Single-cell Transcriptomic Architecture Unraveling the Complexity of Tumor Heterogeneity in Distal Cholangiocarcinoma

BACKGROUND & AIMS: Distal cholangiocarcinoma (dCCA) are a group of epithelial cell malignancies that occurs at the distal common bile duct, and account for approximately 40% of all cholangiocarcinoma cases. dCCA remains a highly lethal disease as it typically features remarkable cellular heterogeneity. A comprehensive exploration of cellular diversity and the tumor microenvironment is essential to depict the mechanisms driving dCCA progression. METHODS: Single-cell RNA sequencing was used here to dissect the heterogeneity landscape and tumor microenvironment composition of human dCCAs. Seven human dCCAs and adjacent normal bile duct samples were included in the current study for single-cell RNA sequencing and subsequent validation approaches. Additionally, the results of the analyses were compared with bulk transcriptomic datasets from extrahepatic cholangiocarcinoma and single-cell RNA data from intrahepatic cholangiocarcinoma. RESULTS: We sequenced a total of 49,717 single cells derived from human dCCAs and adjacent tissues, identifying 11 distinct cell types. Malignant cells displayed remarkable inter- and intra-tumor heterogeneity with 5 distinct subsets were defined in tumor samples. The malignant cells displayed variable degree of aneuploidy, which can be classified into low- and high-copy number variation groups based on either amplification or deletion of chr17q12 - chr17q21.2. Additionally, we identified 4 distinct T lymphocytes subsets, of which cytotoxic CD8+ T cells predominated as effectors in tumor tissues, whereas tumor infiltrating FOXP3+ CD4+ regulatory T cells exhibited highly immunosuppressive characteristics. CONCLUSION: Our single-cell transcriptomic dataset depicts the inter- and intra-tumor heterogeneity of human dCCAs at the expression level.