A peroxisome proliferator-activated receptor γ ligand inhibits adipocyte differentiation

The peroxisome proliferator-activated receptors (PPARs) are nuclear hormone receptors that regulate glucose and lipid homeostasis. The PPARγ subtype plays a central role in the regulation of adipogenesis and is the molecular target for the 2,4-thiazolidinedione class of antidiabetic drugs. Structural studies have revealed that agonist ligands activate the PPARs through direct interactions with the C-terminal region of the ligand-binding domain, which includes the activation function 2 helix. GW0072 was identified as a high-affinity PPARγ ligand that was a weak partial agonist of PPARγ transactivation. X-ray crystallography revealed that GW0072 occupied the ligand-binding pocket by using different epitopes than the known PPAR agonists and did not interact with the activation function 2 helix. In cell culture, GW0072 was a potent antagonist of adipocyte differentiation. These results establish an approach to the design of PPAR ligands with modified biological activities.

A selective peroxisome proliferator-activated receptor δ agonist promotes reverse cholesterol transport

The peroxisome proliferator-activated receptors (PPARs) are dietary lipid sensors that regulate fatty acid and carbohydrate metabolism. The hypolipidemic effects of the fibrate drugs and the antidiabetic effects of the glitazone drugs in humans are due to activation of the α (NR1C1) and γ (NR1C3) subtypes, respectively. By contrast, the therapeutic potential of the δ (NR1C2) subtype is unknown, due in part to the lack of selective ligands. We have used combinatorial chemistry and structure-based drug design to develop a potent and subtype-selective PPARδ agonist, GW501516. In macrophages, fibroblasts, and intestinal cells, GW501516 increases expression of the reverse cholesterol transporter ATP-binding cassette A1 and induces apolipoprotein A1-specific cholesterol efflux. When dosed to insulin-resistant middle-aged obese rhesus monkeys, GW501516 causes a dramatic dose-dependent rise in serum high density lipoprotein cholesterol while lowering the levels of small-dense low density lipoprotein, fasting triglycerides, and fasting insulin. Our results suggest that PPARδ agonists may be effective drugs to increase reverse cholesterol transport and decrease cardiovascular disease associated with the metabolic syndrome X.

Peroxisome proliferator-activated receptor alpha expression is regulated by estrogen receptor alpha and modulates the response of MCF-7 cells to sodium butyrate

Peroxisome proliferator-activated receptors are ligand-activated transcription factors with a potential role in cancer. We investigated peroxisome proliferator-activated receptor alpha expression in breast cancer cell lines and showed a relationship between mean peroxisome proliferator-activated receptor alpha and estrogen receptor alpha mRNA levels in estrogen receptor alpha positive breast cancer cells. Transfection of estrogen receptor alpha into the estrogen receptor alpha negative cell line, MDA-MB-231 decreased peroxisome proliferator-activated receptor a mRNA and conversely inhibition of estrogen receptor alpha by ICI-182 780 in estrogen receptor a positive, MCF-7 cells increased peroxisome proliferator-activated receptor a mRNA levels. Estrogen receptor alpha levels can be modulated by histone deacetylase inhibitors and such agents are in clinical trials for cancer treatment. We found the histone deacetylase inhibitor, sodium butyrate, increased peroxisome proliferator-activated receptor alpha mRNA levels within 4 h of treatment. Peroxisome proliferator-activated receptor a modulation was independent of estrogen receptor alpha, as a similar increase was observed in the estrogen receptor a negative MDA-MB-231 cells. To further investigate the relationship between sodium butyrate and peroxisome proliferator-activated receptor alpha expression, we created an MCF-7 cell line that conditionally over-expresses human peroxisome proliferator-activated receptor alpha. Over-expression of the peroxisome proliferator-activated receptor protected MCF-7 cells from sodium butyrate-mediated inhibition of proliferation and attenuated sodium butyrate-mediated induction of histone deacetylase 3 mRNA, indicating that elevated levels of peroxisome proliferator-activated receptor alpha may reduce the sensitivity of cells to histone deacetylase inhibitors. The estrogen receptor alpha dependence of peroxisome proliferator-activated receptor alpha levels may be significant since estrogen receptor alpha negative breast cancer cells are associated with a more aggressive phenotype. Our studies also suggest that peroxisome proliferator-activated receptor alpha levels may be a marker of breast cancer cell sensitivity to histone deacetylase inhibitors. (c) 2004 Elsevier Ltd. All rights reserved.

A new selective peroxisome proliferator-activated receptor gamma antagonist with antiobesity and antidiabetic activity.

Peroxisome proliferator-activated receptor gamma (PPAR-gamma) plays a key role in adipocyte differentiation and insulin sensitivity. Its synthetic ligands, the thiazolidinediones (TZD), are used as insulin sensitizers in the treatment of type 2 diabetes. These compounds induce both adipocyte differentiation in cell culture models and promote weight gain in rodents and humans. Here, we report on the identification of a new synthetic PPARgamma antagonist, the phosphonophosphate SR-202, which inhibits both TZD-stimulated recruitment of the coactivator steroid receptor coactivator-1 and TZD-induced transcriptional activity of the receptor. In cell culture, SR-202 efficiently antagonizes hormone- and TZD-induced adipocyte differentiation. In vivo, decreasing PPARgamma activity, either by treatment with SR-202 or by invalidation of one allele of the PPARgamma gene, leads to a reduction of both high fat diet-induced adipocyte hypertrophy and insulin resistance. These effects are accompanied by a smaller size of the adipocytes and a reduction of TNFalpha and leptin secretion. Treatment with SR-202 also dramatically improves insulin sensitivity in the diabetic ob/ob mice. Thus, although we cannot exclude that its actions involve additional signaling mechanisms, SR-202 represents a new selective PPARgamma antagonist that is effective both in vitro and in vivo. Because it yields both antiobesity and antidiabetic effects, SR-202 may be a lead for new compounds to be used in the treatment of obesity and type 2 diabetes.

Medium Chain Fatty Acids Are Selective Peroxisome Proliferator Activated Receptor (PPAR) gamma Activators and Pan-PPAR Partial Agonists

Thiazolidinediones (TZDs) act through peroxisome proliferator activated receptor (PPAR) gamma to increase insulin sensitivity in type 2 diabetes (T2DM), but deleterious effects of these ligands mean that selective modulators with improved clinical profiles are needed. We obtained a crystal structure of PPAR gamma ligand binding domain (LBD) and found that the ligand binding pocket (LBP) is occupied by bacterial medium chain fatty acids (MCFAs). We verified that MCFAs (C8-C10) bind the PPAR gamma LBD in vitro and showed that they are low-potency partial agonists that display assay-specific actions relative to TZDs; they act as very weak partial agonists in transfections with PPAR gamma LBD, stronger partial agonists with full length PPAR gamma and exhibit full blockade of PPAR gamma phosphorylation by cyclin-dependent kinase 5 (cdk5), linked to reversal of adipose tissue insulin resistance. MCFAs that bind PPAR gamma also antagonize TZD-dependent adipogenesis in vitro. X-ray structure B-factor analysis and molecular dynamics (MD) simulations suggest that MCFAs weakly stabilize C-terminal activation helix (H) 12 relative to TZDs and this effect is highly dependent on chain length. By contrast, MCFAs preferentially stabilize the H2-H3/beta-sheet region and the helix (H) 11-H12 loop relative to TZDs and we propose that MCFA assay-specific actions are linked to their unique binding mode and suggest that it may be possible to identify selective PPAR gamma modulators with useful clinical profiles among natural products.

Promoter rearrangements cause species-specific hepatic regulation of the glyoxylate reductase/hydroxypyruvate reductase gene by the peroxisome proliferator-activated receptor alpha.

In liver, the glyoxylate cycle contributes to two metabolic functions, urea and glucose synthesis. One of the key enzymes in this pathway is glyoxylate reductase/hydroxypyruvate reductase (GRHPR) whose dysfunction in human causes primary hyperoxaluria type 2, a disease resulting in oxalate accumulation and formation of kidney stones. In this study, we provide evidence for a transcriptional regulation by the peroxisome proliferator-activated receptor alpha (PPARalpha) of the mouse GRHPR gene in liver. Mice fed with a PPARalpha ligand or in which PPARalpha activity is enhanced by fasting increase their GRHPR gene expression via a peroxisome proliferator response element located in the promoter region of the gene. Consistent with these observations, mice deficient in PPARalpha present higher plasma levels of oxalate in comparison with their wild type counterparts. As expected, the administration of a PPARalpha ligand (Wy-14,643) reduces the plasma oxalate levels. Surprisingly, this effect is also observed in null mice, suggesting a PPARalpha-independent action of the compound. Despite a high degree of similarity between the transcribed region of the human and mouse GRHPR gene, the human promoter has been dramatically reorganized, which has resulted in a loss of PPARalpha regulation. Overall, these data indicate a species-specific regulation by PPARalpha of GRHPR, a key gene of the glyoxylate cycle.

Altered growth in male peroxisome proliferator-activated receptor gamma (PPARgamma) heterozygous mice: involvement of PPARgamma in a negative feedback regulation of growth hormone action.

The peroxisome proliferator-activated receptor gamma (PPARgamma) plays a major role in fat tissue development and physiology. Mutations in the gene encoding this receptor have been associated to disorders in lipid metabolism. A thorough investigation of mice in which one PPARgamma allele has been mutated reveals that male PPARgamma heterozygous (PPARgamma +/-) mice exhibit a reduced body size associated with decreased body weight, reflecting lean mass reduction. This phenotype is reproduced when treating the mice with a PPARgamma- specific antagonist. Monosodium glutamate treatment, which induces weight gain and alters body growth in wild-type mice, further aggravates the growth defect of PPARgamma +/- mice. The levels of circulating GH and that of its downstream effector, IGF-I, are not altered in mutant mice. However, the IGF-I mRNA level is decreased in white adipose tissue (WAT) of PPARgamma +/- mice and is not changed by acute administration of recombinant human GH, suggesting an altered GH action in the mutant animals. Importantly, expression of the gene encoding the suppressor of cytokine signaling-2, which is an essential negative regulator of GH signaling, is strongly increased in the WAT of PPARgamma +/- mice. Although the relationship between the altered GH signaling in WAT and reduced body size remains unclear, our results suggest a novel role of PPARgamma in GH signaling, which might contribute to the metabolic disorder affecting insulin signaling in PPARgamma mutant mice.

Role of genetic polymorphism peroxisome proliferator-activated receptor-gamma2 Pro12Ala on ethnic susceptibility to diabetes in South-Asian and Caucasian subjects: Evidence for heterogeneity

OBJECTIVE: To determine whether the peroxisome proliferator-activated receptor (PPAR)-gamma Pro12ala polymorphism modulates susceptibility to diabetes in South Asians. RESEARCH DESIGN AND METHODS: South Asians (n = 697) and Caucasians (n = 457) living in Dallas/Forth Worth, Texas, and South Asians living in Chennai, India (n = 1,619), were enrolled for this study. PPAR-gamma Pro12Ala was determined using restriction fragment-length polymorphism. Insulin responsiveness to an oral glucose tolerance test (OGTT) was measured in nondiabetic subjects. RESULTS: The Caucasian diabetic subjects had significantly lower prevalence of PPAR-gamma 12Ala when compared with the Caucasian nondiabetic subjects (20 vs. 9%, P = 0.006). However, there were no significant differences between diabetic and nondiabetic subjects with reference to the Pro12Ala polymorphism among the South Asians living in Dallas (20 vs. 23%) and in India (19 vs. 19.3%). Although Caucasians carrying PPAR-gamma Pro12Ala had lower plasma insulin levels at 2 h of OGTT than the wild-type (Pro/Pro) carriers (76 +/- 68 and 54 +/- 33 microU/ml, respectively, P = 0.01), no differences in either fasting or 2-h plasma insulin concentrations were found between South Asians carrying the PPAR-gamma Pro12Ala polymorphism and those with the wild-type genotype at either Chennai or Dallas. CONCLUSIONS: Although further replication studies are necessary to test the validity of the described genotype-phenotype relationship, our study supports the hypothesis that the PPAR-gamma Pro12Ala polymorphism is protective against diabetes in Caucasians but not in South Asians.