Dual PPARalpha /gamma activation provides enhanced improvement of insulin sensitivity and glycemic control in ZDF rats

Improvement of insulin sensitivity and lipid and glucose metabolism by coactivation of both nuclear peroxisome proliferator-activated receptor (PPAR)gamma and PPARalpha potentially provides beneficial effects over existing PPARgamma and alpha preferential drugs, respectively, in treatment of type 2 diabetes. We examined the effects of the dual PPARalpha/gamma agonist ragaglitazar on hyperglycemia and whole body insulin sensitivity in early and late diabetes stages in Zucker diabetic fatty (ZDF) rats and compared them with treatment with the PPARgamma preferential agonist rosiglitazone. Despite normalization of hyperglycemia and Hb A(1c) and reduction of plasma triglycerides by both compounds in both prevention and early intervention studies, ragaglitazar treatment resulted in overall reduced circulating insulin and improved insulin sensitivity to a greater extent than after treatment with rosiglitazone. In late-intervention therapy, ragaglitazar reduced Hb A(1c) by 2.3% compared with 1.1% by rosiglitazone. Improvement of insulin sensitivity caused by the dual PPARalpha/gamma agonist ragaglitazar seemed to have beneficial impact over that of the PPARgamma-preferential activator rosiglitazone on glycemic control in frankly diabetic ZDF rats.     

A peroxisome proliferator-activated receptor alpha/gamma dual agonist with a unique in vitro profile and potent glucose and lipid effects in rodent models of type 2 diabetes and dyslipidemia

LSN862 is a novel peroxisome proliferator-activated receptor (PPAR)alpha/gamma dual agonist with a unique in vitro profile that shows improvements on glucose and lipid levels in rodent models of type 2 diabetes and dyslipidemia. Data from in vitro binding, cotransfection, and cofactor recruitment assays characterize LSN862 as a high-affinity PPARgamma partial agonist with relatively less but significant PPARalpha agonist activity. Using these same assays, rosiglitazone was characterized as a high-affinity PPARgamma full agonist with no PPARalpha activity. When administered to Zucker diabetic fatty rats, LSN862 displayed significant glucose and triglyceride lowering and a significantly greater increase in adiponectin levels compared with rosiglitazone. Expression of genes involved in metabolic pathways in the liver and in two fat depots from compound-treated Zucker diabetic fatty rats was evaluated. Only LSN862 significantly elevated mRNA levels of pyruvate dehydrogenase kinase isozyme 4 and bifunctional enzyme in the liver and lipoprotein lipase in both fat depots. In contrast, both LSN862 and rosiglitazone decreased phosphoenol pyruvate carboxykinase in the liver and increased malic enzyme mRNA levels in the fat. In addition, LSN862 was examined in a second rodent model of type 2 diabetes, db/db mice. In this study, LSN862 demonstrated statistically better antidiabetic efficacy compared with rosiglitazone with an equivalent side effect profile. LSN862, rosiglitazone, and fenofibrate were each evaluated in the humanized apoA1 transgenic mouse. At the highest dose administered, LSN862 and fenofibrate reduced very low-density lipoprotein cholesterol, whereas, rosiglitazone increased very low-density lipoprotein cholesterol. LSN862, fenofibrate, and rosiglitazone produced maximal increases in high-density lipoprotein cholesterol of 65, 54, and 30%, respectively. These findings show that PPARgamma full agonist activity is not necessary to achieve potent and efficacious insulin-sensitizing benefits and demonstrate the therapeutic advantages of a PPARalpha/gamma dual agonist.     

4-(2H-1,2,3-benzotriazol-2-yl)phenoxy]alkanoic acids as agonists of peroxisome proliferator-activated receptors (PPARs)

A series of [4-(2H-1,2,3-benzotriazol-2-yl)phenoxy]alkanoic acids has been synthesized and tested as agonists of Peroxisome Proliferator-Activated Receptor (PPAR) alpha, gamma, and delta. Three compounds displayed 56 to 96% of maximal activity of the reference drug Wy-14643 on PPARalpha, and two of these, i.e., 1 and 5, exhibited also moderate activity on either PPARgamma or delta with efficacy equal to 50% and 46% of that of rosiglitazone and GW 501516, respectively. Thus, compounds 1 and 5 represent interesting starting point for preparing novel agents for the treatment of dyslipidemia or of dyslipidemic type-2 diabetes.     

Combination therapy with PPARgamma and PPARalpha agonists increases glucose-stimulated insulin secretion in db/db mice

Although peroxisome proliferator-activated receptor (PPAR)gamma agonists ameliorate insulin resistance, they sometimes cause body weight gain, and the effect of PPAR agonists on insulin secretion is unclear. We evaluated the effects of combination therapy with a PPARgamma agonist, pioglitazone, and a PPARalpha agonist, bezafibrate, and a dual agonist, KRP-297, for 4 wk in male C57BL/6J mice and db/db mice, and we investigated glucose-stimulated insulin secretion (GSIS) by in situ pancreatic perfusion. Body weight gain in db/db mice was less with KRP-297 treatment than with pioglitazone or pioglitazone + bezafibrate treatment. Plasma glucose, insulin, triglyceride, and nonesterified fatty acid levels were elevated in untreated db/db mice compared with untreated C57BL/6J mice, and these parameters were significantly ameliorated in the PPARgamma agonist-treated groups. Also, PPARgamma agonists ameliorated the diminished GSIS and insulin content, and they preserved insulin and GLUT2 staining in db/db mice. GSIS was further increased by PPARgamma and -alpha agonists. We conclude that combination therapy with PPARgamma and PPARalpha agonists may be more useful with respect to body weight and pancreatic GSIS in type 2 diabetes with obesity.     

WY14,643, a peroxisome proliferator-activated receptor alpha (PPARalpha ) agonist, improves hepatic and muscle steatosis and reverses insulin resistance in lipoatrophic A-ZIP/F-1 mice

WY14,643 is a specific peroxisome proliferator-activated receptor alpha (PPARalpha) agonist with strong hypolipidemic effects. Here we have examined the effect of WY14,643 in the A-ZIP/F-1 mouse, a model of severe lipoatrophic diabetes. With 1 week of treatment, all doses of WY14,643 that were tested normalized serum triglyceride and fatty acid levels. Glucose and insulin levels also improved but only with high doses and longer treatment duration. WY14,643 reduced liver and muscle triglyceride content and increased levels of mRNA encoding fatty acid oxidation enzymes. In liver, the elevated lipogenic mRNA profile (including PPARgamma) in A-ZIP/F-1 mice remained unchanged. These results suggest that WY14,643 acts by increasing beta-oxidation rather by than decreasing lipogenesis or lipid uptake. Hyperinsulinemic euglycemic clamp studies indicated that WY14,643 treatment improved liver more than muscle insulin sensitivity and that hepatic mRNA levels of gluconeogenic enzymes were reduced. Combination treatment with both WY14,643 and a PPARgamma ligand, rosiglitazone, did not lower glucose levels more effectively than did treatment with WY14,643 alone. These data support the hypothesis that reducing intracellular triglycerides in non-adipose tissues improves insulin sensitivity and suggest that further investigation of the role of PPARalpha agonists in the treatment of lipoatrophic diabetes is warranted.     

Synergistic activation of UCP-3 expression in cultured fetal rat brown adipocytes by PPARalpha and PPARgamma ligands

Rat brown adipocytes express mRNAs for Uncoupling Proteins (UCP) 1, 2 and 3 and the Peroxisome Proliferator Activated Receptors (PPAR) alpha and gamma. We have examined the effects of selective PPARalpha or -gamma activation on changes in UCP-1 and UCP-3 mRNA levels in cultured fetal rat brown adipocytes (FBA). Rosiglitazone (1.0 microM), a selective PPARgamma agonist, elicited 5- and 3-fold increases in UCP-1 and UCP-3, respectively. The PPARalpha ligand, Wy14643 (10.0 microM) increased UCP-3 tenfold, but decreased UCP-1. A synergistic effect on UCP-3 expression (30-fold increase; P < 0. 05) was observed when FBA were exposed to a combination of Wy14643 (10.0 microM) and rosiglitazone (10.0 microM). Thus, activation of PPARgamma increases UCP-1 and UCP-3 levels which are differentially regulated by PPARalpha. A synergistic interaction occurs between PPARalpha and PPARgamma in the regulation of UCP-3 in FBA, probably via co-activator recruitment, suppression of co-repressor proteins or through a direct interaction at the level of the PPRE.     

SAR-oriented discovery of peroxisome proliferator-activated receptor pan agonist with a 4-adamantylphenyl group as a hydrophobic tail

3-(4-Alkoxyphenyl)propanoic acid derivatives were prepared as candidate peroxisome proliferator-activated receptor (PPAR) alpha/delta/gamma pan agonists, based on our previous SAR studies directed toward the development of subtype-selective PPAR agonists. Those studies indicated that the steric bulkiness of substituents introduced at the distal benzene ring had an important influence on PPAR activity. The finding that a 4-adamantyl derivative exhibited not only PPARalpha/delta activity but also significant PPARgamma activity prompted us to search for structurally novel phenylpropanoic acid derivatives with more potent adipocyte differentiation activity than the well-known PPARgamma agonist, rosiglitazone, as well as well-balanced PPARalpha and PPARdelta agonistic activities. A representative phenylpropanoic acid derivative (12) bearing a 4-adamantylphenyl substituent proved to be a well-balanced PPAR-pan agonist with activities to regulate the expression of genes involved in lipid and glucose homeostasis, and should be useful as a candidate drug for the treatment of altered PPAR function.     

Liver peroxisome proliferator-activated receptor gamma contributes to hepatic steatosis,triglyceride clearance,and regulation of body fat mass

Peroxisome proliferator-activated receptor gamma (PPAR gamma) is a nuclear receptor that mediates the antidiabetic effects of thiazolidinediones. PPAR gamma is present in adipose tissue and becomes elevated in fatty livers, but the roles of specific tissues in thiazolidinedione actions are unclear. We studied the function of liver PPAR gamma in both lipoatrophic A-ZIP/F-1 (AZIP) and wild type mice. In AZIP mice, ablation of liver PPAR gamma reduced the hepatic steatosis but worsened the hyperlipidemia, triglyceride clearance, and muscle insulin resistance. Inactivation of AZIP liver PPAR gamma also abolished the hypoglycemic and hypolipidemic effects of rosiglitazone, demonstrating that, in the absence of adipose tissue, the liver is a primary and major site of thiazolidinedione action. In contrast, rosiglitazone remained effective in non-lipoatrophic mice lacking liver PPAR gamma, suggesting that adipose tissue is the major site of thiazolidinedione action in typical mice with adipose tissue. Interestingly, mice without liver PPAR gamma, but with adipose tissue, developed relative fat intolerance, increased adiposity, hyperlipidemia, and insulin resistance. Thus, liver PPAR gamma regulates triglyceride homeostasis, contributing to hepatic steatosis, but protecting other tissues from triglyceride accumulation and insulin resistance.     

A novel PPARalpha agonist ameliorates insulin resistance in dogs fed a high-fat diet

Agonism of peroxisome proliferator-activated receptor (PPAR) alpha, a key regulator of lipid metabolism, leads to amelioration of lipid abnormalities in dyslipidemic patients. However, whether PPARalpha agonism is an effective form of therapy for obesity-related insulin resistance associated with lipid abnormalities is unclear. The present study investigated the effects of a potent and subtype-selective PPARalpha agonist, KRP-101, in a nonrodent insulin-resistant animal model under pair-fed conditions. Beagle dogs were fed a high-fat diet for 24 wk to induce insulin resistance. During the final 12 wk, 0.03 mg x kg(-1) x day(-1) KRP-101 (n = 5) or vehicle (n = 5) was administered orally once a day. KRP-101 administration resulted in a significantly lower weight of overall visceral fat, which is associated with increased adiponectin and decreased leptin in serum. KRP-101 administration improved hyperglycemia and hyperinsulinemia as well as dyslipidemia in dogs fed a high-fat diet. Oral glucose tolerance test showed that KRP-101 administration improved glucose intolerance. The KRP-101 group showed a markedly lower hepatic triglyceride concentration. Lipid oxidation was increased in the liver and skeletal muscles of the KRP-101 group. These findings in the dog model suggest that the use of potent and subtype-selective PPARalpha agonists as a potentially relevant therapeutic approach to treat human insulin resistance associated with visceral obesity.     

Protein expression changes in the Sprague Dawley rat liver proteome following administration of peroxisome proliferator activated receptor alpha and gamma ligands

Peroxisome proliferator activated receptors (PPARs) are members of the nuclear receptor superfamily and are intimately involved in lipid metabolism and energy homeostasis. Activation of these receptors in rodents can lead to hepatomegaly and ultimately hepatic carcinogenesis although the mechanisms by which these processes occur are poorly understood. To further our understanding of these processes and to discriminate between different PPAR mediated signalling pathways, a proteomic approach has been undertaken to identify changes in protein expression patterns in Sprague Dawley rat liver following dosing with a PPARalpha agonist (Wyeth 14643), a PPARgamma agonist (Troglitazone) and a compound with mixed PPARalpha/gamma agonist activity (SB-219994). Using one-and-two-dimensional electrophoresis of tissue lysates a diverse range of protein abundance changes was observed in these tissues. Whilst a number of these proteins have PPAR response elements (PPREs) in their respective promoters, another group was detected whose expression has been documented to be sensitive to peroxisome proliferator administration. Most notably within these groups, proteins involved in lipid catabolism displayed increased expression following drug administration. A further subset of proteins, with less obvious biological implications, also showed altered expression patterns. Where available, sequences upstream of the coding regions of genes not previously known to have PPREs were searched with positional consensus matrices for the presence of PPREs in an attempt to validate these changes. Using such an approach putative PPARgamma and PPARdelta response elements were discovered upstream of the tubulin beta coding region. There was limited overlap in observed protein abundance changes between the three groups, and where this was the case (cytosolic epoxide hydrolase, peroxisomal bifunctional enzyme, hydroxymethyl glutaryl CoA, synthase, long chain acyl-CoA thioesterase), expression of these proteins had previously been shown to be under the control of PPAR activity.     

Cardiovascular drugs inhibit MMP-9 activity from human THP-1 macrophages

It is now recognized that atherosclerosis complications are related to the unstable character of the plaque rather than its volume. Vulnerable plaques often contain a large lipid core, a reduced content of smooth muscle cells, and accumulation of inflammatory cells. Colocalization of macrophages and active matrix metalloproteinases (MMPs) is likely relevant for atherosclerotic lesion disruption. Nevertheless, MMP activity and regulation by cardiovascular drugs remains poorly defined. In this study, we evaluated the effects of avasimibe, fluvastatin, and peroxisome proliferator-activated receptor (PPAR) ligands on 92-kDa gelatinase B (MMP-9) secretion by human THP-1 macrophages. THP-1 macrophages were treated with compounds for 48 h, and secreted MMP-9 protein was quantified by immunoassay. Avasimibe, fluvastatin, and PPARalpha agonists (fenofibric acid and Wy-14643) significantly reduced, in a concentration-dependent manner, MMP-9 protein (up to 67 +/- 5% for fenofibric acid). In these assays, the PPARgamma selective agonist rosiglitazone displayed a lower efficacy than other compounds. Enzymatic activity of MMP-9 was also decreased by all cardiovascular drugs tested. MMP-9 protein/activity inhibition by cardiovascular drugs was due, at least in part, to a decrease in MMP-9 mRNA. These results show that THP-1 macrophages could be an useful cellular model to investigate effects of compounds on plaque vulnerability through MMP-9 activity.