HBV-related hepatocarcinogenesis: the role of signalling pathways and innovative ex vivo research models

Abstract

Background: Hepatitis B virus (HBV) is the leading cause of liver cancer, but the mechanisms by which HBV causes liver cancer are poorly understood and chemotherapeutic strategies to cure liver cancer are not available. A better understanding of how HBV requisitions cellular components in the liver will identify novel therapeutic targets for HBV associated hepatocellular carcinoma (HCC).

Main body: The development of HCC involves deregulation in several cellular signalling pathways including Wnt/FZD/β-catenin, PI3K/Akt/mTOR, IRS1/IGF, and Ras/Raf/MAPK. HBV is known to dysregulate several hepatocyte pathways and cell cycle regulation resulting in HCC development. A number of these HBV induced changes are also mediated through the Wnt/FZD/β-catenin pathway. The lack of a suitable human liver model for the study of HBV has hampered research into understanding pathogenesis of HBV. Primary human hepatocytes provide one option; however, these cells are prone to losing their hepatic functionality and their ability to support HBV replication. Another approach involves induced-pluripotent stem (iPS) cell-derived hepatocytes. However, iPS technology relies on retroviruses or lentiviruses for effective gene delivery and pose the risk of activating a range of oncogenes. Liver organoids developed from patient-derived liver tissues provide a significant advance in HCC research. Liver organoids retain the characteristics of their original tissue, undergo unlimited expansion, can be differentiated into mature hepatocytes and are susceptible to natural infection with HBV.

Conclusion: By utilizing new ex vivo techniques like liver organoids it will become possible to develop improved and personalized therapeutic approaches that will improve HCC outcomes and potentially lead to a cure for HBV.

Keywords: Cell cycle; Hepatitis B virus; Liver cancer; Organoids; Wnt signalling.

Fig. 1

HBV associated HCC. HBV infection of hepatocytes is thought to impact on a number of cellular signalling pathways to regulate expression and function of genes that control cellular processes but also HBV replication and persistence, that ultimately leads to oncogenic transformation

Fig. 2

Wnt/β-catenin signal transduction pathway. Wnt binding to the FZD/LRP5/6 receptor complex leads to inhibition of GSK3 enzyme activity and the β-catenin destruction complex, which allows newly synthesized β-catenin to accumulate and translocate to the nucleus (orange arrows), where it binds with co-factors to form a transcriptionally active complex. Wnt signalling can be inhibited at the cell surface by various naturally occurring pathway inhibitors such as sFRPs and DKK, which bind to Wnt and LRP5/6 respectively. [secreted Frizzled Related Protein (sFRP); Frizzled (FZD); Dickkopf (DKK); Glycogen Synthase Kinase 3 (GSK3); Adenomatous Polyposis Coli (APC), Casein Kinase 1 (CK1)]

Fig. 3

Targeted phosphorylation’s are required for ubiquitylation and degradation of β-catenin. Axin acts as scaffold that brings CK1, GSK3 and β-catenin in close proximity. CK1 initiates the process by phosphorylating Ser45 at the N-terminus of β-catenin, followed by sequential phosphorylation at Thr41, Ser37 and Ser33 by GSK3. β-Trcp recognizes the phosphorylated residues Ser33 and Ser37, targeting β-catenin for ubiquitylation and proteasomal degradation. These regulatory phosphorylation sites are commonly mutated in liver cancer. [Casein Kinase 1 (CK1); Glycogen Synthase Kinase 3 (GSK3); β-transducin-repeat-containing protein (β-Trcp)]

Fig. 4

In vitro models for studying HBV infection. Primary human hepatocytes derived from liver tissue provide the best material for HBV studies; however, human liver tissue is not readily available and is expensive to source and process. However, the discovery of human NTCP as one of the membrane receptors for HBV binding has allowed for the development of immortalized cell lines susceptible to HBV infection. iPS technology has helped to create better models that resemble functional mature hepatocytes and yield better HBV infection. But these two in vitro models still have several limitations, especially in regard to the genetic and epigenetic profiles of cells arising from different individual sources. Recently, a newly developed technique allows for the production of liver organoids directly from hepatic stem cells in liver tissue, creating a superior model for future HBV studies

Fig. 5

Animal models for studying HBV infection. Primates are the best models for studying HBV infection, but the associated high cost and regulations with animal ethics present significant limitations in the use of primates for future research. Alternative models using treeshrew, woodchuck, duck and mouse are useful, but these models are limited in progressing studies on host-pathogen interactions, immune response and viral clearance in humans. New potential models using transgenic macaques or pigs expressing human NTCP may help bridge this gap