TY - JOUR
T1 - Synthetic FXR agonist GW4064 is a modulator of multiple G protein-coupled receptors
AU - Singh, Nidhi
AU - Yadav, Manisha
AU - Singh, Abhishek Kumar
AU - Kumar, Harish
AU - Dhar Dwivedi, Shailendra Kumar
AU - Mishra, Jay Sharan
AU - Gurjar, Anagha
AU - Manhas, Amit
AU - Chandra, Sharat
AU - Yadav, Prem Narayan
AU - Jagavelu, Kumaravelu
AU - Siddiqi, Mohammad Imran
AU - Trivedi, Arun Kumar
AU - Chattopadhyay, Naibedya
AU - Sanyal, Sabyasachi
PY - 2014/5
Y1 - 2014/5
N2 - The synthetic nuclear bile acid receptor (farnesoid X receptor [FXR]) agonist GW4064 is extensively used as a specific pharmacological tool to illustrate FXR functions. We noticed that GW4064 activated empty luciferase reporters in FXR-deficient HEK-293T cells.Wepostulated that this activity of GW4064 might be routed through as yet unknown cellular targets and undertook an unbiased exploratory approach to identify these targets. Investigations revealed that GW4064 activated cAMP and nuclear factor for activated T-cell response elements (CRE and NFAT-RE, respectively) present on these empty reporters. Whereas GW4064-induced NFAT-RE activation involved rapid intracellular Ca2+ accumulation and NFAT nuclear translocation, CRE activation involved soluble adenylyl cyclase-dependent cAMP accumulation and Ca2+-calcineurin-dependent nuclear translocation of transducers of regulated CRE-binding protein 2. Use of dominant negative heterotrimeric G-protein minigenes revealed that GW4064 caused activation of Gαi/o and Gq/11 G proteins. Sequential pharmacological inhibitor- based screening and radioligand-binding studies revealed that GW4064 interacted with multiple G protein-coupled receptors. Functional studies demonstrated that GW4064 robustly activated H1 and H4and inhibitedH2histamine receptor signaling events.Wealso found that MCF-7 breast cancer cells, reported to undergo GW4064-induced apoptosis in an FXR-dependent manner, did not express FXR, and the GW4064-mediated apoptosis, also apparent in HEK-293T cells, could be blocked by selective histamine receptor regulators. Taken together, our results demonstrate identification of histamine receptors as alternate targets for GW4064, which not only necessitates cautious interpretation of the biological functions attributed to FXR using GW4064 as a pharmacological tool but also provides a basis for the rational designing of new pharmacophores for histamine receptor modulation.
AB - The synthetic nuclear bile acid receptor (farnesoid X receptor [FXR]) agonist GW4064 is extensively used as a specific pharmacological tool to illustrate FXR functions. We noticed that GW4064 activated empty luciferase reporters in FXR-deficient HEK-293T cells.Wepostulated that this activity of GW4064 might be routed through as yet unknown cellular targets and undertook an unbiased exploratory approach to identify these targets. Investigations revealed that GW4064 activated cAMP and nuclear factor for activated T-cell response elements (CRE and NFAT-RE, respectively) present on these empty reporters. Whereas GW4064-induced NFAT-RE activation involved rapid intracellular Ca2+ accumulation and NFAT nuclear translocation, CRE activation involved soluble adenylyl cyclase-dependent cAMP accumulation and Ca2+-calcineurin-dependent nuclear translocation of transducers of regulated CRE-binding protein 2. Use of dominant negative heterotrimeric G-protein minigenes revealed that GW4064 caused activation of Gαi/o and Gq/11 G proteins. Sequential pharmacological inhibitor- based screening and radioligand-binding studies revealed that GW4064 interacted with multiple G protein-coupled receptors. Functional studies demonstrated that GW4064 robustly activated H1 and H4and inhibitedH2histamine receptor signaling events.Wealso found that MCF-7 breast cancer cells, reported to undergo GW4064-induced apoptosis in an FXR-dependent manner, did not express FXR, and the GW4064-mediated apoptosis, also apparent in HEK-293T cells, could be blocked by selective histamine receptor regulators. Taken together, our results demonstrate identification of histamine receptors as alternate targets for GW4064, which not only necessitates cautious interpretation of the biological functions attributed to FXR using GW4064 as a pharmacological tool but also provides a basis for the rational designing of new pharmacophores for histamine receptor modulation.
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U2 - 10.1210/me.2013-1353
DO - 10.1210/me.2013-1353
M3 - Article
C2 - 24597548
AN - SCOPUS:84899706348
SN - 0888-8809
VL - 28
SP - 659
EP - 673
JO - Molecular Endocrinology
JF - Molecular Endocrinology
IS - 5
ER -