ZG
Zhiyuan Gong
  • National University of Singapore
In vivo Drug Screening to Identify Anti-metastatic Drugs in Twist1a-ERT2 Transgenic Zebrafish
Twist1a-ERT2转基因斑马鱼中进行体内药物筛选以确定抗转移药物
作者:Joji Nakayama, Hideki Makinoshima and Zhiyuan Gong日期:05/20/2023,浏览量:581,Q&A: 1

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