Usefulness of p53 gene mutations in the supernatant of bile for diagnosis of biliary tract carcinoma: comparison with K-ras mutation
The sensitivity of bile cytology for the diagnosis of biliary tract carcinoma (BTCa) is still low. In addition, the incidence of detection of genetic mutations in the bile of BTCa is not satisfactory yet. To improve the molecular diagnosis of BTCa, we analyzed p53 and K-ras mutations in DNA extracted from not only the sediment but the supernatant of bile samples. Methods: Polymerase chain reaction-single-strand conformation polymorphism and direct sequencing were used for analyses of p53 mutations in exons 5 through 8. K-ras mutations at codon 12 were examined by mutant allele-specific amplification. Results: In bile supernatant from patients with BTCa, p53 and K-ras mutations were detected in 50.0% (15/30) and 56.7% (17/30) of cases, respectively. The incidence of p53 and K-ras mutations in the sediment was 33.3% and 43.3%, respectively. When a combination assay with both genes was used, molecular abnormalities were detected in 80.0% of cases, including 3 in which p53 alone was positive. In addition, either p53 or K-ras mutations were detected in 12 of 15 (80.0%) cases of BTCa in which the cytologic diagnoses were negative. p53 mutations were detected in neither supernatant nor sediment in 20 patients with cholelithiasis, although the incidence of K-ras mutations in the sediment was 20%. Conclusions: The incidence of p53 and K-ras mutations is higher in the supernatant than in the sediment, and simultaneous analyses of p53 and K-ras in the two bile fractions could enhance the genetic diagnosis of BTCa. Notably, the specificity of p53 mutations for cancer was very high in bile samples, and the sensitivity was also relatively high.
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