Clinicopathologic features, tumor immune microenvironment and genomic landscape of Epstein-Barr virus-associated intrahepatic cholangiocarcinoma

J Hepatol. 2021 Apr;74(4):838-849. doi: 10.1016/j.jhep.2020.10.037. Epub 2020 Nov 17.


Background & aims: Little is known about Epstein-Barr virus (EBV)-associated intrahepatic cholangiocarcinoma (EBVaICC) because of its rarity. We aimed to comprehensively investigate the clinicopathology, tumor immune microenvironment (TIME) and genomic landscape of this entity in southern China.

Methods: We evaluated 303 intrahepatic cholangiocarcinomas (ICCs) using in situ hybridization for EBV. We compared clinicopathological parameters between EBVaICC and nonEBVaICC, and we analyzed EBV infection status, tumor-infiltrating lymphocytes (TILs) and genomic features of EBVaICC by immunohistochemistry, double staining, nested PCR, multiplex immunofluorescence staining, fluorescence in situ hybridization and whole-exome sequencing.

Results: EBVaICC accounted for 6.6% of ICCs and was associated with EBV latency type I infection and clonal EBV isolates. Patients with EBVaICC were more often female and younger, with solitary tumors, higher HBV infection rates and less frequent cirrhosis; the lymphoepithelioma-like (LEL) subtype was more common in EBVaICC. EBVaICC was associated with a significantly larger TIME component than nonEBVaICC. The LEL subtype of EBVaICC - associated with a significantly increased density and proportion of CD20+ B cells and CD8+ T cells - was associated with significantly higher 2-year survival rates than conventional EBVaICC and nonEBVaICC. Both PD-1 and PD-L1 in TILs, and PD-L1 in tumor cells, were overexpressed in EBVaICC. High PD-L1 expression in tumor cells and high CD8+ TIL densities were significantly more common in EBVaICC than in nonEBVaICC. Seven genes (MUC4, DNAH1, GLI2, LIPE, MYH7, RP11-766F14.2 and WDR36) were mutated in at least 3 patients. EBVaICC had a different mutational pattern to liver fluke-associated cholangiocarcinoma and HBV-associated ICC.

Conclusions: EBVaICC, as a subset of ICC, has unique etiological, clinicopathological and genetic characteristics, with a significantly larger TIME component. Paradoxically, patients with EBVaICC could be candidates for immune checkpoint therapy.

Lay summary: Epstein-Barr virus (EBV) is associated with a subtype of intrahepatic cholangiocarcinoma, with unique clinicopathological and genetic characteristics. The tumor immune microenvironment is also different in this tumor subtype and patients with EBV-associated intrahepatic cholangiocarcinoma may respond well to immune checkpoint inhibitors.

Keywords: Clinicopathology; Epstein-Barr virus; Genomic landscape; Intrahepatic cholangiocarcinoma; Tumor immune microenvironment.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • B7-H1 Antigen / genetics*
  • Bile Duct Neoplasms* / genetics
  • Bile Duct Neoplasms* / mortality
  • Bile Duct Neoplasms* / pathology
  • Bile Duct Neoplasms* / therapy
  • CD8-Positive T-Lymphocytes / pathology
  • China / epidemiology
  • Cholangiocarcinoma* / genetics
  • Cholangiocarcinoma* / mortality
  • Cholangiocarcinoma* / pathology
  • Cholangiocarcinoma* / therapy
  • Epstein-Barr Virus Infections* / complications
  • Epstein-Barr Virus Infections* / diagnosis
  • Epstein-Barr Virus Infections* / epidemiology
  • Female
  • Herpesvirus 4, Human* / genetics
  • Herpesvirus 4, Human* / isolation & purification
  • Humans
  • Immune Checkpoint Inhibitors* / immunology
  • Immune Checkpoint Inhibitors* / therapeutic use
  • Immunohistochemistry
  • Lymphocytes, Tumor-Infiltrating / immunology
  • Male
  • Middle Aged
  • Prognosis
  • Programmed Cell Death 1 Receptor / genetics*
  • Survival Analysis
  • Tumor Microenvironment / immunology*
  • Whole Exome Sequencing / methods


  • B7-H1 Antigen
  • CD274 protein, human
  • Immune Checkpoint Inhibitors
  • PDCD1 protein, human
  • Programmed Cell Death 1 Receptor