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Cell-active dual specificity phosphatase inhibitors identified by high-content screening
Andreas Vogt1, Kathleen A Cooley, Marni Brisson
1Department of Pharmacology, University of Pittsburgh, Pittsburgh, PA 15261, USA.
Chemistry & Biology
|September 5, 2003
Summary
Researchers identified compounds that inhibit mitogen-activated protein kinase phosphatase-3 (MKP-3), a key enzyme controlling extracellular signal-regulated kinase (Erk) phosphorylation. This discovery offers a new approach for developing dual specificity phosphatase (DSPase) inhibitors.
Area of Science:
- Biochemistry
- Molecular Biology
- Pharmacology
Background:
- Extracellular signal-regulated kinase (Erk) phosphorylation is regulated by dual specificity phosphatases (DSPases).
- Identifying inhibitors for Erk dephosphorylation is crucial for understanding cellular signaling pathways.
Purpose of the Study:
- To discover novel inhibitors of DSPases, specifically targeting Erk dephosphorylation.
- To evaluate the efficacy of identified compounds against mitogen-activated protein kinase phosphatase-3 (MKP-3).
Main Methods:
- Utilized a high-content, fluorescence-based cellular assay to screen the National Cancer Institute's 1990 agent Diversity Set.
- Employed an in vitro assay to assess the inhibitory activity of compounds against MKP-3, VHR, and PTP1B.
- Developed a novel fluorescence-based multiparameter chemical complementation assay for MKP-3 inhibition.
Main Results:
- Identified ten compounds (0.5%) that increased phospho-Erk cytonuclear differences in intact cells.
- Three compounds inhibited MKP-3 in vitro, with no significant effect on VHR or PTP1B.
- The most potent MKP-3 inhibitor exhibited an IC(50) of <10 microM.
Conclusions:
- The phospho-Erk nuclear accumulation assay is a valuable tool for discovering DSPase inhibitors.
- The identified compounds represent potential leads for developing novel therapeutics targeting DSPase activity.