Structure-activity studies on 1,3-dioxane-2-carboxylic acid derivatives, a novel class of subtype-selective peroxisome proliferator-activated receptor alpha (PPARalpha) agonists

A series of 1,3-dioxane carboxylic acid derivatives was synthesized and evaluated for human PPAR transactivation activity. Structure-activity relationships on the phenyloxazole moiety of the lead compound 3 revealed that the introduction of small hydrophobic substituents at the 4-position of the terminal phenyl ring increased the PPARalpha agonist activity, and that the oxazole heterocycle was essential to the maintenance of both potency and PPARalpha subtype-selectivity. This investigation led to the identification of 14d (NS-220) and 14i as highly potent and selective human PPARalpha agonists. In KK-A(y) type 2 diabetic mice, these compounds significantly lowered plasma triglyceride and very-low-density plus low-density lipoprotein cholesterol levels while simultaneously raising HDL cholesterol levels. Our results suggest that highly potent and subtype-selective PPARalpha agonists will be promising drugs for the treatment of metabolic disorders in type 2 diabetes.     

Design and synthesis of novel oxazole containing 1,3-dioxane-2-carboxylic acid derivatives as PPAR alpha/gamma dual agonists

A few novel 1,3-dioxane carboxylic acid derivatives were designed and synthesized to aid in the characterization of PPAR alpha/gamma dual agonists. Structural requirements for PPARalpha/gamma dual agonism of 1,3-dioxane carboxylic acid derivatives included the structural similarity with potent glitazones in fibric acid chemotype. The compounds with this pharmacophore and substituted oxazole as a lipophilic heterocyclic tail were synthesized and evaluated for their in vitro PPAR agonistic potential and in vivo hypoglycemic and hypolipidemic efficacy in animal models. Lead compound 2-methyl-c-5-[4-(5-methyl-2-(4-methylphenyl)-oxazol-4-ylmethoxy)-benzyl]-1,3-dioxane-r-2-carboxylic acid 13b exhibited potent hypoglycemic, hypolipidemic and insulin sensitizing effects in db/db mice and Zucker fa/fa rats.     

The peroxisome proliferator-activated receptor alpha (PPARalpha) regulates bile acid biosynthesis

Fibrates are a group of hypolipidemic agents that efficiently lower serum triglyceride levels by affecting the expression of many genes involved in lipid metabolism. These effects are exerted via the peroxisome proliferator-activated receptor alpha (PPARalpha). In addition, fibrates also lower serum cholesterol levels, suggesting a possible link between the PPARalpha and cholesterol metabolism. Bile acid formation represents an important pathway for elimination of cholesterol, and the sterol 12alpha-hydroxylase is a branch-point enzyme in the bile acid biosynthetic pathway, which determines the ratio of cholic acid to chenodeoxycholic acid. Treatment of mice for 1 week with the peroxisome proliferator WY-14,643 or fasting for 24 h both induced the sterol 12alpha-hydroxylase mRNA in liver. Using the PPARalpha knockout mouse model, we show that the induction by both treatments was dependent on the PPARalpha. A reporter plasmid containing a putative peroxisome proliferator-response element (PPRE) identified in the rat sterol 12alpha-hydroxylase promoter region was activated by treatment with WY-14,643 in HepG2 cells, being dependent on co-transfection with a PPARalpha expression plasmid. The rat 12alpha-hydroxylase PPRE bound in vitro translated PPARalpha and retinoid X receptor alpha, albeit weakly, in electrophoretic mobility shift assay. Treatment of wild-type mice with WY-14,643 for 1 week resulted in an increased relative amount of cholic acid, an effect that was abolished in the PPARalpha null mice, verifying the functionality of the PPRE in vivo.     

Design and synthesis of substituted phenylpropanoic acid derivatives as human peroxisome proliferator-activated receptor alpha/delta dual agonists

A series of phenylpropanoic acids was prepared as candidate dual agonists of peroxisome proliferator-activated receptors (PPAR) alpha and delta. Structure-activity relationship studies indicated that the shape of the linker moiety and the nature of the substituent at the distal benzene ring play key roles in determining the potency and selectivity of PPAR subtype transactivation. Optically active alpha-ethylphenylpropanoic acid derivatives were identified as potent human PPAR alpha and delta dual agonists with potential for the treatment of metabolic syndrome.     

Effects of peroxisome proliferator-activated receptor alpha (PPARalpha) agonists on leucine-induced phosphorylation of translational targets in C2C12 cells

Effect of peroxisome proliferator-activated receptor alpha (PPARalpha) agonists, WY-14,643 (WY) and/or clofibrate, on the leucine-induced phosphorylation of translational targets in C2C12 myoblasts was studied. C2C12 cells were treated with WY or clofibrate for 24 h prior to stimulation with leucine. Western blot analyses revealed that the leucine-induced phosphorylation of p70 S6 kinase (p70S6K), a key regulator of translation initiation, was significantly higher in WY-treated cells than in control and clofibrate-treated cells. Phosphorylation of extracellular-regulated kinase (ERK1/2) was higher in WY-treated cells. WY treatment also increased the leucine-induced phosphorylation of ribosomal protein S6 and eukaryotic initiation factor 4B. In contrast, eukaryotic elongation factor 2, a marker for peptide chain elongation process, was significantly activated (dephosphorylated) only in leucine-stimulated control cells. Pre-treatment of the cells with PD98059 (ERK1/2 kinase inhibitor) prevented the phosphorylation of ERK1/2 and decreased the leucine-induced phosphorylation of p70S6K. It is concluded that WY increased the leucine-induced phosphorylation of target proteins involving in translation initiation via ERK/p70S6K pathway, but impaired the signaling for elongation process, suggesting that p70S6K phosphorylation may be essential, but not sufficient for the activation of entire targets for protein translation in WY-treated cells.     

Expression and purification of the ligand-binding domain of peroxisome proliferator-activated receptor alpha (PPARalpha)

We describe the expression and purification of the C-terminally His-tagged ligand-binding domain of the peroxisome proliferator-activated receptor alpha (PPARα-LBD). Using a modified expression and purification strategy a five times higher protein yield was achieved compared to existing protocols. In summary, a modified Escherichia coli strain was used, which allows rare codons to be expressed more efficiently, and moreover, conditions for cell growth and cell lysis were improved. Protein purification was achieved in a two-step approach using nickel affinity chromatography followed by anion exchange chromatography. The identity of PPARα-LBD was confirmed by online-nano-high performance liquid chromatography/matrix-assisted laser desorption/ionization time-of-flight/time-of-flight mass spectrometry (nano-HPLC/MALDI-TOF/TOF-MS).     

Pancreatic islet response to hyperglycemia is dependent on peroxisome proliferator-activated receptor alpha (PPARalpha)

This study tests the hypothesis that islet peroxisome proliferator-activated receptor alpha (PPARalpha) influences insulin secretion. Freshly isolated islets of normoglycemic PPARalpha-null mice display no major alteration of glucose-stimulated insulin release. However, after 24 h of culture in high glucose, PPARalpha-null islets exhibit elevated basal insulin secretion and fail to increase insulin mRNA. 24-h culture with palmitate replicates this phenotype in wild-type islets. The data suggest that PPARalpha is needed to ensure appropriate insulin secretory response in situation of short-term hyperglycemia, likely by maintaining islet lipid homeostasis. As such, islet PPARalpha could contribute to delay the progression of type 2 diabetes.     

Fatty acid regulation of liver X receptors (LXR) and peroxisome proliferator-activated receptor alpha (PPARalpha ) in HEK293 cells

Fatty acids bind to and regulate the activity of peroxisome proliferator-activated (PPAR) and liver X receptors (LXR). However, the role lipid metabolism plays in the control of intracellular fatty acid ligands is poorly understood. We have identified two strains of HEK293 cells that display differences in fatty acid regulation of nuclear receptors. Using full-length and Gal4-LBD chimeric receptors in functional assays, 20:4,n6 induced PPARalpha activity approximately 2.2-fold and suppressed LXRalpha activity by 80% (ED50 approximately 25-50 microm) in HEK293-E (early passage) cells but had no effect on PPARalpha or LXRalpha receptor activity in HEK293-L (late passage) cells. LXRbeta was insensitive to fatty acid regulation in both HEK293 strains. Metabolic labeling studies using [14C]20:4,n6 (at 100 microm) indicated that the uptake of 20:4,n6 and its assimilation into triacylglycerol, diacylglycerol, and polar lipids revealed no difference between the two strains. Such treatment increased total cellular 20:4,n6 ( approximately 11-fold) and its elongation product, 22:4,n6 ( approximately 3.6-fold), within 6 h. Non-esterified 20:4,n6 and 22:4,n6 represented 相似文献   

Concise and efficient asymmetric synthesis of (S)-2-ethylphenylpropanoic acid derivatives: dual agonists for human peroxisome proliferator-activated receptor alpha and delta

Optically active (S)-2-ethylphenylpropanoic acid derivatives, dual agonists for human peroxisome proliferator-activated receptor (PPAR) alpha and delta, were efficiently prepared by using Evan's chiral oxazolidinone technique and reductive amide N-alkylation as key steps.     

The discovery of novel isoflavone pan peroxisome proliferator-activated receptor agonists

Twenty three dual PPARα and γ molecules of natural product origin, previously reported by our group, were further investigated for pan PPAR transactivation against PPARδ. The in vitro cell toxicity profile, as well as, in silico study of the most active molecules within this new class of pan PPAR agonists are also described. 3′,5′ Dimethoxy-7 hydroxyisoflavone 6, Ψ-baptigenin 7, 4′ fluoro-7 hydroxyisoflavone 8, and 3′ methoxy-7 hydroxyisoflavone 9 were identified as the most potent molecules studied within the set compared to the commercially available pan PPAR agonist, bezafibrate 1. These novel active molecules may thus be useful as future leads in PPAR-related disorders, including type II diabetes mellitus and metabolic syndrome.     

Molecular cloning and characterization of two mouse peroxisome proliferator-activated receptor alpha (PPARalpha)-regulated peroxisomal acyl-CoA thioesterases

Peroxisomes are organelles that function in the beta-oxidation of long- and very long-chain acyl-CoAs, bile acid-CoA intermediates, prostaglandins, leukotrienes, thromboxanes, dicarboxylic fatty acids, pristanic acid, and xenobiotic carboxylic acids. The very long- and long-chain acyl-CoAs are mainly chain-shortened and then transported to mitochondria for further metabolism. We have now identified and characterized two peroxisomal acyl-CoA thioesterases, named PTE-Ia and PTE-Ic, that hydrolyze acyl-CoAs to the free fatty acid and coenzyme A. PTE-Ia and PTE-Ic show 82% sequence identity at the amino acid level, and a putative peroxisomal type 1 targeting signal of -AKL was identified at the carboxyl-terminal end of both proteins. Localization experiments using green fluorescent fusion protein showed PTE-Ia and PTE-Ic to be localized in peroxisomes. Despite their high level of sequence identity, we show that PTE-Ia is mainly active on long-chain acyl-CoAs, whereas PTE-Ic is mainly active on medium-chain acyl-CoAs. Lack of regulation of enzyme activity by free CoASH suggests that PTE-Ia and PTE-Ic regulate intraperoxisomal levels of acyl-CoA, and they may have a function in termination of beta-oxidation of fatty acids of different chain lengths. Tissue expression studies revealed that PTE-Ia is highly expressed in kidney, whereas PTE-Ic is most highly expressed in spleen, brain, testis, and proximal and distal intestine. Both PTE-Ia and PTE-Ic were highly up-regulated in mouse liver by treatment with the peroxisome proliferator WY-14,643 and by fasting in a peroxisome proliferator-activated receptor alpha-dependent manner. These data show that PTE-Ia and PTE-Ic have different functions based on different substrate specificities and tissue expression.     

Design, synthesis and evaluation of trifluoromethane sulfonamide derivatives as new potent and selective peroxisome proliferator-activated receptor alpha agonists

Starting from the structure of 5, a two-step strategy was applied to identify a new generation of trifluoromethane sulfonamides as potent PPARalpha agonists. Synthesis, in vitro and in vivo evaluation of the most potent compound are reported.     

Design, synthesis, and evaluation of a novel series of alpha-substituted phenylpropanoic acid derivatives as human peroxisome proliferator-activated receptor (PPAR) alpha/delta dual agonists for the treatment of metabolic syndrome

A series of alpha-alkyl-substituted phenylpropanoic acids was prepared as dual agonists of peroxisome proliferator-activated receptors alpha and delta (PPARalpha/delta). Structure-activity relationship studies indicated that the shape of the linking group and the shape of the substituent at the distal benzene ring play key roles in determining the potency and the selectivity of PPAR subtype transactivation. Structure-activity relationships among the amide series (10) and the reversed amide series (13) are similar, but not identical, especially in the case of the compounds bearing a bulky hydrophobic substituent at the distal benzene ring, indicating that the hydrophobic tail part of the molecules in these two series binds at somewhat different positions in the large binding pocket of PPAR. alpha-Alkyl-substituted phenylpropanoic acids of (S)-configuration were identified as potent human PPARalpha/delta dual agonists. Representative compounds exhibited marked nuclear receptor selectivity for PPARalpha and PPARdelta. Subtype-selective PPAR activation was also examined by analysis of the mRNA expression of PPAR-regulated genes.