This post highlights the approach and finding of a new research article published by Disease Models and Mechanisms (DMM). This feature is written by Joseph I. Kaluzny as apart of a seminar at The University of Alabama (taught by DMM Editorial Board member, Prof. Guy Caldwell) on current topics related to use of animal and cellular model systems in studies of human disease.
Fibrolamellar Carcinoma (FLC) is a hepatocellular carcinoma that disproportionately affects young patients and is characterized by a fusion transcript, DNAJB1-PRKACA, which acts as a unique molecular driver and is sufficient for diagnosis (Graham et al., 2015). While liver resection and transplantation remain common management approaches (Kassahun, 2016), the lack of available therapy has motivated molecular mechanistic studies of the fusion.
Previous work has shown that the fusion is sufficient to drive FLC tumorigenesis in murine models (Engelholm et al., 2017). In a recent Disease Models & Mechanisms article, de Oliveira and colleagues chose to study the fusion in zebrafish due to their transparent larvae that provide non-invasive live imaging of liver morphology and inflammatory responses (de Oliveira et al., 2020). The researchers used a hepatocyte-specific promoter to overexpress the fusion and establish an FLC zebrafish line. Liver visualization in adults was achieved via outcrossing with a transgenic line expressing agfp-I10a (Fig. 2A in de Oliveira et al., 2020). The livers of 8- and 12-month-old FLC and control fish were resected for standard histopathological evaluation, which confirmed liver enlargement and abnormal hepatocellular architecture in FLC livers (Fig. 2B-C in de Oliveira et al., 2020).
The investigators then sought to determine if overexpression of the fusion caused alterations indicative of malignancy in larval zebrafish. The researchers confirmed hepatomegaly 7 days post-fertilization, suggesting the potential for zebrafish larvae to be used as a model to study the progression of early FLC, an area of interest for a progressive condition that primarily affects young patients (Fig. 3 in de Oliveira et al., 2020). Overexpression of the fusion increased neutrophil and macrophage infiltration into the liver, TNFα-positive macrophages, and caspase-a activity, confirming an inflammatory response in FLC larvae (de Oliveira et al., 2020). Targeting this inflammation with TNFα and caspase-a inhibitors limited FLC progression.
Despite this potential for therapy, there are many outstanding issues with the zebrafish FLC model, such as the presence of two fusion forms due to the genome duplication in zebrafish, the lack of fibrosis markers characteristic of human FLC progression (Kastenhuber et al., 2017), and alternate pro-inflammatory pathways that are unexplored or understudied in the field (Rigutto et al., 2009), which warrant further study.
de Oliveira, S., Houseright, R. A., Korte, B. G. and Huttenlocher A. (2020). DnaJ-PKAc Fusion Induces Liver Inflammation in a Zebrafish Model of Fibrolamellar Carcinoma. Disease Models & Mechanisms, 26 Feb. 2020, doi:10.1242/dmm.042564.
Engelholm, L. H., Riaz, A., Serra, D., Dagnaes-Hansen, F., Johansen, J. V., Santoni-Rugiu, E., Hansen, S. H., Niola, F. and Frodin, M. (2017). CRISPR/Cas9 Engineering of Adult Mouse Liver Demonstrates That the Dnajb1-Prkaca Gene Fusion Is Sufficient to Induce Tumors Resembling Fibrolamellar Hepatocellular Carcinoma. Gastroenterology 153, 1662-1673 e10.
Graham, R. P., Jin, L., Knutson, D. L., Kloft-Nelson, S. M., Greipp, P. T., Waldburger, N., Roessler, S., Longerich, T., Roberts, L. R., Oliveira, A. M. et al. (2015). DNAJB1-PRKACA is specific for fibrolamellar carcinoma. Mod Pathol 28, 822-9.
Kassahun, W. T. (2016). Contemporary management of fibrolamellar hepatocellular carcinoma: diagnosis, treatment, outcome, prognostic factors, and recent developments. World J Surg Oncol 14, 151.
Kastenhuber, E. R., Lalazar, G., Houlihan, S. L., Tschaharganeh, D. F., Baslan, T., Chen, C. C., Requena, D., Tian, S., Bosbach, B., Wilkinson, J. E. et al. (2017). DNAJB1-PRKACA fusion kinase interacts with beta-catenin and the liver regenerative response to drive fibrolamellar hepatocellular carcinoma. Proc Natl Acad Sci U S A 114, 13076-13084.
Rigutto, S., Hoste, C., Grasberger, H., Milenkovic, M., Communi, D., Dumont, J. E., Corvilain, B., Miot, F. and De Deken, X. (2009). Activation of dual oxidases Duox1 and Duox2: differential regulation mediated by camp-dependent protein kinase and protein kinase C-dependent phosphorylation. J Biol Chem 284, 6725-34.