DNA flow cytometric analysis of paraffin-embedded tissue for the diagnosis of malignancy in bile duct biopsies

Abstract

Differentiation of reactive versus neoplastic epithelial changes can be challenging in bile duct biopsies. The samples are often scant, distorted, and mixed with significant inflammation, ulceration, and/or debris. Histological confirmation of malignancy is often required before the initiation of surgical therapy, and an erroneous diagnosis of malignancy can lead to unnecessary clinical management. Aneuploidy assessment by DNA flow cytometry was performed on formalin-fixed paraffin-embedded (FFPE) tissue from 63 bile duct biopsies: 10 with a malignant diagnosis (7 with adenocarcinoma and 3with at least high-grade dysplasia [HGD]); 3 with an atypicaldiagnosis showing rare atypical glands/cells, concerning but not definite for malignancy; 28 likely reactivebiopsies with acute/chronic inflammation, ulceration, and/or mild nuclear atypia; and 22 additional benign biopsies without significant inflammation, ulceration, or nuclear atypia. Aneuploidy was detected in 7 (70%) of the 10 biopsies with definite neoplasia (5 of 7 adenocarcinoma cases and 2 of 3at least HGD cases), all 3 (100%) atypicalbiopsies, and none of the 50 benign biopsies. All 3 atypicalcases with aneuploidy were subsequently found to have adenocarcinoma (n=2) or HGD (n=1). Among the 2 cases of at least HGD with aneuploidy, 1 case developed adenocarcinoma, but no follow-up information was available in the other case. The remaining 1 case of at least HGD, despite having normal DNA content, was found to have adenocarcinoma on follow-up. None of the 50 benign cases (further supported by normal DNA content) developed adenocarcinoma within a mean follow-up time of 37 months (range: 0-282 months). The estimated sensitivity of aneuploidy as a diagnostic marker of malignancy (adenocarcinoma and HGD) was 70%, with the specificity of 100%, positive predictive value of 100%, and negative predictive value of 94%. In conclusion, DNA flow cytometry using FFPE tissue from bile duct biopsies demonstrates a high rate of aneuploidy (70%) in malignant casesand normal DNA content in all benign biopsies. Although the sample size is small, the results indicate that this assay can be potentially useful in challenging atypicalcases, where morphological evaluation is limited by scarcity of atypical glands/cells, inflammation, and/or ulceration.