Transplantation of Zebrafish Pediatric Brain Tumors into Immune-competent Hosts for Long-term Study of Tumor Cell Behavior and Drug Response

Abstract

Tumor cell transplantation is an important technique to define the mechanisms controlling cancer cell growth, migration, and host response, as well as to assess potential patient response to therapy. Current methods largely depend on using syngeneic or immune-compromised animals to avoid rejection of the tumor graft. Such methods require the use of specific genetic strains that often prevent the analysis of immune-tumor cell interactions and/or are limited to specific genetic backgrounds. An alternative method in zebrafish takes advantage of an incompletely developed immune system in the embryonic brain before 3 days, where tumor cells are transplanted for use in short-term assays (i.e., 3 to 10 days). However, these methods cause host lethality, which prevents the long-term study of tumor cell behavior and drug response. This protocol describes a simple and efficient method for the long-term orthotopic transplantation of zebrafish brain tumor tissue into the fourth ventricle of a 2-day-old immune-competent zebrafish. This method allows: 1) long-term study of tumor cell behaviors, such as invasion and dissemination; 2) durable tumor response to drugs; and 3) re-transplantation of tumors for the study of tumor evolution and/or the impact of different host genetic backgrounds. In summary, this technique allows cancer researchers to assess engraftment, invasion, and growth at distant sites, as well as to perform chemical screens and cell competition assays over many months. This protocol can be extended to studies of other tumor types and can be used to elucidate mechanisms of chemoresistance and metastasis.

Astrocytes influence medulloblastoma phenotypes and CD133 surface expression

“…Recent methodologies have established zebrafish as an effective vertebrate system to visualize xenografted tumor cell behavior and movement [ 21 ]. Specifically, pediatric brain tumors have been successfully xenografted in zebrafish for drug screening purposes [ 22 , 23 ]. A zebrafish xenotransplant model combined with confocal time-lapse imaging were used to determine the behavior of astrocyte conditioned MB cells in vivo .…”

Modeling Cancer Using Zebrafish Xenografts: Drawbacks for Mimicking the Human Microenvironment

“…Furthermore, zebrafish orthotopic brain xenografts offer the notable advantage of performing high-throughput drug screenings and, at the same time, studying the ability of potential drugs to penetrate the blood–brain barrier [ 133 , 134 , 135 ]. Alternative strategies have also been developed to allow for long-term study of tumor cell behavior and drug response by the transplantation of zebrafish-derived tumors into immune-competent hosts [ 136 ]. For example, Casey et al described an orthotopic implantation method where pediatric zebrafish brain tumors were injected into 2-day-old zebrafish host embryos to study tumor cell behavior and drug response over many months [ 136 ].…”

“…Alternative strategies have also been developed to allow for long-term study of tumor cell behavior and drug response by the transplantation of zebrafish-derived tumors into immune-competent hosts [ 136 ]. For example, Casey et al described an orthotopic implantation method where pediatric zebrafish brain tumors were injected into 2-day-old zebrafish host embryos to study tumor cell behavior and drug response over many months [ 136 ]. This alternate syngeneic approach could be applied and extended to other zebrafish tumor types.…”

Preclinical Models of Pediatric Brain Tumors—Forging Ahead

“…An ideal alternative to mice may be zebrafish (Table 1). The cancer genomes between zebrafish and humans are highly conserved [79]. Patient tumors can be transplanted to form PDXs.…”