By using a modified polymerase chain reaction strategy, we have devised an approach to detect a K-ras oncogene mutated at codon 12 in the presence of 1000 normal alleles. This is a considerable improvement in sensitivity on previous assays. Application of this assay to 15 cholangiocarcinomas showed that all contained a K-ras mutation at codon 12 and that nine of the tumors contained two or more mutations. In 11 cases, mutations were present in less than 10% of the cells in the sample. In common with pancreatic adenocarcinomas, in which 75 to 95% of cases contain a mutation in K-ras, cholangiocarcinomas show a very high frequency of ras gene mutation, but within a tumor only a fraction of cells contain a ras mutation. The presence of multiple mutations and the low frequency of mutant alleles in the samples argue against K-ras mutations being the initiating genetic lesion in this tumor, but suggest that ras gene mutation is involved in the stepwise progression of neoplastic cells to full malignancy.
Work in the laboratory of C. J. M. is supported by a grant to the Institute of Cancer Research from the Cancer Research Campaign and Medical Research Council. S. L. was supported by a Cancer Research Campaign Clinical Research Fellowship.