Ligand Activation of ERR? by Cholesterol Mediates Statin and Bisphosphonate Effects.
ABSTRACT: Nuclear receptors (NRs) are key regulators of gene expression and physiology. Nearly half of all human NRs lack endogenous ligands including estrogen-related receptor ? (ERR?). ERR? has important roles in cancer, metabolism, and skeletal homeostasis. Affinity chromatography of tissue lipidomes with the ERR? ligand-binding domain (LBD) and subsequent transcriptional assays identified cholesterol as an endogenous ERR? agonist. Perturbation of cholesterol biosynthesis or inhibition of ERR? revealed the interdependence of cholesterol and ERR?. In bone, the effects of cholesterol, statin, and bisphosphonate on osteoclastogenesis require ERR?; and consequently, cholesterol-induced bone loss or bisphosphonate osteoprotection is lost in ERR? knockout mice. Furthermore, statin induction of muscle toxicity and cholesterol suppression of macrophage cytokine secretion are impaired by loss or inhibition of ERR?. These findings reveal a key step in ERR? regulation and explain the actions of two highly prescribed drugs, statins and bisphosphonates.
Project description:Breast cancer is the 2nd leading cause of cancer-related death among women. Increased risk of breast cancer has been associated with high dietary cholesterol intake. However, the underlying mechanisms are not known. The nuclear receptor, estrogen-related receptor alpha (ERR?), plays an important role in breast cancer cell metabolism, and its overexpression has been linked to poor survival. Here we identified cholesterol as an endogenous ligand of ERR? by purification from human pregnancy serum using a GST-ERR? affinity column and liquid chromatography-tandem mass spectrometry (LC-MS/MS). We show that cholesterol interacts with ERR? and induces its transcriptional activity in estrogen receptor positive (ER+) and triple negative breast cancer (TNBC) cells. In addition, we show that cholesterol enhances ERR?-PGC-1? interaction, induces ERR? expression itself, augments several metabolic target genes of ERR?, and increases cell proliferation and migration in both ER+ and TNBC cells. Furthermore, the stimulatory effect of cholesterol on metabolic gene expression, cell proliferation, and migration requires the ERR? pathway. These findings provide a mechanistic explanation for the increased breast cancer risk associated with high dietary cholesterol and possibly the pro-survival effect of statins in breast cancer patients, highlighting the clinical relevance of lowering cholesterol levels in breast cancer patients overexpressing ERR?.
Project description:Transcriptional regulators include a superfamily of nuclear proteins referred to as co-activators and co-repressors, both of which are involved in controlling the functions of several nuclear receptors (NRs). The Nuclear Receptor Signaling Atlas (NURSA) has cataloged the composition of NRs, co-regulators, and ligands present in the human cell and their effort has been identified in more than 600 potential molecules. Given the importance of co-regulators in steroid, retinoid, and thyroid hormone signaling networks, hypothesizing that NRs/co-regulators are implicated in a wide range of pathologies are tempting. The co-activators known as peroxisome proliferator-activated receptor gamma co-activator 1 (PGC-1) and their key nuclear partner, the estrogen-related receptor (ERR), are emerging as pivotal transcriptional signatures that regulate an extremely broad repertoire of mitochondrial and metabolic genes, making them very attractive drug targets for cancer. Several studies have provided an increased understanding of the functional and structural biology of nuclear complexes. However, more comprehensive work is needed to create different avenues to explore the therapeutic potential of NRs/co-activators in precision oncology. Here, we discuss the emerging data associated with the structure, function, and molecular biology of the PGC-1/ERR network and address how the concepts evolving from these studies have deepened our understanding of how to develop more effective treatment strategies. We present an overview that underscores new biological insights into PGC-1/ERR to improve cancer outcomes against therapeutic resistance. Finally, we discuss the importance of exploiting new technologies such as single-particle cryo-electron microscopy (cryo-EM) to develop a high-resolution biological structure of PGC-1/ERR, focusing on novel drug discovery for precision oncology.
Project description:<h4>Background</h4>PGC-1? is a crucial regulator of cellular metabolism and energy homeostasis that functionally acts together with the estrogen-related receptors (ERR? and ERR?) in the regulation of mitochondrial and metabolic gene networks. Dimerization of the ERRs is a pre-requisite for interactions with PGC-1? and other coactivators, eventually leading to transactivation. It was suggested recently (Devarakonda et al) that PGC-1? binds in a strikingly different manner to ERR? ligand-binding domains (LBDs) compared to its mode of binding to ERR? and other nuclear receptors (NRs), where it interacts directly with the two ERR? homodimer subunits.<h4>Methods/principal findings</h4>Here, we show that PGC-1? receptor interacting domain (RID) binds in an almost identical manner to ERR? and ERR? homodimers. Microscale thermophoresis demonstrated that the interactions between PGC-1? RID and ERR LBDs involve a single receptor subunit through high-affinity, ERR-specific L3 and low-affinity L2 interactions. NMR studies further defined the limits of PGC-1? RID that interacts with ERRs. Consistent with these findings, the solution structures of PGC-1?/ERR? LBDs and PGC-1?/ERR? LBDs complexes share an identical architecture with an asymmetric binding of PGC-1? to homodimeric ERR.<h4>Conclusions/significance</h4>These studies provide the molecular determinants for the specificity of interactions between PGC-1? and the ERRs, whereby negative cooperativity prevails in the binding of the coactivators to these receptors. Our work indicates that allosteric regulation may be a general mechanism controlling the binding of the coactivators to homodimers.
Project description:As transcriptional factors, nuclear receptors (NRs) function as major regulators of gene expression. In particular, dysregulation of NR activity has been shown to significantly alter metabolic homeostasis in various contexts leading to metabolic disorders and cancers. The orphan estrogen-related receptor (ERR) subfamily of NRs, comprised of ERR?, ERR?, and ERR?, for which a natural ligand has yet to be identified, are known as central regulators of energy metabolism. If AMP-activated protein kinase (AMPK) and mechanistic target of rapamycin (mTOR) can be viewed as sensors of the metabolic needs of a cell and responding acutely via post-translational control of proteins, then the ERRs can be regarded as downstream effectors of metabolism via transcriptional regulation of genes for a long-term and sustained adaptive response. In this review, we will focus on recent findings centered on the transcriptional roles played by ERR? in hepatocytes. Modulation of ERR? activity in both in vitro and in vivo models via genetic or pharmacological manipulation coupled with chromatin-immunoprecipitation (ChIP)-on-chip and ChIP-sequencing (ChIP-seq) studies have been fundamental in delineating the direct roles of ERR? in the control of hepatic gene expression. These studies have identified crucial roles for ERR? in lipid and carbohydrate metabolism as well as in mitochondrial function under both physiological and pathological conditions. The regulation of ERR? expression and activity via ligand-independent modes of action including coregulator binding, post-translational modifications (PTMs) and control of protein stability will be discussed in the context that may serve as valuable tools to modulate ERR? function as new therapeutic avenues for the treatment of hepatic metabolic dysfunction and related diseases.
Project description:Bile acids are primarily synthesized from cholesterol in the liver and have important roles in dietary lipid absorption and cholesterol homoeostasis. Detailed roles of the orphan nuclear receptors regulating cholesterol 7?-hydroxylase (CYP7A1), the rate-limiting enzyme in bile acid synthesis, have not yet been fully elucidated. In the present study, we report that oestrogen-related receptor ? (ERR?) is a novel transcriptional regulator of CYP7A1 expression. Activation of cannabinoid receptor type 1 (CB1 receptor) signalling induced ERR?-mediated transcription of the CYP7A1 gene. Overexpression of ERR? increased CYP7A1 expression in vitro and in vivo, whereas knockdown of ERR? attenuated CYP7A1 expression. Deletion analysis of the CYP7A1 gene promoter and a ChIP assay revealed an ERR?-binding site on the CYP7A1 gene promoter. Small heterodimer partner (SHP) inhibited the transcriptional activity of ERR? and thus regulated CYP7A1 expression. Overexpression of ERR? led to increased bile acid levels, whereas an inverse agonist of ERR?, GSK5182, reduced CYP7A1 expression and bile acid synthesis. Finally, GSK5182 significantly reduced hepatic CB1 receptor-mediated induction of CYP7A1 expression and bile acid synthesis in alcohol-treated mice. These results provide the molecular mechanism linking ERR? and bile acid metabolism.
Project description:<h4>Objective</h4>To seek evidence for the association of bisphosphonate use with diffuse musculoskeletal pain (MSKP) in a large national cohort, controlling for conditions associated with MSKP.<h4>Patients and methods</h4>This retrospective cohort study enrolled all US veterans aged 65 years or older with a vertebral or hip fracture who were treated for at least 1 year between October 1, 1998, and September 30, 2006 (N=26,545). All International Classification of Diseases, Ninth Revision, Clinical Modification (ICD-9-CM) diagnostic codes, demographics, and pharmaceutical data were obtained from national databases. A composite end point, based on ICD-9-CM codes compatible with diffuse MSKP, was constructed. The primary outcome was time until MSKP. We performed regression analysis using the Cox proportional hazards model, controlling for age, sex, race, alcoholism, depression, anxiety, smoking, recent 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitor (statin) use, rheumatic disease, and comorbidity score.<h4>Results</h4>The univariate regression identified an association of bisphosphonate exposure and MSKP (hazard ratio, 1.22; 95% confidence interval, 1.04-1.44). In the multivariate regression, however, patients prescribed a bisphosphonate were not more likely to be assigned an ICD-9-CM code compatible with diffuse MSKP (hazard ratio, 1.10; 95% confidence interval, 0.93-1.30). Consistent with prior studies, we found that female sex, depression, anxiety, comorbidity score, and the presence of a rheumatic disease were all associated with a greater risk of a diagnosis of diffuse MSKP. There was no demonstrable association with statin exposure.<h4>Conclusion</h4>Bisphosphonate use was not associated with a statistically higher rate of MSKP in this cohort. Individual patients may rarely report MSKP while taking bisphosphonates; however, for our studied cohort, incident MSKP does not appear to explain bisphosphonate discontinuation rates.
Project description:Breast cancer is the second greatest cause of cancer-related death in women. Resistance to endocrine treatments or chemotherapy is a limiting drawback. In this context, this work aims to evaluate the effects of cholesterol and mevalonate during tumor progression and their contribution in the onset of resistance to clinical treatments in use today. In this study, we demonstrated that cholesterol and mevalonate treatments were able to activate the estrogen-related receptor alpha (ERR?) pathway, increasing the expression levels of peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1?), ERbB2/human epithelial receptor (HER2), tumor protein D52 (TPD52), and NOTCH2 proteins in breast cancer cells. The activation of this pathway is shown to be responsible for intense metabolic switching, higher proliferation rates, sustained motility, the propagation of cancer stem-like cells (CSCs), and lipid droplet formation. All of these events are related to greater tumor propagation, aggressiveness, and drug resistance. Furthermore, the activation and expression of proteins induced by the treatment with cholesterol or mevalonate are consistent with those obtained from the MCF-7/TAMr cell line, which is largely used as a breast cancer model of acquired endocrine therapy resistance. Altogether, our data indicate that cholesterol and mevalonate are two metabolites implicated in breast cancer progression, aggressiveness, and drug resistance, through the activation of the ERR? pathway. Our findings enable us to identify the ERR? receptor as a poor prognostic marker in patients with breast carcinoma, suggesting the correlation between cholesterol/mevalonate and ERR? as a new possible target in breast cancer treatment.
Project description:Statins and bisphosphonates are increasingly recognized as anti-cancer drugs, especially because of their cholesterol-lowering properties. However, these drugs act differently on various types of cancers. Thus, the aim of this study was to compare the effects of statins and bisphosphonates on the metabolism (NADP?/NADPH-relation) of highly proliferative tumor cell lines from different origins (PC-3 prostate carcinoma, MDA-MB-231 breast cancer, U-2 OS osteosarcoma) versus cells with a slower proliferation rate like MG-63 osteosarcoma cells. Global gene expression analysis revealed that after 6 days of treatment with pharmacologic doses of the statin simvastatin and of the bisphosphonate ibandronate, simvastatin regulated more than twice as many genes as ibandronate, including many genes associated with cell cycle progression. Upregulation of starvation-markers and a reduction of metabolism and associated NADPH production, an increase in autophagy, and a concomitant downregulation of H3K27 methylation was most significant in the fast-growing cancer cell lines. This study provides possible explanations for clinical observations indicating a higher sensitivity of rapidly proliferating tumors to statins and bisphosphonates.
Project description:BACKGROUND:Peroxisome proliferator-activated receptor-? co-activator-1? (PGC-1?) is a member of the transcriptional coactivator family that plays a central role in the regulation of cellular energy metabolism under various physiological stimuli. During fasting, PGC-1? is induced in the liver and together with estrogen-related receptor a and ? (ERR? and ERR?, orphan nuclear receptors with no known endogenous ligand, regulate sets of genes that participate in the energy balance program. We found that PGC-1?, ERR? and ERR? was highly expressed in human kidney HK2 cells and that PGC-1? induced dynamic protein interactions on the ERR? chromatin. However, the effect of fasting on the expression of endogenous PGC-1?, ERR? and ERR? in the kidney is not known. METHODOLOGY/PRINCIPAL FINDINGS:In this study, we demonstrated by qPCR that the expression of PGC-1?, ERR? and ERR? was increased in the mouse kidney after fasting. By using immunohistochemistry (IHC), we showed these three proteins are co-localized in the outer stripe of the outer medulla (OSOM) of the mouse kidney. We were able to collect this region from the kidney using the Laser Capture Microdissection (LCM) technique. The qPCR data showed significant increase of PGC-1?, ERR? and ERR? mRNA in the LCM samples after fasting for 24 hours. Furthermore, the known ERR? target genes, mitochondrial oxidative phosphorylation gene COX8H and the tricarboxylic acid (TCA) cycle gene IDH3A also showed an increase. Taken together, our data suggest that fasting activates the energy balance program in the OSOM of the kidney.
Project description:GSK5182 (4) is currently one of the lead compounds for the development of estrogen-related receptor gamma (ERR?) inverse agonists. Here, we report the design, synthesis, pharmacological and in vitro absorption, distribution, metabolism, excretion, toxicity (ADMET) properties of a series of compounds related to 4. Starting from 4, a series of analogs were structurally modified and their ERR? inverse agonist activity was measured. A key pharmacophore feature of this novel class of ligands is the introduction of a heterocyclic group for A-ring substitution in the core scaffold. Among the tested compounds, several of them are potent ERR? inverse agonists as determined by binding and functional assays. The most promising compound, 15g, had excellent binding selectivity over related subtypes (IC50 = 0.44, >10, >10, and 10 ?M at the ERR?, ERR?, ERR?, and ER? subtypes, respectively). Compound 15g also resulted in 95% transcriptional repression at a concentration of 10 ?M, while still maintaining an acceptable in vitro ADMET profile. This novel class of ERR? inverse agonists shows promise in the development of drugs targeting ERR?-related diseases.