Emergence of Intrahepatic Cholangiocarcinoma: How High-Throughput Technologies Expedite the Solutions for a Rare Cancer Type

Abstract

Intrahepatic cholangiocarcinoma (ICC) is the cancer of the intrahepatic bile ducts, and together with hepatocellular carcinoma (HCC), constitute the majority of primary liver cancers. ICC is a rare disorder as its overall incidence is < 1/100,000 in the United States and Europe. However, it shows much higher incidence in particular geographical regions, such as northeastern Thailand, where liver fluke infection is the most common risk factor of ICC. Since the early stages of ICC are often asymptomatic, the patients are usually diagnosed at advanced stages with no effective treatments available, leading to the high mortality rate. In addition, unclear genetic mechanisms, heterogeneous nature, and various etiologies complicate the development of new efficient treatments. Recently, a number of studies have employed high-throughput approaches, including next-generation sequencing and mass spectrometry, in order to understand ICC in different biological aspects. In general, the majority of recurrent genetic alterations identified in ICC are enriched in known tumor suppressor genes and oncogenes, such as mutations in TP53, KRAS, BAP1, ARID1A, IDH1, IDH2, and novel FGFR2 fusion genes. Yet, there are no major driver genes with immediate clinical solutions characterized. Interestingly, recent studies utilized multi-omics data to classify ICC into two main subgroups, one with immune response genes as the main driving factor, while another is enriched with driver mutations in the genes associated with epigenetic regulations, such as IDH1 and IDH2. The two subgroups also show different hypermethylation patterns in the promoter regions. Additionally, the immune response induced by host-pathogen interactions, i.e., liver fluke infection, may further stimulate tumor growth through alterations of the tumor microenvironment. For in-depth functional studies, although many ICC cell lines have been globally established, these homogeneous cell lines may not fully explain the highly heterogeneous genetic contents of this disorder. Therefore, the advent of patient-derived xenograft and 3D patient-derived organoids as new disease models together with the understanding of evolution and genetic alterations of tumor cells at the single-cell resolution will likely become the main focus to fill the current translational research gaps of ICC in the future.

Keywords: disease model; high-throughput technology; integrative multi-omics analysis; intrahepatic cholangiocarcinoma; molecular biomarker; precision oncology; translational medicine.

Figure 1

Overview of the anatomical structures, macroscopic subtypes, and recurrent genetic alterations in ICCs. Left panel; an illustration showing the anatomical structures of biliary system and their associated malignancies. HCC, hepatocellular carcinoma; ICC, intrahepatic cholangiocarcinoma; PCC, perihilar cholangiocarcinoma; GB, gallbladder cancer; DCC, distal cholangiocarcinoma; PDAC, pancreatic ductal adenocarcinoma. Middle panel; an illustration showing the three macroscopic subtypes of ICC. MF, mass-forming type; PDI, periductal-infiltrating type; IDG, intraductal growth type. Right panel; a summary of recurrent genetic alterations and their reported frequencies in ICCs. ^aThe mutation frequency of each gene is calculated by dividing the combined number of ICC cases presenting the mutation with the total number of ICC cases analyzed in all four cohorts included in the cBioPortal for Cancer Genomics database (www.cbioportal.org). ^bThe frequency of each fusion gene were obtained from previous literatures (Nakamura et al., ; Moeini et al., 2016). ^cDifferent hypermethylation patterns of liver fluke-associated and non-liver fluke-associated ICCs and their associated alterations were summarized based on a previous study (Jusakul et al., 2017).

Figure 2

Current disease models for studying ICC. (A) ICC cancer cell lines. There are many cell lines established from primary tumor cells. Three representative cell lines are listed. HuCCA-1 was derived from a Thai patient with liver fluke infection. MT-CHCO1 and KKU-213L5 were both established from patient-derived xenografts (PDX). (B) 3D patient-derived tissue-like organoids. Organoids preserve the properties of primary tumor cells as well as tissue heterogeneity. (C) Genetically engineered mouse model (GEMM). A GEMM of ICC was generated by inducing oncogenic KRAS mutation and homozygous PTEN deletion in mouse liver. (D) Orthotopic patient-derived xenograft (PDX). In orthotopic PDX mouse models, patient-derived tumor cells are transplanted into the same organ from which the patient's cancer originated, followed by stabilizing the tumors in the animals. (E) A mouse model of ICC created by CRISPR/Cas9 gene editing. CRISPR/Cas9 is used to introduce mutations to the selected tumor suppressor genes including Arid1a, Trp53, Tet2, Pten, Cdkn2a, Apc, Brca1/2, and Smad4, which lead to ICC in the gene-edited mice.

Figure 3

A schematic diagram proposing the application of precision oncology in ICC through the use of high-throughput technologies and disease models. By applying high-throughput technologies on large numbers of patient samples, different levels of omics data can be obtained and provide information of the molecular changes in the tumor cells or microenvironments (Left panel). Aberrant alterations identified from omics data can then be functionally validated in disease models. Organoids and patient-derived xenograft (PDX) mouse are new disease models (Right panel). The two “next-generation” tumor avatars provide “patient-like” models for integrative multi-omics analyses to study the underlying mechanisms of disorders. The avatars can be used for the following studies: single cell sequencing for understanding clonal evolution and heterogeneity of tumors, disease models for gene editing, tumor microenvironments, and high-throughput systematic drug screening and testing. They can further be biobanked for future studies (Far right panel).