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Identification of Small-Molecule Regulators of Testicular Receptor 4 via a Drug Repurposing Screening
Liqun Xia1, Danyang Shen1, Huan Wang1
1Department of Urology, Sir Run Run Shaw Hospital, School of Medicine, Zhejiang University, Hangzhou 310016, China.
Abstract:
The testicular receptor 4 (TR4) is a nuclear receptor implicated in multiple pathological processes, including cancer development, chemotherapy, and radiotherapy resistance. However, no effective TR4 small-molecule regulator is available to date. Here, we assessed a physical-interaction-based surface plasmon resonance imaging assay for discovery of TR4 regulators. We screened 1018 FDA-approved drugs and obtained 126 drugs with K D values below 10-6 M. The dual-luciferase-based biological assay verified four activatory compounds and two inhibitory compounds against TR4. Among them, nilotinib exhibited the most potent inhibitor, with an EC50 of 1.05 μM, while genistein represented the most potent activator, with an EC50 of 2.42 μM. Both drugs were predicted to bind in the ligand binding pocket of TR4. The circular dichroism spectroscopic assay revealed differed conformation changes upon nilotinib or genistein binding. These results established our combined physical and biological approaches as a highly effective way to identify and develop new TR4 regulators.
Insights
Researchers identified new regulators for testicular receptor 4 (TR4), a key factor in cancer resistance. Using a novel screening method, they found potent activators and inhibitors, including nilotinib and genistein, offering potential therapeutic strategies.
Area of Science:
- Molecular Endocrinology
- Drug Discovery
- Nuclear Receptor Signaling
Background:
- Testicular receptor 4 (TR4) is a nuclear receptor involved in cancer development and resistance to therapies like chemotherapy and radiotherapy.
- Currently, no effective small-molecule regulators targeting TR4 are available for therapeutic use.
Purpose of the Study:
- To develop and validate a novel assay for discovering small-molecule regulators of TR4.
- To identify compounds that can modulate TR4 activity for potential therapeutic applications.
Main Methods:
- Employed a physical-interaction-based surface plasmon resonance imaging assay to screen 1018 FDA-approved drugs for TR4 binding.
- Utilized a dual-luciferase-based biological assay to verify the activity of identified compounds.
- Confirmed drug binding to the TR4 ligand-binding pocket and analyzed conformational changes using circular dichroism spectroscopy.
Main Results:
- Identified 126 compounds with dissociation constants (KD) below 10-6 M, indicating TR4 binding.
- Validated four activators and two inhibitors of TR4, with genistein as the most potent activator (EC50 = 2.42 μM) and nilotinib as the most potent inhibitor (EC50 = 1.05 μM).
- Observed distinct conformational changes in TR4 upon binding with nilotinib and genistein, suggesting specific interaction mechanisms.
Conclusions:
- Established a combined physical and biological screening approach as a highly effective method for identifying TR4 regulators.
- Nilotinib and genistein represent promising lead compounds for developing novel TR4-targeted therapies to overcome cancer resistance.
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