Phenotype-driven chemical screening in zebrafish for collective cell migration inhibitors identifies multiple potential pathways for targeting metastasis

Gallardo, V.E., Varshney, G.K., Lee, M., Bupp, S., Xu, L., Shinn, P., Crawford, N.P., Inglese, J., Burgess, S.M.
Disease models & mechanisms   8(6): 565-76 (Journal)
Registered Authors
Burgess, Shawn, Gallardo, Viviana, Varshney, Gaurav, Xu, Lisha
MeSH Terms
  • Amino Acid Sequence
  • Animals
  • CRISPR-Cas Systems/genetics
  • Cell Line, Tumor
  • Cell Movement*/drug effects
  • Drug Evaluation, Preclinical*
  • Embryo, Nonmammalian/drug effects
  • Female
  • Humans
  • Indoles/pharmacology
  • Lateral Line System/cytology
  • Lateral Line System/metabolism
  • Mice, Inbred BALB C
  • Models, Biological
  • Molecular Sequence Data
  • Neoplasm Metastasis
  • Neoplasms/metabolism
  • Neoplasms/pathology*
  • Phenotype
  • RNA, Messenger/genetics
  • RNA, Messenger/metabolism
  • Reproducibility of Results
  • Signal Transduction*/drug effects
  • Small Molecule Libraries/pharmacology
  • Sulfonamides/pharmacology
  • Zebrafish/embryology
  • Zebrafish/metabolism*
  • Zebrafish Proteins/chemistry
  • Zebrafish Proteins/metabolism
  • src-Family Kinases/antagonists & inhibitors
  • src-Family Kinases/metabolism
25810455 Full text @ Dis. Model. Mech.
In the last decade, high-throughput chemical screening has become the dominant approach for discovering novel compounds with therapeutic properties. Automated screening using in vitro or cultured cell assays have yielded thousands of candidate drugs for a variety of biological targets, but these approaches have not resulted in an increase in drug discovery despite major increases in expenditures. In contrast, phenotype-driven screens have shown a much stronger success rate, so we developed an in vivo assay using transgenic zebrafish with a GFP marked migrating posterior lateral line primordium (PLLp) to identify compounds influencing collective cell migration. We then conducted a high-throughput screen using a compound library of 2,160 annotated bioactive synthetic compounds and 800 natural products to identify molecules that blocked normal PLLp migration. We identified 165 compounds interfering with primordium migration without overt toxicity in vivo. Selected compounds were confirmed in their migration blocking activity using additional assays for cell migration. We then proved the screen could be successful in identifying anti-metastatic compounds active in vivo by performing orthotopic tumor implantation assays in mice. We demonstrated that the Src inhibitor SU6656, identified in our screen, could be used to suppress the metastatic capacity of a highly aggressive mammary tumor cell line. Finally we used CRISPR/Cas9 targeted mutagenesis in zebrafish to genetically validate predicted targets for compounds. This approach demonstrates that the migrating PLLp in zebrafish can be used for large-scale, high-throughput screening for compounds that inhibit collective cell migration and potentially anti-metastatic compounds.
Genes / Markers
Show all Figures
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Engineered Foreign Genes