Pubblicazioni 2015
- Home
- Pubblicazioni 2015
1 – Di Leva FS, Festa C, Renga B, Sepe V, Novellino E, Fiorucci S, Zampella A, Limongelli V. Structure-based drug design targeting the cell membrane receptor GPBAR1: exploiting the bile acid scaffold towards selective agonism. Sci Rep. 2015 Nov 16;5:16605.
Abstract
Bile acids can regulate nutrient metabolism through the activation of the cell membrane receptor GPBAR1 and the nuclear receptor FXR. Developing an exogenous control over these receptors represents an attractive strategy for the treatment of enterohepatic and metabolic disorders. A number of dual GPBAR1/FXR agonists are known, however their therapeutic use is limited by multiple unwanted effects due to activation of the diverse downstream signals controlled by the two receptors. On the other hand, designing selective GPBAR1 and FXR agonists is challenging since the two proteins share similar structural requisites for ligand binding. Here, taking advantage of our knowledge of the two targets, we have identified through a rational drug design study a series of amine lithocholic acid derivatives as selective GPBAR1 agonists. The presence of the 3α-NH2 group on the steroidal scaffold is responsible for the selectivity over FXR unveiling unprecedented structural insights into bile acid receptors activity modulation.
2 – Renga B, Cipriani S, Carino A, Simonetti M, Zampella A, Fiorucci S. Reversal of Endothelial Dysfunction by GPBAR1 Agonism in Portal Hypertension Involves a AKT/FOXOA1 Dependent Regulation of H2S Generation and Endothelin-1. PLoS One. 2015 Nov 5;10(11):e0141082.
Abstract
Background
GPBAR1 is a bile acids activated receptor expressed in entero-hepatic tissues. In the liver expression of GPBAR1 is restricted to sinusoidal and Kuppfer cells. In the systemic circulation vasodilation caused by GPBAR1 agonists is abrogated by inhibition of cystathione-γ-liase (CSE), an enzyme essential to the generation of hydrogen sulfide (H2S), a vasodilatory agent. Portal BAR501 is a semisynthetic bile acid derivative endowed with a potent and selective agonistic activity toward GPBAR1.
Methods
Cirrhosis was induced in mice by carbon tetrachloride (CCL4) administration for 9 weeks. Liver endothelial dysfunction was induced by feeding wild type and Gpbar1-/- mice with methionine for 4 weeks. In both models, mice were administered BAR501, 15 mg/kg/day.
Results
By transactivation assay we demonstrate that BAR501 is a selective GPBAR1 agonist devoid of any FXR agonistic activity. In naïve rats, BAR501 effectively reduced hepatic perfusion pressure and counteracted the vasoconstriction activity of norepinephrine. In the CCl4 model, 9 weeks treatment with BAR501 effectively protected against development of endothelial dysfunction by increasing liver CSE expression and activity and by reducing endothelin (ET)-1 gene expression. In mice feed methionine, treatment with BAR501 attenuated endothelial dysfunction and caused a GPBAR1-dependent regulation of CSE. Using human liver sinusoidal cells, we found that modulation of CSE expression/activity is mediated by both genomic (recruitment of CREB to CRE in the CSE promoter) and non-genomic effects, involving a Akt-dependent phosporylation of CSE and endothelial nitric oxide (NO) synthase (eNOS). BAR501, phosphorylates FOXO1 and inhibits ET-1 transcription in liver sinusoidal cells.
Conclusions
BAR501, a UDCA-like GPBAR1 agonist, rescues from endothelial dysfunction in rodent models of portal hypertension by exerting genomic and non-genomic effects on CSE, eNOS and ET-1 in liver sinusoidal cells.
3 – Hodnik Ž, Tomašič T, Smodiš D, D’Amore C, Mašič LP, Fiorucci S, Kikelj D. Diethylstilbestrol-scaffold-based pregnane X receptor modulators. Eur J Med Chem. 2015 Oct 20;103:551-62.
Abstract
Due to its function as a regulator of drug-metabolizing enzymes and transporters, pregnane X receptor (PXR) represents an important factor involved in drug metabolism. In this work, we describe the discovery of diethylstilbestrol-based PXR modulators, which were designed from marine sulfated steroids with PXR agonistic activity, solomonsterols A and B, and our recently reported bazedoxifene scaffold-derived PXR antagonists. The methylated diethylstilbestrol derivative 1 displayed potent PXR agonistic activity with an EC50 value of 10.5 μM, whereas compounds 3, 4 and 6 (IC50 for 6 = 27.4 μM) and diethylstilbestrol (2) itself (IC50 = 14.6 μM) exhibited PXR antagonistic effects in HepG2 cells. The PXR modulatory effects of the synthesized diethylstilbestrol derivatives were further confirmed by the induction of PXR-regulated CYP3A4 expression with compound1, as well as by the inhibition of the rifaximin-promoted up-regulation of CYP3A4 expression with 2 and its derivative 6.
4 – Renga B, Francisci D, Carino A, Marchianò S, Cipriani S, Chiara Monti M, Del Sordo R, Schiaroli E, Distrutti E, Baldelli F, Fiorucci S. The HIV matrix protein p17 induces hepatic lipid accumulation via modulation of nuclear receptor transcriptoma. Sci Rep. 2015 Oct 15;5:15403
Abstract
Liver disease is the second most common cause of mortality in HIV-infected persons. Exactly how HIV infection per se affects liver disease progression is unknown. Here we have investigated mRNA expression of 49 nuclear hormone receptors (NRs) and 35 transcriptional coregulators in HepG2 cells upon stimulation with the HIV matrix protein p17. This viral protein regulated mRNA expression of some NRs among which LXRα and its transcriptional co-activator MED1 were highly induced at mRNA level. Dissection of p17 downstream intracellular pathway demonstrated that p17 mediated activation of Jak/STAT signaling is responsible for the promoter dependent activation of LXR. The treatment of both HepG2 as well as primary hepatocytes with HIV p17 results in the transcriptional activation of LXR target genes (SREBP1c and FAS) and lipid accumulation. These effects are lost in HepG2 cells pre-incubated with a serum from HIV positive person who underwent a vaccination with a p17 peptide as well as in HepG2 cells pre-incubated with the natural LXR antagonist gymnestrogenin. These results suggest that HIV p17 affects NRs and their related signal transduction thus contributing to the progression of liver disease in HIV infected patients.
5 – Sepe V, Distrutti E, Fiorucci S, Zampella A. Farnesoid X receptor modulators (2011 – 2014): a patent review. Expert Opin Ther Pat. 2015;25(8):885-96.
Abstract
Introduction: Farnesoid-X-receptor (FXR) is the receptor for primary bile acids expressed in enterohepatic tissues where it regulates bile acid uptake, metabolism and disposal. For its role as a bile acid sensor, FXR has been thought to be an important target in the treatment of cholestatic disorders, a family of diseases in which endogenous bile acids accumulate in the body. Cholestasis might occur as a consequence of inborn metabolic errors and three major disorders, intra-hepatic cholestasis in pregnancy, primary biliary cirrhosis (PBC) and primary sclerosing cholangitis account for the vast majority of clinical cholestasis occurring in adulthood. In addition, FXR agonists are gaining attention as potential regulators of lipid and glucose metabolism and therefore as new therapeutical approaches to the treatment of fatty liver disease, type 2 diabetes and obesity.
Areas covered: New chemical entities as FXR modulators and their in vitro and in vivoefficacy are reviewed with particular focus on patents and peer-reviewed publications in the period 2011 – 2014.
Expert opinion: FXR agonists have shown robust therapeutic potential and results from clinical trials have supported their use in the treatment of liver disorders including PBC and fatty liver disease despite side effects.
6 – Cipriani S, Renga B, D’Amore C, Simonetti M, De Tursi AA, Carino A, Monti MC, Sepe V, Zampella A, Fiorucci S. Impaired Itching Perception in Murine Models of Cholestasis Is Supported by Dysregulation of GPBAR1 Signaling. PLoS One. 2015 Jul 15;10(7):e0129866.
Abstract
Background & Aims
In cholestatic syndromes, body accumulation of bile acids is thought to cause itching. However, the mechanisms supporting this effect remain elusive. Recently, GPBAR1 (TGR5) a G-protein coupled receptor has been shown to mediate itching caused by intradermal administration of DCA and LCA. 6α-ethyl-3α, 7α-dihydroxy-24-nor-5β-cholan-23-ol (BAR502) is a non-bile acid dual ligand for FXR and GPBAR1.
Methods
Cholestasis was induced in wild type and GPBAR1-/- mice by administration of α-naphthyl-isothiocyanate (ANIT) or 17α-ethynylestradiol.
Results.
In naïve mice skin application of DCA, TLCA, 6-ECDCA, oleanolic and betulinic acid induces a GPBAR1 dependent pruritogenic response that could be desensitized by re-challenging the mice with the same GPBAR1 agonist. In wild type and GPBAR1-/- mice cholestasis induced by ANIT fails to induce spontaneous itching and abrogates scratching behavior caused by intradermal administration of DCA. In this model, co-treatment with BAR502 increases survival, attenuates serum alkaline phosphatase levels and robustly modulates the liver expression of canonical FXR target genes including OSTα, BSEP, SHP and MDR1, without inducing pruritus. Betulinic acid, a selective GPBAR1 ligand, failed to rescue wild type and GPBAR1-/- mice from ANIT cholestasis but did not induced itching. In the 17α-ethynylestradiol model BAR502 attenuates cholestasis and reshapes bile acid pool without inducing itching.
Conclusions
The itching response to intradermal injection of GPBAR1 agonists desensitizes rapidly and is deactivated in models of cholestasis, explain the lack of correlation between bile acids levels and itching severity in cholestatic syndromes. In models of non-obstructive cholestasis, BAR502 attenuates liver injury without causing itching.
7 – Renga B, Festa C, De Marino S, Di Micco S, D’Auria MV, Bifulco G, Fiorucci S, Zampella A. Molecular decodification of gymnemic acids from Gymnema sylvestre. Discovery of a new class of liver X receptor antagonists. Steroids. 2015 Apr;96:121-31.
Abstract
The individual chemical components of commercial extract of Gymnema sylvestre, a medicinal plant used in the traditional systems of the Indian medicine for its antidiabeticand hypolipidemic properties, were isolated and evaluated for their capability to act as modulators of nuclear and membrane receptors involved in glucose and lipidhomeostasis. The study disclosed for the first time that individual gymnemic acids are potent and selective antagonists for the β isoform of LXR. Indeed the above activity was shared by the most abundant aglycone gymnemagenin (10) whereas gymnestrogenin (11) was endowed with a dual LXRα/β antagonistic profile. Deep pharmacological investigation demonstrated that gymnestrogenin, reducing the expression of SREBP1c and ABCA1 in vitro, is able to decrease lipid accumulation in HepG2 cells. The results of this study substantiate the use of G. sylvestre extract in LXR mediated dislypidemic diseases.
8 – Talero E, Bolivar S, Ávila-Román J, Alcaide A, Fiorucci S, Motilva V. Inhibition of chronic ulcerative colitis-associated adenocarcinoma development in mice by VSL#3. Inflamm Bowel Dis. 2015 May;21(5):1027-37.
Abstract
BACKGROUND:
Colorectal cancer is the most severe complication in inflammatory bowel disease. This study aimed to investigate the effects of the probiotic VSL#3 when administered as either preventive or concurrent treatment in the progression from chronic colitis to colon cancer.
METHODS:
Mice were exposed to 5, 10, and 15 cycles of dextran sulfate sodium (DSS); each cycle consisted of 0.7% DSS for 1 week followed by distilled water for 10 days. VSL#3 was administered either from 2 weeks before the colitis induction or from the first day of the colitis until being killed. After each period, macroscopic and histological studies, as well as analysis of inflammatory and tumor biomarkers, were performed.
RESULTS:
Prophylactic or concurrent VSL#3 administration attenuated the disease activity index score and colon inflammation after 5, 10, and 15 cycles of DSS, as well as reduced the histological alterations and the incidence of colonic dysplastic lesions at the 3 periods studied. None of the animals receiving VSL#3 as a concurrent treatment developed carcinoma, which is in contrast to 5% and 20% of the mice following preventiveVSL#3 administration, developing carcinoma at the 10th and the 15th cycles of DSS, respectively. In addition, the probiotic reduced the proliferating cell nuclear antigen labeling index, tumor necrosis factor alpha, interleukin-1β, interleukin-6 production, cyclooxygenase-2 expression, and increased interleukin-10 levels in colon tissue at the 3 periods assayed.
CONCLUSIONS:
VSL#3 administration reduced chronic inflammation and prevented or delayed the development of dysplasia and carcinoma in a mouse model of chronic colitis-associated cancer.
9 – Distrutti E, Santucci L, Cipriani S, Renga B, Schiaroli E, Ricci P, Donini A, Fiorucci S. Bile acid activated receptors are targets for regulation of integrity of gastrointestinal mucosa. J Gastroenterol. 2015 Jul;50(7):707-19.
Abstract
Bile acids are the end product of cholesterol metabolism. Synthesized in the liver, primary bile acids are secreted by hepatocytes and are transformed by intestinal microbiota into secondary bile acids. In addition to their role in cholesterol and lipid absorption, bile acids act as signaling molecules activating a family of nuclear and G-protein-coupled receptors collectively known as bile acid activated receptors (BARs). These receptors are expressed at high density in enterohepatic tissues, but their expression occurs throughout the body and their activation mediates regulatory functions of bile acids on lipids and glucose metabolism and immunity. In the gastrointestinal tract, BARs maintain intestinal integrity, and their deletion makes the intestine more susceptible to the damage caused by acetylsalicylic acid and nonsteroidal anti-inflammatory drugs (NSAIDs). Deficiency in farnesoid X receptor and G-protein-coupled bile acid receptor 1 genes alters the expression/activity of cystathione γ-lyase and endothelial nitric oxide synthase, two genes involved in the synthesis of hydrogen sulfide and nitric oxide, i.e., two gaseous mediators that have been shown to be essential in maintaining the intestinal homeostasis. In addition, farnesoid X receptor regulates the expression of transporters required for secretion of phospholipid by hepatocytes. Because phospholids attenuate intestinal injury caused by acetylsalicylic acid and NSAIDs, BAR agonism could be exploited to protect the intestinal mucosa against injury caused by anti-inflammatory medications. This approach might be useful in the prevention of so-called NSAID enteropathy, a common clinical condition occurring in long-term users of NSAIDs, which is not effectively prevented either by cotreatment with proton pump inhibitors or by the use of coxibs.
10 – Fiorucci S, Distrutti E. Bile Acid-Activated Receptors, Intestinal Microbiota, and the Treatment of Metabolic Disorders. Trends Mol Med. 2015 Nov;21(11):702-14.
Abstract
The composition of the bile acid pool is a function of the microbial metabolism of bile acids in the intestine. Perturbations of the microbiota shape the bile acid pool and modulate the activity of bile acid-activated receptors (BARs) even beyond the gastrointestinal tract, triggering various metabolic axes and altering host metabolism. Bile acids, in turn, can also regulate the composition of the gut microbiome at the highest taxonomic levels. Primary bile acids from the host are preferential ligands for the farnesoid X receptor (FXR), while secondary bile acids from the microbiota are ligands for G-protein-coupled bile acid receptor 1 (GPBAR1). In this review, we examine the role of bile acid signaling in the regulation of intestinal microbiota and how changes in bile acid composition affect human metabolism. Bile acids may offer novel therapeutic modalities in inflammation, obesity, and diabetes.
11 – Sepe V, Distrutti E, Limongelli V, Fiorucci S, Zampella A. Steroidal scaffolds as FXR and GPBAR1 ligands: from chemistry to therapeutical application. Future Med Chem. 2015;7(9):1109-35.
Abstract
Bile acids (BAs) are experiencing a new life. Next to their ancestral roles in lipid digestion and solubilization, BAs are today recognized signaling molecules involved in many physiological functions. These signaling pathways involve the activation of metabolic nuclear receptors, mainly the BA sensor FXR, and the dedicated membrane G protein-coupled receptor, GPBAR1 (TGR5). As a consequence, the discovery of GPBAR1/FXR selective or dual modulators represents an important answer to the urgent demand of new pharmacological opportunity for several human diseases including dyslipidemia, cholestasis, nonalcoholic steatohepatitis, Type 2 diabetes and inflammation. Targeted oriented discovery of natural compounds and medicinal chemistry manipulation have allowed the development of promising drug candidates.
12 – Renga B, Bucci M, Cipriani S, Carino A, Monti MC, Zampella A, Gargiulo A, d’Emmanuele di Villa Bianca R, Distrutti E, Fiorucci S. Cystathionine γ-lyase, a H2S-generating enzyme, is a GPBAR1-regulated gene and contributes to vasodilation caused by secondary bile acids. Am J Physiol Heart Circ Physiol. 2015 Jul 1;309(1):H114-26.
Abstract
GPBAR1 is a bile acid-activated receptor (BAR) for secondary bile acids, lithocholic (LCA) and deoxycholic acid (DCA), expressed in the enterohepatic tissues and in the vasculature by endothelial and smooth muscle cells. Despite that bile acids cause vasodilation, it is unclear why these effects involve GPBAR1, and the vascular phenotype of GPBAR1 deficient mice remains poorly defined. Previous studies have suggested a role for nitric oxide (NO) in regulatory activity exerted by GPBAR1 in liver endothelial cells. Hydrogen sulfide (H2S) is a vasodilatory agent generated in endothelial cells by cystathionine-γ-lyase (CSE). Here we demonstrate that GPBAR1 null mice had increased levels of primary and secondary bile acids and impaired vasoconstriction to phenylephrine. In aortic ring preparations, vasodilation caused by chenodeoxycholic acid (CDCA), a weak GPBAR1 ligand and farnesoid-x-receptor agonist (FXR), was iberiotoxin-dependent and GPBAR1-independent. In contrast, vasodilation caused by LCA was GPBAR1 dependent and abrogated by propargyl-glycine, a CSE inhibitor, and by 5β-cholanic acid, a GPBAR1 antagonist, but not by N5-(1-iminoethyl)-l-ornithine (l-NIO), an endothelial NO synthase inhibitor, or iberiotoxin, a large-conductance calcium-activated potassium (BKCa) channels antagonist. In venular and aortic endothelial (HUVEC and HAEC) cells GPBAR1 activation increases CSE expression/activity and H2S production. Two cAMP response element binding protein (CREB) sites (CREs) were identified in the CSE promoter. In addition, TLCA stimulates CSE phosphorylation on serine residues. In conclusion we demonstrate that GPBAR1 mediates the vasodilatory activity of LCA and regulates the expression/activity of CSE. Vasodilation caused by CDCA involves BKCa channels. The GPBAR1/CSE pathway might contribute to endothelial dysfunction and hyperdynamic circulation in liver cirrhosis.
13 – Tsuchiya H, da Costa KA, Lee S, Renga B, Jaeschke H, Yang Z, Orena SJ, Goedken MJ, Zhang Y, Kong B, Lebofsky M, Rudraiah S, Smalling R, Guo G, Fiorucci S, Zeisel SH, Wang L. Interactions Between Nuclear receptor SHP and FOXA1 Maintain Oscillatory Homocysteine Homeostasis in Mice. Gastroenterology. 2015 May;148(5):1012-1023.e14.
Abstract
BACKGROUND & AIMS
Hyperhomocysteinemia is often associated with liver and metabolic diseases. We studied nuclear receptors that mediate oscillatory control of homocysteine homeostasis in mice.
METHODS
We studied mice with disruptions in Nr0b2 (called SHP-null mice) Bhmt, or both genes (BHMT-null/SHP-null mice), along with mice with wild-type copies of these genes (controls). Hyperhomocysteinemia was induced by feeding mice alcohol (the NIAAA binge model) or chow diets along with water containing 0.18% DL-homocysteine. Some mice were placed on diets containing cholic acid (1%) or cholestyramine (2%), or high-fat diets (60%). Serum and livers were collected over a 24 hr light–dark cycle and analyzed by RNA-seq, metabolomic, and quantitative PCR, immunoblot, and chromatin immunoprecipitation assays.
RESULTS
SHP-null mice had altered timing in expression of genes that regulate homocysteine metabolism, compared with control mice. Oscillatory production of S-adenosylmethionine, betaine, choline, phosphocholine, glyceophosphocholine, cystathionine, cysteine, hydrogen sulfide, glutathione disulfide, and glutathione, differed between SHP-null mice and control mice. SHP inhibited transcriptional activation of Bhmt and Cthby FOXA1. Expression of Bhmt and Cth was decreased when mice were fed cholic acid but increased when they were placed on diets containing cholestyramine or high-fat content. Diets containing ethanol or homocysteine induced hyperhomocysteinemia and glucose intolerance in control but not SHP-null mice. In BHMT-null and BHMT-null/SHP-null mice fed a control liquid, lipid vacuoles were observed in livers. Ethanol feeding induced accumulation of macrovesicular lipid vacuoles to the greatest extent in BHMT-null and BHMT-null/SHP-null mice.
CONCLUSIONS
Disruption of Shp in mice alters timing of expression of genes that regulate homocysteine metabolism and the liver responses to ethanol and homocysteine. SHP inhibits the transcriptional activation of Bhmt and Cth by FOXA1.