Whole Transcriptome Sequencing Reveals a GATA4-INHA Program and Progenitor Cell of Origin in Cholangioblastic Cholangiocarcinoma

Cholangioblastic cholangiocarcinoma (CBCA) is a rare hepatobiliary neoplasm defined by the NIPBL::NACC1 fusion. Its cell of origin and nomenclature remain unresolved, and some investigators favor reclassification as a fusion-positive hepatic carcinoma rather than a distinctive cholangiocarcinoma variant. We used whole transcriptome sequencing (WTS) to define the CBCA transcriptional program and differentiation state relative to other hepatobiliary malignancies. In total, 334 hepatobiliary neoplasms were subjected to WTS from multiple institutions, spanning eight diagnostic categories including 10 CBCA and 163 intrahepatic cholangiocarcinoma (iCCA). Clustering, differential expression, gene set enrichment, immune deconvolution, and cell-of-origin analyses were performed, and GATA4 immunohistochemistry was validated on an independent cohort of 445 cases. The NIPBL::NACC1 fusion gene was identified in all 10 CBCA and in none of the remaining 324 neoplasms. Diagnostic category accounted for 52.7% of transcriptomic variance (p=0.001), and CBCA formed a discrete cluster nearest iCCA despite limited gene-level overlap, and harbored more differentially expressed genes than any category (n=1,451 vs. all combined). GATA4 was upregulated (log₂FC=3.23, p<0.001), with upregulation of 14 of 33 GATA4 target genes including INHA (largest fold change), corresponding to GATA4 protein overexpression in 6 of 6 CBCA, 2% of iCCA, and 0 of 278 HCC. Synaptic and neuroendocrine programs were the most enriched pathways, and cell-of-origin analysis placed CBCA in an intermediate biliary progenitor state with the highest Bipotency Index of any category (p≤0.001) and near-complete immune exclusion. Together, these findings define a GATA4-INHA transcriptional program downstream of NIPBL::NACC1 that accounts for the inhibin A immunophenotype of CBCA. This biliary progenitor state supports the cholangioblastic designation over a hepatocellular carcinoma origin, confirmed at the protein level by GATA4 immunohistochemistry.