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Rapid degradation of hypoxia-inducible factor-1alpha by KRH102053, a new activator of prolyl hydroxylase 2.

ABSTRACT: Hypoxia-inducible factor (HIF) is a transcription factor induced by hypoxia and degraded by ubiquitin-dependent proteasomes in normoxic conditions. Under hypoxic conditions, hydroxylation of HIF-1alpha subunit by prolyl hydroxylase (PHD) is suppressed, thus leading to increased levels of HIF. Although PHD2 plays a key role in regulating the levels of HIF, chemical activators of PHD2 are relatively unknown. The aim of this study was to identify small molecule activators of PHD2 that could be used, eventually, to suppress the level of HIF-1alpha.By using the overproduced PHD2 as a target, a molecular library consisting of more than 600 small molecules with a benzopyran structure was screened with an HPLC assay method.We found a potent activator of PHD2, KRH102053 (2-amino-4-methylsulphanyl-butylic acid-4-methoxy-6-(4-methoxy-benzylamino)-2,2-dimethyl-chroman-3-yl ester). The effects of KRH102053 on controlling HIF were studied in human HOS osteosarcoma, rat PC12 phaeochromocytoma and human HepG2 hepatoma cells. Under our experimental conditions, KRH102053 decreased the protein level of HIF-1alpha and the mRNA levels of HIF-regulated downstream target genes, such as vascular endothelial growth factor, aldolase A, enolase 1 and monocarboxylate transporter 4. Consistent with these results, KRH102053 also inhibited the rates of HIF-related migration and invasion of HOS cells as well as the degree of tube formation in human umbilical vein endothelium cells.These results suggest that KRH102053 and its structural analogues have the potential for use as therapeutic agents against various diseases associated with HIF.

SUBMITTER: Choi HJ

PROVIDER: S-EPMC2438970 | biostudies-literature | 2008 May

REPOSITORIES: biostudies-literature

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Rapid degradation of hypoxia-inducible factor-1alpha by KRH102053, a new activator of prolyl hydroxylase 2.

Choi H J HJ Song B-J BJ Gong Y-D YD Gwak W J WJ Soh Y Y

British journal of pharmacology 20080310 1

<h4>Background and purpose</h4>Hypoxia-inducible factor (HIF) is a transcription factor induced by hypoxia and degraded by ubiquitin-dependent proteasomes in normoxic conditions. Under hypoxic conditions, hydroxylation of HIF-1alpha subunit by prolyl hydroxylase (PHD) is suppressed, thus leading to increased levels of HIF. Although PHD2 plays a key role in regulating the levels of HIF, chemical activators of PHD2 are relatively unknown. The aim of this study was to identify small molecule activa ...[more]

PMID: 18332861

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Project description:BackgroundThe endothelin B receptor (ET(B)R) promotes tumorigenesis and melanoma progression through activation by endothelin (ET)-1, thus representing a promising therapeutic target. The stability of hypoxia-inducible factor (HIF)-1alpha is essential for melanomagenesis and progression, and is controlled by site-specific hydroxylation carried out by HIF-prolyl hydroxylase domain (PHD) and subsequent proteosomal degradation.Principal findingsHere we found that in melanoma cells ET-1, ET-2, and ET-3 through ET(B)R, enhance the expression and activity of HIF-1alpha and HIF-2alpha that in turn regulate the expression of vascular endothelial growth factor (VEGF) in response to ETs or hypoxia. Under normoxic conditions, ET-1 controls HIF-alpha stability by inhibiting its degradation, as determined by impaired degradation of a reporter gene containing the HIF-1alpha oxygen-dependent degradation domain encompassing the PHD-targeted prolines. In particular, ETs through ET(B)R markedly decrease PHD2 mRNA and protein levels and promoter activity. In addition, activation of phosphatidylinositol 3-kinase (PI3K)-dependent integrin linked kinase (ILK)-AKT-mammalian target of rapamycin (mTOR) pathway is required for ET(B)R-mediated PHD2 inhibition, HIF-1alpha, HIF-2alpha, and VEGF expression. At functional level, PHD2 knockdown does not further increase ETs-induced in vitro tube formation of endothelial cells and melanoma cell invasiveness, demonstrating that these processes are regulated in a PHD2-dependent manner. In human primary and metastatic melanoma tissues as well as in cell lines, that express high levels of HIF-1alpha, ET(B)R expression is associated with low PHD2 levels. In melanoma xenografts, ET(B)R blockade by ET(B)R antagonist results in a concomitant reduction of tumor growth, angiogenesis, HIF-1alpha, and HIF-2alpha expression, and an increase in PHD2 levels.ConclusionsIn this study we identified the underlying mechanism by which ET-1, through the regulation of PHD2, controls HIF-1alpha stability and thereby regulates angiogenesis and melanoma cell invasion. These results further indicate that targeting ET(B)R may represent a potential therapeutic treatment of melanoma by impairing HIF-1alpha stability.

Dataset Information

Rapid degradation of hypoxia-inducible factor-1alpha by KRH102053, a new activator of prolyl hydroxylase 2.

Publications

Rapid degradation of hypoxia-inducible factor-1alpha by KRH102053, a new activator of prolyl hydroxylase 2.

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