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Liu et al. Hepatoma Res 2020;6:7 I http://dx.doi.org/10.20517/2394-5079.2019.39 Page 5 of 16
[35]
genetically applicable to human disease . Patient-derived xenograft (PDX) models in which cells from
[36]
a specific patient with HCC are transplanted into immunocompromised mice have been established .
PDXs faithfully recapitulate histologic, genomic and biological characteristics of the primary tumour and
have been shown to predict drug response in HCC patients. However, this model is limited by engraftment
failure rates of up to 60%, long time to engraftment (several months) and high cost, which make it
unsuitable for large-scale drug screening [36,37] . Furthermore, the major drawback of xenograft models (PDX
or otherwise) is the lack of a tumoural immune response, which has become increasingly important as we
enter the era of immunotherapies for HCC. Attempts at overcoming this with double humanised mouse
models which express human hepatocytes and haematopoietic stem cells (and hence human immune
[21]
cells) are technically intensive, expensive and not yet widely adopted . Finally, xenograft of human HCC
models which develop metastases (more readily than GEM models) have been established, providing the
[38]
opportunity to study late-stage disease .
Replicability in human disease
Human HCCs are highly complex and heterogeneous and thus cannot be adequately represented by any
single mouse model. For example, gene expression profiles of tumours from the commonly-used DEN
model was previously shown to be most similar to a subgroup of human HCC with poorer survival [39,40] .
Correspondingly, many poor prognostic markers in human HCC are also highly expressed in DEN-induced
tumours, e.g., alpha-foetoprotein (AFP). However, the DEN model lacks other hallmarks of human HCC,
particularly fibrosis in the surrounding microenvironment . In a more recent integrative genomic analysis
[41]
of four separate mouse models and 987 human HCC samples, DEN tumours were found to be histologically
hard to classify and least similar to human disease while Stelic Animal Model (STAM) tumours (discussed
further in the NAFLD-associated HCC models section) were most molecularly similar to human HCC,
[34]
especially high-grade, proliferative tumours with poor prognosis . The authors further argued that DEN
models should be avoided since they are dominated by mutational mechanisms not seen in human HCC.
In contrast to DEN-induced and STAM tumours, MDR2 knockout tumours are most similar to human
[42]
HCCs associated with better survival . However, the MDR2 knockout model produces a phenotype
resembling humans with primary sclerosing cholangitis or primary biliary cholangitis rather than chronic
[43]
“hepatitis” diseases caused by alcohol excess and HBV or HCV infection . Very recently, experimental
hepatocyte-specific activation of β-catenin also resulted the development of a phenotype that resembled
[32]
the low proliferative subclass of human HCC . Interestingly these tumours also had few intratumoural
immune cells and were resistant to immune checkpoint inhibitor therapy. Therefore, it is clear that different
models (and their combinations) are required to simulate specific subgroups of human HCC. Furthermore,
drugs with known anti-tumour activity against human HCC do not demonstrate activity in some animal
models and vice versa. Indeed, the current Food and Drug Administration-approved drugs for treating
advanced HCC such as sorafenib and anti-programmed cell death receptor 1 (PD1) antibodies were trialled
based on success in other cancers (advanced renal cell carcinoma and melanoma, respectively) rather than
positive results in HCC animal models per se.

SPECIFIC MOUSE MODELS FOR ALCOHOLIC AND METABOLIC LIVER DISEASE-

ASSOCIATED HCC
ALD-associated HCC models
In general, mice and other species (except the golden hamster) dislike alcohol and avoid ingestion when it
is offered ad libitum [21,44] . Therefore, ALD mouse models are established by one of three ways: (1) replacing
the food and water source with a liquid diet in which 5% ethanol accounts for 36% of total calories (Lieber-
DeCarli model); (2) binge feeding mice with ethanol via gavage in addition to chronic ingestion [National
Institute of Alcohol Abuse and Alcoholism (NIAAA) model]; or (3) intragastric ethanol infusion via a
surgically inserted infusion pump (Tsukamoto-French model) [Table 1].