DETECTION OF COLORECTAL-CANCER K-RAS MUTATIONS USING A SIMPLIFIED OLIGONUCLEOTIDE LIGATION ASSAY

It has been suggested that some mutations in codons 12 and 13 of the K -ras gene are associated with the progression of colorectal adenomas t o carcinomas. The aim of this study was to develop a rapid, colorimetr ic assay for K-ras point mutations commonly associated with colorectal cancer. K-ras exon 1 was amplified from colorectal tumor DNA and K-ra s activating mutations detected using an oligonucleotide ligation assa y (OLA) in combination with immunological and colorimetric detection. Using the OLA with oligonucleotides specific to individual K-ms mutati ons, 6 (of 17 total colorectal adenomas/carcinomas) were found to have K-ras mutations. The assay could detect as little as 10% mutant allel e. A simplified OLA designed to test for either the presence (+) or ab sence (-) of any of the K-rns activating mutations was developed, The assay was further streamlined by use of a dipstick methodology for col orimetric development. If required, assay sensitivity can be increased by the use of the recently described EDNA-ELCA detection system. The simplified (+/-) mutation OLA in combination with a dipstick or EDNA-E LCA detection system provides a rapid, sensitive assay for K-ras point mutations suitable for use as part of the clinical assessment of colo rectal cancer. (C) 1997 Elsevier Science B.V.