K-ras codon 12 mutations in biliary tract tumors detected by polymerase chain reaction denaturing gradient gel electrophoresis

Abstract

Although the prevalence of K-ras codon 12 mutations in biliary tract (BT) tumors has been addressed in previous studies, the results have shown large discrepancies in mutation frequency.K-ras codon 12 mutations were investigated by polymerase chain reaction (PCR)-denaturing gradient gel electrophoresis (DGGE), a sensitive method for detecting DNA base changes, in a large series of BT tumors.In A-549 cells, which are known to contain a G to A change at the first base of K-ras codon 12, the mutation could be detected by DGGE even after 1:16 dilution with normal DNA. Tumor samples were microdissected from paraffin embedded tissue sections to ensure the presence of the tumor cells. K-ras mutations were detected in 13 of 23 bile duct tumors (56.5%) and in 9 of 23 gallbladder tumors (39.1%) by DGGE. However, no mutations were detected in normal, hyperplastic, and dysplastic BT epithelium or in tumorlike lesions, such as adenomyomatous hyperplasia, cholesterol polyps, and cystitis glandularis proliferans. The samples exhibiting abnormalities on DGGE showed a base change at K-ras codon 12 when examined by oligonucleotide hybridization.K-ras codon 12 mutations are seen often in BT tumors, and a combination of microdissection and PCR-DGGE is an effective approach for their detection.