Substituted benzyloxytricyclic compounds as retinoic acid-related orphan receptor gamma t (RORγt) agonists

Substituted benzyloxy aryl compound 2 was identified as an RORγt agonist. Structure based drug design efforts resulted in a potent and selective tricyclic compound 19 which, when administered orally in an MC38 mouse tumor model, demonstrated a desired pharmacokinetic profile as well as a dose-dependent pharmacodynamic response. However, no perceptible efficacy was observed in this tumor model at the doses investigated.     

Identification of bicyclic hexafluoroisopropyl alcohol sulfonamides as retinoic acid receptor-related orphan receptor gamma (RORγ/RORc) inverse agonists. Employing structure-based drug design to improve pregnane X receptor (PXR) selectivity

We disclose the optimization of a high throughput screening hit to yield benzothiazine and tetrahydroquinoline sulfonamides as potent RORγt inverse agonists. However, a majority of these compounds showed potent activity against pregnane X receptor (PXR) and modest activity against liver X receptor α (LXRα). Structure-based drug design (SBDD) led to the identification of benzothiazine and tetrahydroquinoline sulfonamide analogs which completely dialed out LXRα activity and were less potent at PXR. Pharmacodynamic (PD) data for compound 35 in an IL-23 induced IL-17 mouse model is discussed along with the implications of a high Y_max in the PXR assay for long term preclinical pharmacokinetic (PK) studies.     

Annulation reaction enables the identification of an exocyclic amide tricyclic chemotype as retinoic acid Receptor-Related orphan receptor gamma (RORγ/RORc) inverse agonists

RORγt is the master regulator of the IL-23/IL-17 axis, a pathway that is clinically validated for the treatment of various immunological disorders. Over the last few years, our group has reported different chemotypes that potently act as inverse agonists of RORγt. One of them, the tricyclic pyrrolidine chemotype, has demonstrated biologic-like preclinical efficacy and has led to our clinical candidate BMS-986251. In this letter, we discuss the invention of an annulation reaction which enabled the synthesis of a tricyclic exocyclic amide chemotype and the identification of compounds with RORγt inverse agonist activity. Preliminary structure activity relationships are disclosed.     

The retinoic acid receptor-related orphan nuclear receptor γ1 (RORγ1): a novel player determinant of insulin sensitivity in morbid obesity

The orphan nuclear receptors (ONRs), retinoic acid receptor-related orphan receptor γ-1 (RORγ1) and peroxisome proliferator-activated receptor γ-2 (PPARγ2), are central mediators controlling adipocyte (AD) differentiation. Through their distinct tissue distribution and specific target gene activation, ONRs control diverse aspects of fatty acid metabolism and insulin sensitivity. Adding further complexity, obesity begets resistance to insulin signals and can ultimately result in diabetes. In this study, we investigate whether there are differences in the RORγ1 and PPARγ2 expression in visceral adipose tissue (VAT) and subcutaneous adipose tissue (SAT) from morbid obesity (MO) individuals either insulin resistant (high-IR MO) or insulin sensitivity (low-IR MO). Our results indicate for the first time in human the RORγ1 mRNA and protein expression levels and activation with coactivator, such as peroxisome proliferator-activated receptor γ coactivator 1-α (PGC-1α) were higher in the VAT from high-IR MO. In contrast, PPARγ2 expression and activation were higher in the VAT from low-IR MO. In this way, we have also found a positive association between RORγ1 mRNA and protein expression with many components of metabolic syndrome, with a strong dependence of insulin and HOMA(IR) index in VAT, but not in SAT. Our data suggest that RORγ1 may be added to the growing list of nuclear receptors in adipose tissue use to modulate the insulin resistance associated to the obesity. Measurement of RORγ1 and PPARγ2 in adipose tissue might be useful for evaluating the outcomes of various clinical interventions for obesity-related diabetes type II.     

Structural development of tetrachlorophthalimides as liver X receptor β (LXRβ)-selective agonists with improved aqueous solubility

LXRβ-selective agonists are promising candidates to improve atherosclerosis without increasing plasma or hepatic TG levels. We have reported a series of tetrachlorophthalimide analogs as an LXRβ-selective agonist. However, they exhibited poor aqueous solubility probably due to its high hydrophobicity and highly rigid and plane structure. In this report, we present further structural development of tetrachloro(styrylphenyl)phthalimides as the LXRβ-selective agonists with improved aqueous solubility.     

Identification of pyridazin-3-one derivatives as potent, selective histamine H₃ receptor inverse agonists with robust wake activity

H(3)R structure-activity relationships on a novel class of pyridazin-3-one H(3)R antagonists/inverse agonists are disclosed. Modifications of the pyridazinone core, central phenyl ring and linker led to the identification of molecules with excellent target potency, selectivity and pharmacokinetic properties. Compounds 13 and 21 displayed potent functional H(3)R antagonism in vivo in the rat dipsogenia model and demonstrated robust wake activity in the rat EEG/EMG model.     

Structural development of 1H-pyrazolo-[3,4-b]pyridine-4-carboxylic acid derivatives as human peroxisome proliferator-activated receptor alpha (PPARα)-selective agonists

We previously reported that 1H-pyrazolo-[3,4-b]pyridine-4-carboxylic acid derivative 6 is an agonist of human peroxisome proliferator-activated receptor alpha (hPPARα). Here, we prepared a series of 1H-pyrazolo-[3,4-b]pyridine-4-carboxylic acid derivatives in order to examine the structure-activity relationships (SAR). SAR studies clearly indicated that the steric bulkiness of the substituent on 1H-pyrazolo-[3,4-b]pyridine ring, the position of the distal hydrophobic tail part, and the distance between the distal hydrophobic tail part and the acidic head part are critical for hPPARα agonistic activity. These SAR results are somewhat different from those reported for fibrate-class hPPARα agonists. A representative compound (10f) was as effective as fenofibrate in reducing the elevated plasma triglyceride levels in a high-fructose-fed rat model.     

Discovery and SAR of cinnolines/quinolines as liver X receptor (LXR) agonists with binding selectivity for LXRβ

A series of cinnolines/quinolines was prepared and it was found that 4-phenyl-cinnoline/quinolines with either a 2′,3′ or 2′,5′-disubstituted benzyloxy moiety or the 1-Me-7-indole methoxy moiety on the meta position of the 4-phenyl ring showed good binding selectivity for LXRβ over LXRα. The LXRβ binding selective modulators displayed good activity for inducing ABCA1 gene expression in J774 macrophage cell line and poor efficacy in the LXRα Gal4 functional assay. 26, 37 and 41 were examined for their ability to induce SREBP-1c gene expression in Huh-7 liver cell line and they were weak partial agonists.     

Design and synthesis of a series of α-benzyl phenylpropanoic acid-type peroxisome proliferator-activated receptor (PPAR) gamma partial agonists with improved aqueous solubility

In the continuing study directed toward the development of peroxisome proliferator-activated receptor gamma (hPPARγ) agonist, we attempted to improve the water solubility of our previously developed hPPARγ-selective agonist 3, which is insufficiently soluble for practical use, by employing two strategies: introducing substituents to reduce its molecular planarity and decreasing its hydrophobicity via replacement of the adamantyl group with a heteroaromatic ring. The first approach proved ineffective, but the second was productive. Here, we report the design and synthesis of a series of α-benzyl phenylpropanoic acid-type hPPARγ partial agonists with improved aqueous solubility. Among them, we selected (R)-7j, which activates hPPARγ to the extent of about 65% of the maximum observed with a full agonist, for further evaluation. The ligand-binding mode and the reason for the partial-agonistic activity are discussed based on X-ray-determined structure of the complex of hPPARγ ligand-binding domain (LBD) and (R)-7j with previously reported ligand-LDB structures. Preliminal apoptotic effect of (R)-7j against human scirrhous gastric cancer cell line OCUM-2MD3 is also described.     

Quinoline-3-carboxamide containing sulfones as liver X receptor (LXR) agonists with binding selectivity for LXRβ and low blood–brain penetration

A series of quinoline-3-carboxamide containing sulfones was prepared and found to have good binding affinity for LXRβ and moderate binding selectivity over LXRα. The 8-Cl quinoline analog 33 with a high TPSA score, displayed 34-fold binding selectivity for LXRβ over LXRα (LXRβ IC₅₀ = 16 nM), good activity for inducing ABCA1 gene expression in a THP macrophage cell line, desired weak potency in the LXRα Gal4 functional assay, and low blood–brain barrier penetration in rat.     

Peroxisome proliferator-activated receptor γ (PPARγ) plays a critical role in the development of TGFβ resistance of H460 cell

The primary goal of the study was to investigate how peroxisome proliferator-activated receptor γ (PPARγ) played a critical role in the protection of H460 cell, one of the non-small cell lung cancer (NSCLC) cells with multidrug resistance, from transforming growth factor β (TGFβ)-mediated mitoinhibition. In the study, TGFβ resistance of H460 cell was first confirmed by analyses of PPARγ expression, its interaction with TGFβ-induced Smad3 and phospho-Smad3 (p-Smad3) and survival of H460. Results showed that enable to escape from G2/M phase arrest, H460 cell had higher resistance to TGFβ-mediated mitoinhibition than CH27 (a drug sensitive control). TGFβ significantly increased PPARγ expression of H460 but not of CH27 cell whereas nuclear accumulation of p-Smad3 was only limited to CH27, the latter was believed to contribute to the induction of P^{21 waf1/cip1} and cyclin B1, cell cycle arrest at G2/M phase and TGFβ-mediated mitoinhibition of CH27 cell. TGFβ-induced PPARγ of H460 cell was further demonstrated to bind to Smad3 and p-Smad3, and GW9662 (PPARγ inhibitor) or PPARγ-specific shRNA could disrupt the binding. GW9662 also increased the nuclear accumulation of p-Smad3 that eventually led to the reduction of TGFβ resistance of H460. A transient knockdown of PPARγ with shRNA revealed a similar effect as GW9662. In addition, activation of P³⁸ instead of ERK played a critical role in TGFβ-induced expression of PPARγ, which subsequently activated RhoA in H460 cell.     

The C161T polymorphism in the peroxisome proliferator-activated receptor gamma gene (PPARγ) is associated with risk of coronary artery disease: a meta-analysis

The researches attempting to associate the PPARγ C161T polymorphism with coronary artery disease (CAD) yielded complicated and contradictory results. We aimed for more precise estimate of the relationship and conducted a comprehensive meta-analysis. Publications written in English or Chinese were screened in MEDLINE, Embase, CNKI, Wanfang and CBM. Data on 11 studies including 3,020 cases and 2,853 controls were extracted. A random-effects model was available to synthesize the inconsistent outcomes of the individual studies, while addressing between-study heterogeneity and publication bias. The PPARγ C161T polymorphism followed Hard-Weinberg Equilibrium for all studies (P > 0.05).Overall, there was no evidence for a significant association under all genetic models but with distinct heterogeneity (T vs. C: P = 0.29, OR = 0.91, 95 %CI 0.77–1.08, P _{heterogeneity} = 0.004, I ² = 61.2 %). However, in the subgroup analysis by ethnicity, the T allele carriers showed a prominent 26 % risk reduction of CAD among Chinese (dominant genetic model: P = 0.03, 95 %CI 0.57–0.97, P _{heterogeneity} = 0.03, I ² = 56.1 %). After dividing into population source, the significance of CAD risk reduction was strengthened in hospital-based studies (allele comparison: P = 0.04, OR = 0.82, 95 %CI 0.67–1.00, P _{heterogeneity} = 0.04, I ² = 52.5 %; dominant model: P = 0.01, OR = 0.73, 95 %CI 0.57–0.92, P _{heterogeneity} = 0.05, I ² = 50.8 %). There was no obvious publication bias verified in the method of funnel plot and Egger’s linear regression test (t = ?0.11, P = 0.913). Taken together, our results revealed the PPARγ C161T polymorphism might play a moderate protective effect on developing CAD among Chinese, but not among Caucasians.     

Cladosporols A and B,two natural peroxisome proliferator-activated receptor gamma (PPARγ) agonists,inhibit adipogenesis in 3T3-L1 preadipocytes and cause a conditioned-culture-medium-dependent arrest of HT-29 cell proliferation

BackgroundObesity and type 2 diabetes mellitus, which are widespread throughout the world, require therapeutic interventions targeted to solve clinical problems (insulin resistance, hyperglycaemia, dyslipidaemia and steatosis). Several natural compounds are now part of the therapeutic repertoire developed to better manage these pathological conditions. Cladosporols, secondary metabolites from the fungus Cladosporium tenuissimum, have been characterised for their ability to control cell proliferation in human colon cancer cell lines through peroxisome proliferator-activated receptor gamma (PPARγ)-mediated modulation of gene expression. Here, we report data concerning the ability of cladosporols to regulate the differentiation of murine 3T3-L1 preadipocytes.MethodsCell counting and MTT assay were used for analysing cell proliferation. RT-PCR and Western blotting assays were performed to evaluate differentiation marker expression. Cell migration was analysed by wound-healing assay.ResultsWe showed that cladosporol A and B inhibited the storage of lipids in 3T3-L1 mature adipocytes, while their administration did not affect the proliferative ability of preadipocytes. Moreover, both cladosporols downregulated mRNA and protein levels of early (C/EBPα and PPARγ) and late (aP2, LPL, FASN, GLUT-4, adiponectin and leptin) differentiation markers of adipogenesis. Finally, we found that proliferation and migration of HT-29 colorectal cancer cells were inhibited by conditioned medium from cladosporol-treated 3T3-L1 cells compared with the preadipocyte conditioned medium.ConclusionsTo our knowledge, this is the first report describing that cladosporols inhibit in vitro adipogenesis and through this inhibition may interfere with HT-29 cancer cell growth and migration.General significanceCladosporols are promising tools to inhibit concomitantly adipogenesis and control colon cancer initiation and progression.     

Purification of a new glutathione S-transferase (transferase μ) from human liver having high activity with benzo(α)pyrene-4,5-oxide

A new glutathione $\text{S}$-transferase from human liver has been purified to homogeneity in good yield by use of ion-exchange chromatography on DEAE-cellulose, affinity chromatography on $\text{S}$-hexylglutathione coupled to epoxy-activated Sepharose 6B, and chromatography on hydroxyapatite. This new enzyme, transferase μ, is present in high concentration, but only in some individuals. It has an isoelectric point at about pH 6 to 6.5 and a different substrate specificity than the previously described alkaline transferases α-ε from human liver. Especially noteworthy is the finding of high activity against benzo(α)pyrene-4,5-oxide. Glutathione $\text{S}$-transferase μ has about 20-fold higher activity with this substrate than have the alkaline transferases. The most pronounced difference was found with $\text{trans}$-4-phenyl-3-buten-2-one which was >100-fold better as substrate for transferase μ than for the previously described transferases.