Orthotopic models of pediatric brain tumors in zebrafish

Abstract

High-throughput screens (HTS) of compound toxicity against cancer cells can identify thousands of potential new drug-leads. But only limited numbers of these compounds can progress to expensive and labor intensive efficacy studies in mice, creating a ‘bottle-neck’ in the drug development pipeline. Approaches that triage drug-leads for further study are greatly needed. Here, we provide an intermediary platform between HTS and mice by adapting mouse models of pediatric brain tumors to grow as orthotopic xenografts in the brains of zebrafish. Freshly isolated mouse ependymoma, glioma and choroid plexus carcinoma cells expressing red fluorescence protein (RFP) were conditioned to grow at 34°C. Conditioned tumor cells were then transplanted orthotopically into the brains of zebrafish acclimatized to ambient temperatures of 34°C. Live in vivo fluorescence imaging identified robust, quantifiable and reproducible brain tumor growth as well as spinal metastasis in zebrafish. All tumor xenografts in zebrafish retained the histological characteristics of the corresponding parent mouse tumor and efficiently recruited fish endothelial cells to form a tumor vasculature. Finally, by treating zebrafish harboring ERBB2-driven gliomas with an appropriate cytotoxic chemotherapy (5-fluorouracil) or tyrosine kinase inhibitor (Erlotinib), we show that these models can effectively assess drug efficacy. Our data demonstrate, for the first time, that mouse brain tumors can grow orthtopically in fish and serve as a platform to study drug efficacy. Since large cohorts of brain tumor bearing zebrafish can be generated rapidly and inexpensively, these models may serve as a powerful tool to triage drug-leads from HTS for formal efficacy testing in mice.

The Study of Glioma by Xenotransplantation in Zebrafish Early Life Stages

“…Gene expression patterns of cancer cells were shown to be highly similar in zebrafish and humans (Lam et al 2006;Zheng et al 2014). Similar molecular pathways may lead to tumor development in the two species (Jung et al 2013;Mione and Trede 2010) and tumors in zebrafish were found to be histologically similar to their mammalian counterparts (Armatruda et al 2002;Eden et al 2014;Stern and Zon 2003). Furthermore, zebrafish cells can respond to human signaling molecules (Drabsch et al 2013).…”

“…The absence of pigmentation aids microscopic observations because pigments in the embryo may conceal the fluorescence emitted by fluorescently labeled cells and only non-pigmented fish enable quantitative intravital fluorescence imaging of implanted cells. Despite its benefits, the Casper strain has rarely been used in xenotransplantation studies (Corkery et al 2011;Eden et al 2014). Instead, low concentrations of phenylthiourea (PTU) are often used to inhibit melanin synthesis (Lee et al 2009;Yang et al 2013aYang et al , 2013bYang et al , 2014.…”

“…The assessment of the effects of different treatments on the proliferation of cancer cells through fluorescence quantification has been successfully implemented (Eden et al 2014;Geiger et al 2008;Lally et al 2007;Yang et al 2014). An alternative approach is the use of bioluminescence imaging of implanted cells genetically modified to express luciferase (Zhao et al 2009).…”

“…An alternative to the fluorescence-labeling approach is the establishment of cancer cell lines that stably express fluorescent proteins (Drabsch et al 2012;Eden et al 2014;Geiger et al 2008;He et al 2012;Yang et al 2013a). The fluorescence of endogenous fluorescent proteins is more stable and correlates with cell number, enabling the quantification of proliferation by measuring fluorescence intensity (Geiger et al 2008;Vittori et al 2014;Yang et al 2014).…”

“…Human GBM cells have been successfully implanted into the embryonic brain at two days after fertilization (Kitambi et al 2014;Vittori et al 2014). However, most orthotopic studies have used older developmental stages, such as larvae and juveniles (Eden et al 2014;Lal et al 2012;Rampazzo et al 2013). Lal et al (2012) assessed the invasiveness of GBM cells implanted into the brain of either 4-day-old embryos or 10-day-old larval zebrafish.…”