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Identification of Polo-like kinase 1 interaction inhibitors using a novel cell-based assay
Karine Normandin1, Jean-François Lavallée1, Marie Futter1
1Institute for Research in Immunology and Cancer, Université de Montréal, Montréal, Canada.
Abstract:
Polo-like kinase 1 (Plk1) plays several roles in cell division and it is a recognized cancer drug target. Plk1 levels are elevated in cancer and several types of cancer cells are hypersensitive to Plk1 inhibition. Small molecule inhibitors of the kinase domain (KD) of Plk1 have been developed. Their selectivity is limited, which likely contributes to their toxicity. Polo-like kinases are characterized by a Polo-Box Domain (PBD), which mediates interactions with phosphorylation substrates or regulators. Inhibition of the PBD could allow better selectivity or result in different effects than inhibition of the KD. In vitro screens have been used to identify PBD inhibitors with mixed results. We developed the first cell-based assay to screen for PBD inhibitors, using Bioluminescence Resonance Energy Transfer (BRET). We screened through 112 983 compounds and characterized hits in secondary biochemical and biological assays. Subsequent Structure-Activity Relationship (SAR) analysis on our most promising hit revealed that it requires an alkylating function for its activity. In addition, we show that the previously reported PBD inhibitors thymoquinone and Poloxin are also alkylating agents. Our cell-based assay is a promising tool for the identification of new PBD inhibitors with more drug-like profiles using larger and more diverse chemical libraries.
Insights
Researchers developed a novel cell-based assay to discover Polo-like kinase 1 (Plk1) Polo-Box Domain (PBD) inhibitors. This assay identified alkylating agents as potential PBD inhibitors, offering a new avenue for targeted cancer therapy.
Area of Science:
- Biochemistry
- Cell Biology
- Drug Discovery
Background:
- Polo-like kinase 1 (Plk1) is crucial for cell division and a validated cancer drug target.
- Existing Plk1 kinase domain inhibitors show limited selectivity and toxicity.
- Targeting the Plk1 Polo-Box Domain (PBD) offers potential for improved selectivity and therapeutic effects.
Purpose of the Study:
- To develop and validate a cell-based assay for identifying Polo-Box Domain (PBD) inhibitors of Plk1.
- To screen a large compound library for novel PBD inhibitors.
- To characterize the mechanism of action of identified PBD inhibitors.
Main Methods:
- Development of a novel cell-based assay utilizing Bioluminescence Resonance Energy Transfer (BRET) for PBD inhibitor screening.
- Screening of 112,983 compounds.
- Secondary biochemical and biological assays for hit characterization.
- Structure-Activity Relationship (SAR) analysis.
Main Results:
- The BRET-based cell assay successfully identified inhibitors of Plk1 PBD activity.
- A promising hit compound was identified, requiring an alkylating function for activity.
- Previously reported PBD inhibitors, thymoquinone and Poloxin, were also identified as alkylating agents.
Conclusions:
- The developed cell-based BRET assay is a powerful tool for discovering novel Plk1 PBD inhibitors.
- Alkylating agents represent a class of compounds with potential for selective Plk1 PBD inhibition.
- This approach may lead to the development of new cancer therapeutics with improved drug-like profiles.
