GW4064

GW4064 is an agonist of FXR with EC50 values of 15nM and 90nM, respectively in an isolated receptor activity assay and in cells transfected with human FXR [1].

GW4064 is identified as a highly effective, selective, nonsteroidal agonist of FXR. It shows beneficial effects on cholesterol and TG in various animal species. GW4064 is found to lower serum TG levels in both the KK-Ay and ob/ob mice potently. In KK-Ay mice, a 1-week administration of GW4064 also significantly lowers VLDL secretion. And in the SHP+/+ mice, GW4064 can also lower serum TGs. [2]

Due to some limitations, such as limited solubility, potentially toxic stilbene pharmacophore and UV light instability, GW4064 is not a good candidate of drug. It is now usually used as a tool compound for investigating the physiological functions of FXR [3].

References:
[1] Chiang PC, Thompson DC, Ghosh S, Heitmeier MR. A formulation-enabled preclinical efficacy assessment of a farnesoid X receptor agonist, GW4064, in hamsters and cynomolgus monkeys. J Pharm Sci. 2011 Nov;100(11):4722-33.
[2] Watanabe M, Houten SM, Wang L, Moschetta A, Mangelsdorf DJ, Heyman RA, Moore DD, Auwerx J. Bile acids lower triglyceride levels via a pathway involving FXR, SHP, and SREBP-1c. J Clin Invest. 2004 May;113(10):1408-18.
[3] Li W, Fu J, Cheng F, Zheng M, Zhang J, Liu G, Tang Y. Unbinding pathways of GW4064 from human farnesoid X receptor as revealed by molecular dynamics simulations. J Chem Inf Model. 2012 Nov 26;52(11):3043-52.

Restoration of enterohepatic bile acid pathways in pregnant mice following short term activation of Fxr by GW4064.[Pubmed:27609522]

Toxicol Appl Pharmacol. 2016 Nov 1;310:60-67.

The farnesoid X receptor (Fxr) controls bile acid homeostasis by coordinately regulating the expression of synthesizing enzymes (Cyp7a1, Cyp8b1), conjugating enzymes (Bal, Baat) and transporters in the ileum (Asbt, Ostalpha/beta) and liver (Ntcp, Bsep, Ostbeta). Transcriptional regulation by Fxr can be direct, or through the ileal Fgf15/FGF19 and hepatic Shp pathways. Circulating bile acids are increased during pregnancy due to hormone-mediated disruption of Fxr signaling. While this adaptation enhances lipid absorption, elevated bile acids may predispose women to develop maternal cholestasis. The objective of this study was to determine whether short-term treatment of pregnant mice with GW4064 (a potent FXR agonist) restores Fxr signaling to the level observed in virgin mice. Plasma, liver and ilea were collected from virgin and pregnant mice administered vehicle or GW4064 by oral gavage. Treatment of pregnant mice with GW4064 induced ileal Fgf15, Shp and Ostalpha/beta mRNAs, and restored hepatic Shp, Bal, Ntcp, and Bsep back to vehicle-treated virgin levels. Pregnant mice exhibited 2.5-fold increase in Cyp7a1 mRNA compared to virgin controls, which was reduced by GW4064. Similarly treatment of mouse primary hepatocytes with plasma isolated from pregnant mice induced Cyp7a1 mRNA by nearly 3-fold as compared to virgin plasma, which could be attenuated by co-treatment with either GW4064 or recombinant FGF19 protein. Collectively, these data reveal that repressed activity of intestinal and hepatic Fxr in pregnancy, as previously demonstrated, may be restored by pharmacological activation. This study provides the basis for a novel approach to restore bile acid homeostasis in patients with maternal cholestasis.

Novel heterocyclic scaffolds of GW4064 as farnesoid X receptor agonists.[Pubmed:25499883]

Bioorg Med Chem Lett. 2015 Jan 15;25(2):280-4.

The farnesoid X receptor (FXR) may play a crucial role in a number of metabolic diseases and, as such, could potentially serve as a target for the development of therapeutics as a treatment for those diseases. Previous work has described GW4064 as an FXR agonist with an interesting activity profile. This manuscript will describe the synthesis of novel analogs of GW4064 and the activity profile of those analogs.

GW4064, an agonist of farnesoid X receptor, represses CYP3A4 expression in human hepatocytes by inducing small heterodimer partner expression.[Pubmed:25725071]

Drug Metab Dispos. 2015 May;43(5):743-8.

Farnesoid X receptor (FXR) functions as a regulator of bile acid and lipid homeostasis and is recognized as a promising therapeutic target for metabolic diseases. The biologic function of FXR is mediated in part by a small heterodimer partner (SHP); ligand-activated FXR enhances SHP expression, and SHP in turn represses the activity of multiple transcription factors. This study aimed to investigate the effect of FXR activation on expression of the major drug-metabolizing enzyme CYP3A4. The effects of 3-(2,6-dichlorophenyl)-4-(3'-carboxy-2-chlorostilben-4-yl)oxymethyl-5-isopropylis oxazole (GW4064), a synthetic agonist of FXR, on the expression and activity of CYP3A4 were examined in primary human hepatocytes by using quantitative real-time polymerase chain reaction and S9 phenotyping. In human hepatocytes, treatment of GW4064 (1 muM) for 48 hours resulted in a 75% decrease in CYP3A4 mRNA expression and a 25% decrease in CYP3A4 activity, accompanied by approximately 3-fold increase in SHP mRNA expression. In HepG2 cells, SHP repressed transactivation of CYP3A4 promoter by pregnane X receptor (PXR), constitutive androstane receptor (CAR), and glucocorticoid receptor. Interestingly, GW4064 did not repress expression of CYP2B6, another target gene of PXR and CAR; GW4064 enhanced CYP2B6 promoter activity. In conclusion, GW4064 represses CYP3A4 expression in human hepatocytes, potentially through upregulation of SHP expression and subsequent repression of CYP3A4 promoter activity. Clinically significant drug-drug interaction involving FXR agonists and CYP3A4 substrates may occur.

Fragmentation of GW4064 led to a highly potent partial farnesoid X receptor agonist with improved drug-like properties.[Pubmed:25934227]

Bioorg Med Chem. 2015 Jul 1;23(13):3490-8.

The ligand activated transcription factor farnesoid X receptor (FXR) is a crucial regulator of several metabolic and inflammatory pathways and its activation by agonistic ligands seems a valuable therapeutic approach for many disorders. Most known non-steroidal FXR agonists however, have limitations that hinder their clinical development and novel FXR ligands are required. Evaluation of the co-crystal structures of the widely used FXR agonist GW4064 and related compounds in complex with the FXR ligand binding domain indicated that their disubstituted isoxazole moiety is especially relevant for FXR activation. By investigation of GW4064-fragments missing the aromatic tail, we discovered a highly potent and soluble partial FXR agonist (14, ST-1892) as well as a fluorescent FXR ligand (15) as potential pharmacological tool.

Nutrient-sensing nuclear receptors coordinate autophagy.[Pubmed:25383539]

Nature. 2014 Dec 4;516(7529):112-5.

Autophagy is an evolutionarily conserved catabolic process that recycles nutrients upon starvation and maintains cellular energy homeostasis. Its acute regulation by nutrient-sensing signalling pathways is well described, but its longer-term transcriptional regulation is not. The nuclear receptors peroxisome proliferator-activated receptor-alpha (PPARalpha) and farnesoid X receptor (FXR) are activated in the fasted and fed liver, respectively. Here we show that both PPARalpha and FXR regulate hepatic autophagy in mice. Pharmacological activation of PPARalpha reverses the normal suppression of autophagy in the fed state, inducing autophagic lipid degradation, or lipophagy. This response is lost in PPARalpha knockout (Ppara(-/-), also known as Nr1c1(-/-)) mice, which are partially defective in the induction of autophagy by fasting. Pharmacological activation of the bile acid receptor FXR strongly suppresses the induction of autophagy in the fasting state, and this response is absent in FXR knockout (Fxr(-/-), also known as Nr1h4(-/-)) mice, which show a partial defect in suppression of hepatic autophagy in the fed state. PPARalpha and FXR compete for binding to shared sites in autophagic gene promoters, with opposite transcriptional outputs. These results reveal complementary, interlocking mechanisms for regulation of autophagy by nutrient status.

The farnesoid X receptor modulates adiposity and peripheral insulin sensitivity in mice.[Pubmed:16446356]

J Biol Chem. 2006 Apr 21;281(16):11039-49.

The farnesoid X receptor (FXR) is a bile acid (BA)-activated nuclear receptor that plays a major role in the regulation of BA and lipid metabolism. Recently, several studies have suggested a potential role of FXR in the control of hepatic carbohydrate metabolism, but its contribution to the maintenance of peripheral glucose homeostasis remains to be established. FXR-deficient mice display decreased adipose tissue mass, lower serum leptin concentrations, and elevated plasma free fatty acid levels. Glucose and insulin tolerance tests revealed that FXR deficiency is associated with impaired glucose tolerance and insulin resistance. Moreover, whole-body glucose disposal during a hyperinsulinemic euglycemic clamp is decreased in FXR-deficient mice. In parallel, FXR deficiency alters distal insulin signaling, as reflected by decreased insulin-dependent Akt phosphorylation in both white adipose tissue and skeletal muscle. Whereas FXR is not expressed in skeletal muscle, it was detected at a low level in white adipose tissue in vivo and induced during adipocyte differentiation in vitro. Moreover, mouse embryonic fibroblasts derived from FXR-deficient mice displayed impaired adipocyte differentiation, identifying a direct role for FXR in adipocyte function. Treatment of differentiated 3T3-L1 adipocytes with the FXR-specific synthetic agonist GW4064 enhanced insulin signaling and insulin-stimulated glucose uptake. Finally, treatment with GW4064 improved insulin resistance in genetically obese ob/ob mice in vivo. Although the underlying molecular mechanisms remain to be unraveled, these results clearly identify a novel role of FXR in the regulation of peripheral insulin sensitivity and adipocyte function. This unexpected function of FXR opens new perspectives for the treatment of type 2 diabetes.