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A High-Throughput Method to Identify Novel Senescence-Inducing Compounds

Department of Urology, University of Wisconsin-Madison, Madison, Wisconsin, Paul P. Carbone Comprehensive Cancer Center, University of Wisconsin-Madison, Madison, Wisconsin

Noel Peters

Paul P. Carbone Comprehensive Cancer Center, University of Wisconsin-Madison, Madison, Wisconsin

Joshua A. Desotelle

Department of Environmental Toxicology, University of Wisconsin-Madison, Madison, Wisconsin

F. Michael Hoffmann

Paul P. Carbone Comprehensive Cancer Center, University of Wisconsin-Madison, Madison, Wisconsin, McArdle Laboratory for Cancer Research, University of Wisconsin-Madison, Madison, Wisconsin

David F. Jarrard

Department of Urology, University of Wisconsin-Madison, Madison, Wisconsin, jarrard{at}surgery.wisc.edu, Paul P. Carbone Comprehensive Cancer Center, University of Wisconsin-Madison, Madison, Wisconsin, Department of Environmental Toxicology, University of Wisconsin-Madison, Madison, Wisconsin

Cellular senescence is a persistently growth-arrested phenotype in normal and transformed cells induced by noncytotoxic stress. Cytostasis as a method of cancer treatment has recently generated significant interest. Research into the induction of cellular senescence as cancer therapy has been hindered by a lack of compounds that efficiently induce this response. The authors describe a semiautomated high-throughput method to identify library compounds that induce senescence using prostate cancer cells cultured in 96-well plates. Primary hits are identified by low cell numbers after 3 days in culture, measured by Hoechst 33342 fluorescence. A secondary visual assessment of senescence-associated β-galactosidase staining and cellular morphology in the same wells distinguishes senescence from quiescence, apoptosis, and other false positives. This method was used to screen a 4160-compound library of known bioactive compounds and natural products at a 10-µM dose. Candidate compounds were further selected based on persistent growth arrest after drug removal and increased expression of previously described senescence marker genes. Four lead compounds not previously associated with senescence were identified for further investigation. This is the first successful assay to identify novel agents from compound libraries based on senescence induction in cancer cells. (Journal of Biomolecular Screening 2009:853-858)

Key Words: cellular senescence • high-throughput screen • prostate • prostate cancer

This version was published on August 1, 2009

Journal of Biomolecular Screening, Vol. 14, No. 7, 853-858 (2009)
DOI: 10.1177/1087057109340314


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