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SA-β-Galactosidase-Based Screening Assay for the Identification of Senotherapeutic Drugs
Published on: June 28, 2019
Cell-based screen for altered nuclear phenotypes reveals senescence progression in polyploid cells after Aurora
Mahito Sadaie1, Christian Dillon2, Masako Narita
1Cancer Research UK Cambridge Institute, University of Cambridge, Cambridge CB2 0RE, United Kingdom.
Abstract:
Cellular senescence is a widespread stress response and is widely considered to be an alternative cancer therapeutic goal. Unlike apoptosis, senescence is composed of a diverse set of subphenotypes, depending on which of its associated effector programs are engaged. Here we establish a simple and sensitive cell-based prosenescence screen with detailed validation assays. We characterize the screen using a focused tool compound kinase inhibitor library. We identify a series of compounds that induce different types of senescence, including a unique phenotype associated with irregularly shaped nuclei and the progressive accumulation of G1 tetraploidy in human diploid fibroblasts. Downstream analyses show that all of the compounds that induce tetraploid senescence inhibit Aurora kinase B (AURKB). AURKB is the catalytic component of the chromosome passenger complex, which is involved in correct chromosome alignment and segregation, the spindle assembly checkpoint, and cytokinesis. Although aberrant mitosis and senescence have been linked, a specific characterization of AURKB in the context of senescence is still required. This proof-of-principle study suggests that our protocol is capable of amplifying tetraploid senescence, which can be observed in only a small population of oncogenic RAS-induced senescence, and provides additional justification for AURKB as a cancer therapeutic target.
Insights
Researchers developed a new screening method to identify compounds that induce cellular senescence, a key cancer therapeutic target. This method identified inhibitors of Aurora kinase B (AURKB) that cause a unique tetraploid senescence phenotype.
Area of Science:
- Cell Biology
- Cancer Therapeutics
- Molecular Biology
Background:
- Cellular senescence is a critical stress response and a promising cancer therapeutic strategy.
- Senescence exhibits diverse subphenotypes, unlike apoptosis, influenced by specific effector programs.
- Identifying and characterizing these subphenotypes is crucial for targeted cancer therapies.
Purpose of the Study:
- To establish a sensitive cell-based screening assay for identifying pro-senescence compounds.
- To characterize novel senescence subphenotypes induced by kinase inhibitors.
- To investigate the role of Aurora kinase B (AURKB) in tetraploid senescence and its potential as a cancer therapeutic target.
Main Methods:
- Development and validation of a high-throughput cell-based pro-senescence screen.
- Utilized a focused library of tool compound kinase inhibitors for screening.
- Characterized senescence phenotypes using microscopy and flow cytometry, focusing on nuclear morphology and DNA content.
- Investigated the mechanism of tetraploid senescence induction, including AURKB inhibition.
Main Results:
- Identified several compounds that induce distinct senescence subphenotypes in human diploid fibroblasts.
- Discovered a unique tetraploid senescence phenotype characterized by irregular nuclei and G1 tetraploidy.
- Demonstrated that compounds inducing tetraploid senescence specifically inhibit Aurora kinase B (AURKB).
- Showcased the screen's ability to amplify rare tetraploid senescence phenotypes.
Conclusions:
- The developed screening protocol effectively identifies compounds inducing diverse senescence phenotypes, including tetraploid senescence.
- Aurora kinase B (AURKB) inhibition is a key driver of the identified tetraploid senescence.
- This study provides further evidence supporting AURKB as a viable target for cancer therapy.
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