Utility of DNA flow cytometry in distinguishing between malignant and benign intrahepatic biliary lesions

The distinction between well-differentiated intrahepatic cholangiocarcinoma (iCCA) from its morphological mimics such as bile duct adenoma (BDA) and hamartoma (BDH) can be challenging, particularly in small biopsies. Although a few cases of BDA and BDH have been reported to undergo malignant transformation into iCCA, their neoplastic versus benign nature remains debated. DNA flow cytometry was performed on 47 formalin-fixed paraffin-embedded samples of iCCA, 14 BDA, and 18 BDH. Aneuploidy was detected in 22 iCCA (47%) but in none of the 32 BDA and BDH samples. Among the 34 iCCA patients who underwent complete resection and were followed up to tumor recurrence, tumor-related death, or at least for 1 year, the overall recurrence or death rates (regardless of flow cytometric results) were 18, 56, and 71% within 1, 3, and 5 years, respectively. The 1-, 3-, and 5-year recurrence or death rates in 18 iCCA patients with aneuploidy were 28, 66, and 66%, respectively, whereas 16 iCCA patients in the setting of normal DNA content had 1-, 3-, and 5-year rates of 6, 44, and 72%, respectively. Although aneuploid tumors were associated with worse outcomes during the first 3 years, this difference was not statistically significant (hazard ratio = 1.4, p = 0.473) in the present sample size. In conclusion, the frequency of aneuploidy was significantly higher in iCCA (47%) than in its benign morphological mimics (0%), suggesting that it may potentially serve as a diagnostic marker of malignancy in challenging situations. Our findings also suggest that most BDAs and BDHs, if not all, are benign entities and may not represent precursor lesions to iCCAs that often harbor aneuploidy. Although a larger cohort will be necessary to further determine the prognostic significance of aneuploidy in iCCA patients after resection, the patients with aneuploid tumors may have a higher risk for tumor progression, especially during the first 3 years.