Study of the Role of Circulating Cell-Free DNA in the Diagnosis of Cholangiocarcinoma

EL- Khaiat, Mohsen Mohammed; El Hagary, Heba Mostafa; Aly, Atef Abou Elsoud; Nada, Ali Saadeldin; EI-Abd, Mona Gamal; Abourisha, Khaled Abdelmonem; Teima, Ahmed Abo-Zaid Ahmed

doi:10.31557/APJCP.2026.27.7.2697

Study of the Role of Circulating Cell-Free DNA in the Diagnosis of Cholangiocarcinoma

Document Type : Research Articles

Authors

¹ Tropical Medicine Department, Faculty of Medicine, Menoufia University, Menoufia, Egypt.

² Hepatology and Gastroenterology Department, National Liver Institute, Menoufia University, Menoufia, Egypt.

³ Clinical Pathology department, National Liver Institute, Menoufia University, Menoufia, Egypt.

10.31557/APJCP.2026.27.7.2697

Abstract

Background: Cholangiocarcinoma (CCA) is an aggressive biliary malignancy usually diagnosed at advanced stages because early symptoms are vague. Conventional serum markers, including carcinoembryonic antigen (CEA) and carbohydrate antigen 19-9 (CA19-9), have limited sensitivity and specificity. Circulating cell-free DNA (cfDNA), particularly ALU-115, ALU-247 and DNA integrity index, may provide improved diagnostic accuracy by reflecting tumor-derived DNA fragmentation. Methods: A case-control study enrolled 160 subjects divided into four groups, each comprising 40 individuals: CCA (Group I), other hepatobiliary malignancies including hepatocellular carcinoma and pancreatic head cancer (Group II), benign biliary disorders (Group III), and healthy controls (Group IV). Clinical data, biochemical parameters, tumor markers, and cfDNA fragments were assessed. cfDNA was quantified by real-time PCR using ALU-115 (short fragments) and ALU-247 (long fragments). The DNA integrity index was calculated. Receiver operating characteristic (ROC) curves were used to determine diagnostic performance. Results: Serum CEA and CA19-9 were significantly higher in Groups I and II compared with Groups III and IV (p < 0.001), but they showed limited discrimination between CCA and other malignancies. cfDNA markers demonstrated significantly higher values in CCA than in other groups (p < 0.001). Against Group II, ALU-247 >800 copies/ml yielded 80% sensitivity and 75% specificity, while DNA integrity >0.89 achieved 70% sensitivity and 90% specificity. Against Group III, ALU-115 >700 copies/ml showed 85% sensitivity and 90% specificity. Versus controls, an ALU-115 >580 copies/ml reached 95% sensitivity and 95% specificity. Conclusion: cfDNA concentrations and integrity indices outperform CEA and CA19-9 for distinguishing CCA from other malignancies, benign biliary disease, and healthy subjects. These findings support the incorporation of cfDNA-based assays as minimally invasive diagnostic adjuncts for CCA pending larger validation studies.

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