Name: GW4064
Brand: Selleck Chemicals
SKU: S2782
Rating: 5 (34 reviews)

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Quality Control

Batch: Purity: 99.80%

99.80

Related Targets	Dehydrogenase HSP Transferase P450 (e.g. CYP17) PDE phosphatase PPAR Vitamin Carbohydrate Metabolism Mitochondrial Metabolism
Other FXR Inhibitors	Guggulsterone E&Z Turofexorate Isopropyl (XL335) fexaramine Tropifexor (LJN452) Cilofexor (GS-9674) BAR502 Nidufexor (LMB-763) LY2562175 Vonafexor (EYP001) Mebhydroline

Cell Culture, Treatment & Working Concentration

Cell Lines	Assay Type	Concentration	Incubation Time	Activity Description	PMID
HEK293	Function assay			Agonistic activity at FXR in HEK293 cells by GAL4 transactivation activity, EC50 = 0.07 μM.	17292610
CV1	Function assay			Agonist activity at FXR expressed in african green monkey CV1 cells by luciferase reporter transient transfection assay, EC50 = 0.065 μM.	18621523
CV1	Function assay			Agonist activity at human FXR transfected in african green monkey CV1 cells by luciferase reporter gene transient transfection assay, EC50 = 0.065 μM.	19410460
CV1	Function assay			Increase in human FXR-mediated transient transcription of luciferase reporter gene transfected in african green monkey CV1 cells, EC50 = 0.065 μM.	19586769
HEK293	Function assay			Agonist activity at human full length FXR transfected in HEK293 cells coexpressing pTRexDest/pGL2promotor assessed as luciferase activity by direct reporter cellular assay, EC50 = 0.03 μM.	20638278
HEK293	Function assay			Agonist activity at human GST-fused FXR LBD expressed in HEK293 cells coexpressing GAL4-DNA bindig domain and pFRluc by mammalian one-hybrid assay, EC50 = 0.035 μM.	20638278
CV-1	Function assay			Agonist activity at human FXR LBD iexpressed in monkey CV-1 cells assessed as transactivation of luciferase reporter gene expression, EC50 = 0.065 μM.	21256005
CV1	Function assay			Agonist activity at human FXR LBD transfected in african green monkey CV1 cells after overnight incubation by luciferase reporter gene assay, EC50 = 0.065 μM.	21890356
HEK293	Function assay			Agonist activity at human recombinant FXR expressed in HEK293 cells coexpressing CMX-GAL4N by luciferase reporter gene assay, EC50 = 0.07 μM.	22583617
HEK293	Function assay			Agonist activity at human FXR expressed in HEK293 cells by luciferase reporter gene assay, EC50 = 0.026 μM.	25305688
CV1	Function assay			Agonist activity at FXR (unknown origin) transfected into african green monkey CV1 cells assessed as ligand-mediated transcription by luciferase reporter/ transient transfection assay, EC50 = 0.065 μM.	25499883
HeLa	Function assay		24 hrs	Agonist activity at human full length FXR expressed in HeLa cells cotransfected with pSG5-human RXR after 24 hrs by Dual-Glo luciferase reporter gene assay, EC50 = 0.51 μM.	25934227
HEK293	Function assay			Agonist activity at human FXR transfected in HEK293 cells assessed as transcriptional activity by luciferase reporter gene assay, EC50 = 0.373 μM.	26568144
HEK-293	Function assay			Agonist activity at C-terminal Gal4-tagged human FXR (187 to 472 residues) expressed in HEK-293 cells co-expressing pFRluc by mammalian one hybrid assay, EC50 = 0.035 μM.	27268696
insect cells	Function assay		1 hr	Agonist activity at recombinant human GST-tagged FXR ligand binding domain (193 to 472 residues) expressed in baculovirus infected insect cells assessed as induction of interaction with biotin labelled SRC-1 after 1 hr by HTRF assay, EC50 = 0.019 μM.	29148806
HEK293T	Function assay		24 hrs	Agonist activity at human FXR expressed in HEK293T cells assessed as BSEP promoter driven cellular transcriptional activity after 24 hrs by luciferase reporter gene assay, EC50 = 0.095 μM.	29148806
HepG2	Function assay	1 uM	18 hrs	Decrease in CYP7A1 gene expression in human HepG2 cells at 1 uM after 18 hrs by RT-PCR	17963371
Caco-2	Function assay	1 uM	6 days	Activation of IBABP gene expression in human Caco-2 cells at 1 uM after 6 days by RT-PCR	17963371
HepG2	Function assay	1 uM	18 hrs	Activation of BSEP gene expression in human HepG2 cells at 1 uM after 18 hrs by RT-PCR	17963371
Huh7	Function assay	1 uM	18 hrs	Activation of SHP gene expression in human Huh7 cells at 1 uM after 18 hrs by RT-PCR	17963371
Huh7	Function assay	1 uM	18 hrs	Activation of BSEP gene expression in human Huh7 cells at 1 uM after 18 hrs by RT-PCR	17963371
HepG2	Function assay	10 uM		Induction of FXR-mediated SHP mRNA expression in serum-starved human HepG2 cells at 10 uM measured after overnight incubation by qPCR analysis	27372840
HepG2	Function assay	10 uM	10 mins	Induction of FXR-mediated Akt-phosphorylation at S473 site in serum-starved human HepG2 cells at 10 uM measured after 10 mins by western blot analysis	27372840
HepG2	Function assay	10 uM	10 mins	Induction of FXR-mediated Akt-phosphorylation in serum-starved human HepG2 cells assessed as increase in GSK-3beta-phosphorylation at 10 uM measured after 10 mins by western blot analysis	27372840
HepG2	Function assay	10 uM		Activation of FXR in human HepG2 cells assessed as upregulation of SHP mRNA expression at 10 uM by RT-PCR method	30031618
HepG2	Function assay	10 uM		Activation of FXR in human HepG2 cells assessed as upregulation of FXR mRNA expression at 10 uM by RT-PCR method	30031618
HepG2	Function assay	10 uM		Activation of FXR in human HepG2 cells assessed as downregulation of SREBP-1c mRNA expression at 10 uM by RT-PCR method	30031618
3T3L1	Function assay	10 uM		Reduction in lipid accumulation in mouse 3T3L1 cells at 10 uM by Oil Red O staining-based absorbance method relative to control	30342957
Click to View More Cell Line Experimental Data

Chemical Information, Storage & Stability

Molecular Weight	542.84	Formula	C₂₈H₂₂Cl₃NO₄	Storage (From the date of receipt)	3 years-20°Cpowder
CAS No.	278779-30-9	Download SDF		Storage of Stock Solutions	1 year-80°Cin solvent 1 month-20°Cin solvent
Synonyms	N/A			Smiles	CC(C)C1=C(C(=NO1)C2=C(C=CC=C2Cl)Cl)COC3=CC(=C(C=C3)C=CC4=CC(=CC=C4)C(=O)O)Cl

Solubility

In vitro
Batch:

DMSO : 100 mg/mL (184.21 mM)
(Moisture-contaminated DMSO may reduce solubility. Use fresh, anhydrous DMSO.)

Water : Insoluble

Ethanol : Insoluble

Molarity Calculator

Mass	Concentration	Volume	Molecular Weight
=	×	×

Dilution Calculator Molecular Weight Calculator

In vivo Batch:
Homogeneous suspension	CMC-NA	≥5mg/ml	Taking the 1 mL working solution as an example, add 5 mg of this product to 1 ml of CMC-Na solution, mix evenly to obtain a homogeneous suspension with a final concentration of 5 mg/ml.
Clear solution	5%DMSO 1 40%PEG300 2 5%Tween80 3 50%ddH2O Validated by Selleck labs. Should you need adjustments to this formulation, contact our sales team for custom testing.	2.500mg/ml (4.61mM)	Taking the 1 mL working solution as an example, add 50 μL of 50 mg/ml clarified DMSO stock solution to 400 μL of PEG300, mix evenly to clarify it; add 50 μL of Tween80 to the above system, mix evenly to clarify; then continue to add 500 μL of ddH2O to adjust the volume to 1 mL. The mixed solution should be used immediately for optimal results.
Clear solution	5% DMSO 1 95% Corn oil Validated by Selleck labs. Should you need adjustments to this formulation, contact our sales team for custom testing.	0.600mg/ml (1.11mM)	Taking the 1 mL working solution as an example, add 50 μL of 12 mg/ml clear DMSO stock solution to 950 μL of corn oil and mix evenly. The mixed solution should be used immediately for optimal results.
Homogeneous suspension	CMC-NA	≥5mg/ml	Taking the 1 mL working solution as an example, add 5 mg of this product to 1 ml of CMC-Na solution, mix evenly to obtain a homogeneous suspension with a final concentration of 5 mg/ml.
Homogeneous suspension	CMC-NA	≥5mg/ml	Taking the 1 mL working solution as an example, add 5 mg of this product to 1 ml of CMC-Na solution, mix evenly to obtain a homogeneous suspension with a final concentration of 5 mg/ml.
Homogeneous suspension	0.5% methylcellulose Validated by Selleck labs. Should you need adjustments to this formulation, contact our sales team for custom testing.	10.000mg/ml (18.42mM)	Taking the 1 mL working solution as an example, take 10 mg of this product, add it to 1 ml of 0.5% methylcellulose clear solution, and mix evenly to form a uniform suspension. The mixed solution should be used immediately for optimal results.
Homogeneous suspension	CMC-NA	≥5mg/ml	Taking the 1 mL working solution as an example, add 5 mg of this product to 1 ml of CMC-Na solution, mix evenly to obtain a homogeneous suspension with a final concentration of 5 mg/ml.
Homogeneous suspension	CMC-NA	≥5mg/ml	Taking the 1 mL working solution as an example, add 5 mg of this product to 1 ml of CMC-Na solution, mix evenly to obtain a homogeneous suspension with a final concentration of 5 mg/ml.

In vivo Formulation Calculator (Clear solution)

Step 1: Enter information below (Recommended: An additional animal making an allowance for loss during the experiment)

Dosage: mg/kg Average weight of animals: g Dosing volume per animal:μL Number of animals:

Step 2: Enter the in vivo formulation (This is only the calculator, not formulation. Please contact us first if there is no in vivo formulation at the solubility Section.)

% DMSO + % + % Tween 80 + % ddH₂O

%DMSO+ %

Calculation results:

Working concentration: mg/ml;

Method for preparing DMSO master liquid: mg drug pre-dissolved in μL DMSO ( Master liquid concentration mg/mL, Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug. )

Method for preparing in vivo formulation: Take μL DMSO master liquid, next addμL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH₂O, mix and clarify.

Method for preparing in vivo formulation: Take μL DMSO master liquid, next add μL Corn oil, mix and clarify.

Note: 1. Please make sure the liquid is clear before adding the next solvent.
2. Be sure to add the solvent(s) in order. You must ensure that the solution obtained, in the previous addition, is a clear solution before proceeding to add the next solvent. Physical methods such as vortex, ultrasound or hot water bath can be used to aid dissolving.

Mechanism of Action

Targets/IC₅₀/Ki	FXR (CV1) 65 nM(EC50)
In vitro	GW 4064 is a full agonist with EC50 values of 80 and 90 nM, respectively, in CV-1 cells transfected with mouse and human FXR expression vectors and an established reporter gene. There is no activity of this compound on other nuclear receptors, including the retinoic acid receptor, at concentrations up to 1 μM. Thus, this chemical is a potent and selective nonsteroidal FXR agonist.
Kinase Assay	FXR Transient Transfection Assay
Kinase Assay	CV1 cells are maintained in DMEM high glucose medium. The CV1 cells are harvested 72 hours before transfection and plated in DMEM-F12 phenol red free medium containing 5% charcoal/dextran treated fetal bovine serum and 2 mM glutamine. On the day of transfection, the CV1 cells are harvested in DMEM-F12 phenol red free medium containing 5% charcoal/dextran treated fetal bovine serum and 2 mM glutamine, counted and seeded into a T-175 cm² flask at a density of 14 million cells per flask before adding the transfection mix. The FuGENE® 6 transfection mix containing 0.55 μg pFA-CMV-GAL4-human FXR LBD expression plasmid, 10.92 μg of the (UAS)-tk-Luciferase reporter plasmid, 20.75 μg of pBluescript II KS+, and 3.82 μg of a human steroid receptor co-activator (SRC-1) construct is incubated and then added to the CV1 cells for overnight incubation in incubator. The transfected CV1 cells are harvested in DMEM-F12 phenol red free medium containing 5% charcoal/dextran treated fetal bovine serum and 2 mM glutamine. Transfected cells are added at a concentration of 3750 cells/well to 384-well plates containing ligands. Luciferase expression is measured using Steady-Glo™Luciferase Assay System 24 hr after incubation with ligand. This compound is used as the control, and data are normalized to this compound as % control.
In vivo	Pharmacokinetic analysis in rats shows that GW 4064 possesses an oral bioavailability of 10% with a t_1/2 = 3.5 h. Fisher rats are dosed with this compound by oral gavage. After 7 days, a dose-dependent lowering of serum triglycerides is observed in the rats receiving this chemical, with an ED50 = 20 mg/kg.
References	[1] https://pubmed.ncbi.nlm.nih.gov/18621523/ [2] https://pubmed.ncbi.nlm.nih.gov/10956205/ [3] https://pubmed.ncbi.nlm.nih.gov/35956351/

Applications

Methods	Biomarkers	Images	PMID
Western blot	PTEN / p-AKT / AKT p-EGFR / EGFR / p-ERK / ERK / p-Src / Src SOCS3 / p21 / p-STAT3 / STAT3		25187826
Growth inhibition assay	Cell proliferation		25187826

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Handling Instructions

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GW4064 FXR agonist

Chemical Structure

Molecular Weight: 542.84