PUBLICATION
In vivo Drug Screening to Identify Anti-metastatic Drugs in Twist1a-ERT2 Transgenic Zebrafish
- Authors
- Nakayama, J., Makinoshima, H., Gong, Z.
- ID
- ZDB-PUB-230531-29
- Date
- 2023
- Source
- Bio-protocol 13: e4673e4673 (Journal)
- Registered Authors
- Gong, Zhiyuan
- Keywords
- EMT, In vivo drug screen, Metastasis, Twist1, Zebrafish
- MeSH Terms
- none
- PubMed
- 37251091 Full text @ Bio Protoc
Citation
Nakayama, J., Makinoshima, H., Gong, Z. (2023) In vivo Drug Screening to Identify Anti-metastatic Drugs in Twist1a-ERT2 Transgenic Zebrafish. Bio-protocol. 13:e4673e4673.
Abstract
Here, we present an in vivo drug screening protocol using a zebrafish model of metastasis for the identification of anti-metastatic drugs. A tamoxifen-controllable Twist1a-ERT2 transgenic zebrafish line was established to serve as a platform for the identification. By crossing Twist1a-ERT2 with xmrk (a homolog of hyperactive form of the epidermal growth factor receptor) transgenic zebrafish, which develop hepatocellular carcinoma, approximately 80% of the double transgenic zebrafish show spontaneous cell dissemination of mCherry-labeled hepatocytes from the liver to the entire abdomen and tail regions in five days, through induction of epithelial to mesenchymal transition (EMT). This rapid and high-frequency induction of cell dissemination makes it possible to perform an in vivo drug screen for the identification of anti-metastatic drugs targeting metastatic dissemination of cancer cells. The protocol evaluates the suppressor effect of a test drug on metastasis in five days, by comparing the frequencies of the fish showing abdominal and distant dissemination patterns in the test drug-treated group with those in the vehicle-treated group. Our study previously identified that adrenosterone, an inhibitor for hydroxysteroid (11-beta) dehydrogenase 1 (HSD11β1), has a suppressor effect on cell dissemination in the model. Furthermore, we validated that a pharmacologic and genetic inhibition of HSD11β1 suppressed metastatic dissemination of highly metastatic human cell lines in a zebrafish xenotransplantation model. Taken together, this protocol opens new routes for the identification of anti-metastatic drugs. Graphical overview Timing Day 0: Zebrafish spawning Day 8: Primary tumor induction Day 11: Chemical treatment Day 11.5: Metastatic dissemination induction in the presence of a test chemical Day 16: Data analysis.
Genes / Markers
Expression
Phenotype
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Fish
Orthology
Engineered Foreign Genes
Mapping