A Novel Cell Line Based Orthotopic Xenograft Mouse Model That Recapitulates Human Hepatoblastoma

Sarah E Woodfield,Pavel Sumazin,Zbigniew Starosolski,Karl-Dimiter Bissig,Siddharth S Gupta,Sanjeev A Vasudevan,Dolores López-Terrada,Zhenghu Chen,Stephen F Sarabia,Barry Zorman,Jingling Jin,Ketan B Ghaghada,Yan Shi,Angela Major,Aryana M Ibarra,Roma H Patel

doi:10.1038/s41598-017-17665-8

Sarah E Woodfield, Pavel Sumazin + Show 14 more

Open Access

https://doi.org/10.1038/s41598-017-17665-8

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Abstract

Currently, preclinical testing of therapies for hepatoblastoma (HB) is limited to subcutaneous and intrasplenic xenograft models that do not recapitulate the hepatic tumors seen in patients. We hypothesized that injection of HB cell lines into the livers of mice would result in liver tumors that resemble their clinical counterparts. HepG2 and Huh-6 HB cell lines were injected, and tumor growth was monitored with bioluminescence imaging (BLI) and magnetic resonance imaging (MRI). Levels of human α-fetoprotein (AFP) were monitored in the serum of animals. Immunohistochemical and gene expression analyses were also completed on xenograft tumor samples. BLI signal indicative of tumor growth was seen in 55% of HepG2- and Huh-6-injected animals after a period of four to seven weeks. Increased AFP levels correlated with tumor growth. MRI showed large intrahepatic tumors with active neovascularization. HepG2 and Huh-6 xenografts showed expression of β-catenin, AFP, and Glypican-3 (GPC3). HepG2 samples displayed a consistent gene expression profile most similar to human HB tumors. Intrahepatic injection of HB cell lines leads to liver tumors in mice with growth patterns and biologic, histologic, and genetic features similar to human HB tumors. This orthotopic xenograft mouse model will enable clinically relevant testing of novel agents for HB.

Highlights

To date, in vivo HB research includes studies with hydrodynamic injection of oncogenes for liver specific expression[7], as well as subcutaneous and intrasplenic murine xenograft models[8,9,10]
We injected two million HepG2 or Huh-6 human HB cell lines into either the right median lobe with a right flank incision (Fig. 1a) or the left lateral lobe with a midline, abdominal incision (Fig. 1b) of the liver of NOD/Shi-scid/ IL-2Rγnull (NOG) mice
Regardless of injection technique, the tumors grew as large, exophytic masses originating from the injected lobe and invaded multiple adjacent segments of the liver as seen in children with pre-treatment extent of disease (PRETEXT) I, II, and III tumors (Fig. 1c,d) 17

Summary

Introduction

In vivo HB research includes studies with hydrodynamic injection of oncogenes for liver specific expression[7], as well as subcutaneous and intrasplenic murine xenograft models[8,9,10]. Other groups have examined subcutaneous and intrahepatic growth of patient-derived liver cancer tissues as models of HCC, including an interesting study in which tumors composed of sorted human liver cancer stem cells (hLCSCs) were grown subcutaneously[15,16]. Since these tissues have limited availability due to the rarity of the disease, we wanted to develop and characterize an intrahepatic, orthotopic xenograft model using commercially available HB cell lines. At the conclusion of the study, animals were euthanized and tissues were harvested for protein expression analysis by immunohistochemistry and gene expression profiling by RNA sequencing

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