicate the presence of tumour suppressor genes at affected loci. As the adenomatous polyposis coli gene has been assigned to 5q21, allele loss in this region was expected in carcinomas, however loss of chromosome 5q markers had not previously been reported in premalignant adenomas. The absence of allele loss on chromosome 3p, suggests that the small cell lung cancer tumour suppressor gene, was not involved in colorectal carcinogenesis. Genetic instability was manifest in some carcinoma samples by the generation of novel alleles at various hypervariable loci. The retinoblastoma susceptibility gene was over-represented in two carcinomas and elevated levels of expression were detected in RNA from 90% of adenomas and 50% of carcinomas. Such uncharacteristic findings may indicate that unknown factors are interacting with this tumour suppressor gene in colorectal tumours. Expression of the p53 tumour suppressor gene declined from high levels in adenomas to low levels in carcinomas. Comparison of these findings with published data, suggests that an inverse relationship may exist between gene expression and mutation at this locus. The most striking and consistent change observed in this study, was the loss of carbonic anhydrase 1 gene expression associated with epithelial de-differentiation. Expression of the mucin genes also declined with the progression of colorectal carcinogenesis. In mucinous tumours however, although mucin transcript levels were high in some cases, the pattern of mucin gene expression varied between individual samples. Construction of a normal colonic mucosal cDNA library, allowed a cross hybridization strategy to be employed, in an attempt to isolate clones from human chromosome five that contained sequences expressed in the colon. Six clones were isolated and their preliminary characterization undertaken.

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