Identification of tubulin polymerization inhibitors with a CRISPR-edited cell line with endogenous fluorescent tagging of β-tubulin and Histone 1
GroupMoulder Center for Drug Discovery (Temple University)
Permanent link to this recordhttp://hdl.handle.net/20.500.12613/8250
MetadataShow full item record
AbstractTubulin is an essential protein to maintain the cellular structure and for the cell division process. Inhibiting tubulin polymerization has proven to be an effective method for slowing cancer cell growth. Traditionally, identifying tubulin polymerization inhibitors involved using pure tubulin for in vitro assays or procedures using cells that require cell fixing and anti-tubulin antibody staining. This study explores using a cell line developed via CRISPR genome editing as a cell model to identify tubulin polymerization inhibitors with live cells without using exogenous staining. The cell line has endogenous tagging with fluorescent proteins of β-tubulin and a nuclear protein to facilitate image cellular segmentation by high-content imaging analysis (HCI). The cells were treated with known tubulin polymerization inhibitors, colchicine and vincristine, and the presence of phenotypic changes that indicate tubulin polymerization inhibition were confirmed via HCI. A library of 429 kinase inhibitors was screened to discover tubulin polymerization inhibitors and three compounds that inhibit tubulin polymerization were found (ON-01910, HMN-214, and KX2-391). Live cell tracking analysis confirms that depolymerization of tubulin occurs rapidly after compound treatments. These results suggest that CRISPR-edited cells with fluorescent endogenous tagging of β-tubulin can be used to screen larger compound libraries containing diverse chemical families to identify novel tubulin polymerization inhibitors.
Citation to related workbioRxiv
ADA complianceFor Americans with Disabilities Act (ADA) accommodation, including help with reading this content, please contact email@example.com
Except where otherwise noted, this item's license is described as Attribution-NonCommercial-NoDerivs CC BY-NC-ND