HNF4α Antagonists Discovered by a High-Throughput Screen for Modulators of the Human Insulin Promoter

Abstract

SUMMARY Hepatocyte Nuclear Factor (HNF)4α is a central regulator of gene expression in cell types that play a critical role in metabolic homeostasis, including hepatocytes, enterocytes, and pancreatic β-cells. Although fatty acids were found to occupy the HNF4α ligand-binding pocket and proposed to act as ligands, there is controversy about both the nature of HNF4α ligands as well as the physiological role of the binding. Here, we report the discovery of potent synthetic HNF4α antagonists through a high-throughput screen for effectors of the human insulin promoter. These molecules bound to HNF4α with high affinity and modulated the expression of known HNF4α target genes. Notably, they were found to be selectively cytotoxic to cancer cell lines in vitro and in vivo, although in vivo potency was limited by suboptimal pharmacokinetic properties. The discovery of bioactive modulators for HNF4α raises the possibility that diseases involving HNF4α, such as diabetes and cancer, might be amenable to pharmacologic intervention by modulation of HNF4α activity.

Hepatocyte Nuclear Factors 4αand 1αRegulate Kidney Developmental Expression of Drug-Metabolizing Enzymes and Drug Transporters

“…By using a recently developed small-molecule antagonist which has been shown to selectively inhibit Hnf4a (Kiselyuk et al, 2012), we were able to explore the consequences of Hnf4a downregulation in early proximal tubules in 7-day organ culture, which serves as a model for kidney development. These have been shown to express Slc22 transporters such as Oat1 and Oat3 and eventually become capable of organic anion transport function (Sweet et al, 2006).…”

“…Media was changed every 3 days. After 6 days, cultures were treated in triplicate with either 1:1000 dimethylsulfoxide, 2.5 mM and 5 mM Hnf4a an- Kiselyuk et al, 2012], or 10 mM analogous inactive compound BI6018 for 12 hours (both kindly provided by Dr. Fred Levine, Sanford Burnham Research Institute). Samples were then stored in RNAlater (Life Technologies, Austin, TX) until further processing for quantitative reverse transcription-polymerase chain reaction (PCR).…”

“…There is currently no reported mouse model with kidney-specific ablation of Hnf4a to test this in vivo. Instead, we used a recently developed small-molecule compound that specifically antagonizes Hnf4a activity (Kiselyuk et al, 2012).…”

Identification of the Flavonoid Luteolin as a Repressor of the Transcription Factor Hepatocyte Nuclear Factor 4α

“…More recently, small synthetic molecules, such as BIM5078 and BI6015, were identified as antagonists for HNF4␣. The binding of their antagonists to HNF4␣ resulted in the suppression of HNF4␣ activity (9), suggesting that exogenous small molecules could control HNF4␣ activity. This finding led us to consider additional investigations to find HNF4␣ antagonists.…”

Protein kinase C-dependent regulation of human hepatic drug transporter expression

“…For this purpose, HepaRG cells were treated by growth factors known to trigger EMT, i.e., HGF [42] or TGF-β1 [43], or by the hepatocyte nuclear factor (HNF) 4α antagonist BI6015 [44], because HNF4α suppression results in EMT in hepatocytes [45]. As shown in Fig.…”