Multi‐Regional Comprehensive Genomic Profiling of Diffuse‐Type Cholangiocarcinoma in Five Patients

BACKGROUND/PURPOSE: Differentiating between the monoclonal spread of a single tumor and multicentric carcinogenesis in perihilar cholangiocarcinoma (PHC) with multiple nodules is a major clinical challenge. The clonal origins of these multifocal tumors remain unclear. This study used multi-regional comprehensive genomic profiling (CGP) to investigate its biological basis. METHODS: We retrospectively identified 5 patients with extensive PHC and multiple nodules from 175 resected cases. Fourteen spatially distinct tumor nodules were collected from surgical specimens. Each underwent CGP using a 216-gene panel to compare driver gene alterations and assess clonal relationships. RESULTS: Multi-regional analysis revealed a monoclonal origin in 4 of 5 cases, with identical driver mutations in KRAS, TP53, and ARID1A across nodules within each patient. One case was a polyclonal tumor (a possible collision-type process), with distinct mutation profiles (TP53/BRAF vs. KDM6A/SMARCA4) in separate nodules. CONCLUSIONS: In this five-patient series, four tumors were monoclonal, likely spreading via intraductal seeding or intramural metastasis, whereas one was a polyclonal tumor not reliably distinguished by pathology alone. Although routine multi-regional genomic analysis is not yet feasible, these hypothesis-generating findings suggest two distinct biological pathways and require validation in larger cohorts.