Detection of Ki-ras Mutations by PCR and Differential Hybridization and of p53 Mutations by SSCP Analysis in Endoscopically Obtained Lavage Solution from Patients with Long-Standing Ulcerative Colitis

Objectives : The goal of this study was the early detection of malignant transformation in patients with longstanding ulcerative colitis; therefore, mutations of the Ki-ras and p53 gene were analyzed in lavage solution and biopsies obtained at surveillance colonoscopy. Methods : DNA was isolated from 14 patients (nine female, five male) with a history of pancolitis for more than 10 yr. Exon 1 of the Ki-ras gene and exons 5–8 of the p53 gene were amplified via polymerase chain reaction. Mutations of the p53 gene were detected via single-strand conformation polymorphism analysis; point mutations of the Ki-ras gene were hybridized on dot blots with oligonucleotides marked with digoxigenin. Results : Wild-type Ki-ras and wild-type p53 were detected in all cases of ulcerative colitis and in four of seven control patients. In two ulcerative colitis patients, a mutation was found in the Ki-ras gene (Gly → Asp 12 and Gly → Val 12), and in one patient, a mutation in exon 5 of the p53 gene. Mutations were found only in the lavage fluid, whereas random biopsies were negative. Conclusions : From colonic lavage fluid, it is possible to extract DNA of sufficient quantity and quality for polymerase chain reaction-based amplification and subsequent analysis via single-strand conformation polymorphism or hybridization. Mutations were found in three of 14 patients with long-standing ulcerative colitis but were not found in controls. The method may he useful for the screening of such patients.