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Updated: May 3, 2026
Rapid in vivo Drug Response Prediction Using Leukemia Cell Grafts in Zebrafish Embryos
Published on: May 23, 2025
Quantitative phenotyping-based in vivo chemical screening in a zebrafish model of leukemia stem cell
Beibei Zhang1, Yasuhito Shimada2, Junya Kuroyanagi1
1Department of Molecular and Cellular Pharmacology, Pharmacogenomics and Pharmacoinformatics, Mie University Graduate School of Medicine, Edobashi, Tsu, Mie, Japan.
Abstract:
Zebrafish-based chemical screening has recently emerged as a rapid and efficient method to identify important compounds that modulate specific biological processes and to test the therapeutic efficacy in disease models, including cancer. In leukemia, the ablation of leukemia stem cells (LSCs) is necessary to permanently eradicate the leukemia cell population. However, because of the very small number of LSCs in leukemia cell populations, their use in xenotransplantation studies (in vivo) and the difficulties in functionally and pathophysiologically replicating clinical conditions in cell culture experiments (in vitro), the progress of drug discovery for LSC inhibitors has been painfully slow. In this study, we developed a novel phenotype-based in vivo screening method using LSCs xenotransplanted into zebrafish. Aldehyde dehydrogenase-positive (ALDH+) cells were purified from chronic myelogenous leukemia K562 cells tagged with a fluorescent protein (Kusabira-orange) and then implanted in young zebrafish at 48 hours post-fertilization. Twenty-four hours after transplantation, the animals were treated with one of eight different therapeutic agents (imatinib, dasatinib, parthenolide, TDZD-8, arsenic trioxide, niclosamide, salinomycin, and thioridazine). Cancer cell proliferation, and cell migration were determined by high-content imaging. Of the eight compounds that were tested, all except imatinib and dasatinib selectively inhibited ALDH+ cell proliferation in zebrafish. In addition, these anti-LSC agents suppressed tumor cell migration in LSC-xenotransplants. Our approach offers a simple, rapid, and reliable in vivo screening system that facilitates the phenotype-driven discovery of drugs effective in suppressing LSCs.
Insights
This study introduces a novel zebrafish model for leukemia stem cell (LSC) drug discovery. The in vivo screening method successfully identified compounds that inhibit LSC proliferation and migration, offering a faster approach to developing LSC inhibitors.
Area of Science:
- * Oncology
- * Pharmacology
- * Zebrafish models
Background:
- * Leukemia stem cells (LSCs) are crucial for disease eradication but challenging to target due to low numbers and complex in vitro/in vivo models.
- * Current drug discovery for LSC inhibitors is slow, hindering therapeutic progress.
- * Zebrafish offer a rapid and efficient platform for chemical screening and disease modeling.
Purpose of the Study:
- * To develop and validate a novel phenotype-based in vivo screening method for identifying leukemia stem cell (LSC) inhibitors using zebrafish xenografts.
- * To assess the efficacy of eight therapeutic agents in suppressing LSC proliferation and migration in vivo.
- * To establish a rapid and reliable system for phenotype-driven drug discovery targeting LSCs.
Main Methods:
- * Purification of aldehyde dehydrogenase-positive (ALDH+) leukemia K562 cells tagged with Kusabira-orange.
- * Xenotransplantation of ALDH+ cells into young zebrafish (48 hours post-fertilization).
- * In vivo treatment with eight different therapeutic agents and subsequent high-content imaging to assess cancer cell proliferation and migration.
Main Results:
- * Six of the eight tested compounds, excluding imatinib and dasatinib, selectively inhibited ALDH+ cell proliferation in the zebrafish model.
- * The identified anti-LSC agents effectively suppressed tumor cell migration in LSC-xenotransplants.
- * The developed zebrafish screening system demonstrated reliability and efficiency in identifying LSC-suppressing drugs.
Conclusions:
- * A novel, rapid, and reliable in vivo zebrafish screening system was successfully developed for phenotype-driven LSC drug discovery.
- * The method facilitates the identification of compounds effective in suppressing leukemia stem cells.
- * This approach accelerates the discovery of novel therapeutic agents for leukemia treatment.
