PPARγ Modulators Research Articles

Benzoyl 2‐Methylindoles as Selective PPARγ Modulators.

AbstractFor Abstract see ChemInform Abstract in Full Text.

Selective PPARγ modulators with improved pharmacological profiles

A series of metabolically robust N-benzyl-indole selective PPARγ modulators with either a 3-benzoyl or 3-benzisoxazoyl moiety have been identified. In vitro, these compounds are partial agonists and exhibit reduced adipogenesis in human adipocytes. In vivo, these SPPARγMs result in potent glucose lowering in db/db mice and attenuate increases in heart weight and brown adipose tissue that is typically observed in rats upon treatment with PPARγ full agonists.

Selective activation of PPARγ in breast, colon, and lung cancer cell lines

Peroxisome proliferator-activated receptor gamma (PPARγ) plays a critical albeit poorly defined role in the development and progression of several cancer types including those of the breast, colon, and lung. A PPAR response element (PPRE) reporter assay was utilized to evaluate the selective transactivation of PPARγ in 10 different cell lines including normal mammary epithelial, breast, lung, and colon cancer cells. Cells were treated with one of four compounds including rosglitizone (Ros), ciglitizone (Cig), 15-deoxy-Δ 12,14-prostaglandin J 2 (PGJ 2), or GW 9662 (GW). We observed differences in transactivation between cell lines from different tissue origin, across cell lines from a single tissue type, and selective modulation of PPARγ within a single cell line by different ligands. Interestingly, GW, a PPARγ antagonist in adipocytes, enhanced PPRE reporter activation in normal mammary epithelial cells while it had virtually no effect in any of the cancer cell lines tested. Within each cancer type, individual cell lines were found to respond differently to distinct PPARγ ligands. For instance, Ros, Cig, and PGJ 2 were all potent agonist of PPARγ transactivation in lung adenocarcinoma cell lines while these same ligands had no effect in squamous cell or large cell carcinomas of the lung. Message levels of PPARγ and retinoid X receptor alpha (RXRα) in the individual cell lines were quantitated by real time-polymerase chain reaction (RT-PCR). The ratio of PPARγ to RXRα was predictive of how cells responded to co-treatment of Ros and 9- cis-retinoic acid, an RXRα agonist, in two out of three cell lines tested. These data indicate that PPARγ can be selectively modulated and suggests that it may be used as a therapeutic target for individual tumors.

FK614, a Novel Peroxisome Proliferator-Activated Receptor γ Modulator, Induces Differential Transactivation Through a Unique Ligand-Specific Interaction With Transcriptional Coactivators

Peroxisome proliferator-activated receptor γ (PPARγ) is a ligand-dependent transcriptional factor implicated in regulating adipogenesis, glucose homeostasis, and in mediating the action of the insulin sensitizing anti-diabetic thiazolidinedione (TZD) compounds. [3-(2,4-Dichlorobenzyl)-2-methyl-N-(pentylsulfonyl)-3-H-benzimidazole-5-carboxamide] (FK614) is a structurally novel PPARγ agonist that demonstrates potent anti-diabetic activity in vivo. Herein, we describe that FK614 is a selective PPARγ ligand with specific transactivation properties that are dependent upon the context of coactivators. FK614 dissociates the corepressors NCoR (nuclear receptor corepressor) and SMRT (silencing mediator of retinoid and thyroid hormone receptors) from PPARγ as effectively as rosiglitazone and pioglitazone, but can also differentially induce a ligand specific interaction of PPARγ with coactivators. The amount of CBP (CREB-binding protein) and SRC-1 (steroid receptor coactivator-1) recruited by FK614 was less than that induced by rosiglitazone and pioglitazone, but FK614 caused similar PGC-1α (PPARγ coactivator-1α) recruitment as these compounds. As a consequence of these ligand-specific differences in the strength of ligand-type specific interactions of PPARγ and coactivators, FK614 functions as a partial or full agonist for transcriptional activation depending upon the amount of specific coactivators in cells following overexpression. In conclusion, FK614 is a novel, non-TZD type, and selective PPARγ modulator whose pharmacological properties are distinct from rosiglitazone and pioglitazone.

Impaired lipid accumulation by trans10, cis12 CLA during adipocyte differentiation is dependent on timing and length of treatment

Conjugated linoleic acids (CLAs) are a group of polyunsaturated fatty acids found in ruminant products, where the predominant isomers are cis9, trans11 (c9,t11) and trans10, cis12 (t10,c12) CLA. We have previously shown that t10,c12 CLA prevents lipid accumulation in mature adipocytes in part by acting as a peroxisome proliferator-activated receptor γ (PPARγ) modulator. The objective of this study was to further establish the molecular mechanisms underlying the attenuating effect on lipid accumulation by t10,c12 CLA, with focus on time point and duration of treatment during adipogenesis. We have shown that t10,c12 CLA treatment has its most attenuating effect early (day (D) 0–6) during differentiation. Treatment during this period is sufficient to prevent lipid accumulation in mature adipocytes. The adipogenic marker genes PPARγ and CCAAT/enhancer binding protein α (C/EBPα) are both down-regulated after treatment within the period from D0–6, while additional treatment also down-regulates the expression of sterol regulatory element binding protein-1c (SREBP-1c), liver X receptor α (LXRα), fatty acid binding protein (aP2), fatty acid translocase (CD36) and insulin-sensitive glucose transporter 4 (GLUT4). These effects of t10,c12 CLA reflect the subsequent attenuation of lipid accumulation observed in mature adipocytes. Interestingly, the early B-cell factor (O/E-1), which is known to promote adipogenesis and to be involved in control of genes important for terminal adipocyte differentiation, is unaffected by treatment of t10,c12 CLA. Taken together, our data indicate that inhibition of lipid accumulation induced by t10,c12 CLA treatment during adipocyte differentiation is associated with a tight regulatory cross-talk between early (PPARγ and C/EBPα) and late (LXRα, aP2 and CD36) adipogenic marker genes.

Benzoyl 2-methyl indoles as selective PPARγ modulators

Routine screening for human PPAR ligands yielded compounds 1 and 2, both of which were sub-micromolar hPPARγ agonists. Synthetic modifications of these leads led to a series of potent substituted 3-benzyl-2-methyl indoles, a subset of which were noted to be selective PPARγ modulators (SPPARγMs). SPPARγM 24 displayed robust anti-diabetic activity with an improved therapeutic window in comparison to a PPARγ full agonist in a rodent efficacy model.

A high-capacity assay for PPARγ ligand regulation of endogenous aP2 expression in 3T3-L1 cells

A novel class of insulin-sensitizing agents, the thiazolidinedines (TZDs), has proven effective in the treatment of type 2 diabetes. These compounds, as well as a subclass of non-TZD insulin-sensitizing agents, have been shown to be peroxisome proliferator-activated receptor (PPAR) γ agonists. PPARγ plays a critical role in adipogenesis and PPARγ agonists have been shown to induce adipocyte differentiation. Here, PPARγ ligand activity has been assessed in murine 3T3-L1 cells, a commonly used in vitro model of adipogenesis, by measuring their ability to induce adipocyte fatty acid-binding protein (aP2) mRNA expression. In order to perform this task, we have developed a novel, multiwell assay for the direct detection of aP2 mRNA in cell lysates that is based on hybridization of mRNA to target-specific oligonucleotides. These oligonucleotide probes are conjugated to enzymes that efficiently process unique chemical substrates into robust fluorescent products. Ribosomal protein 36B4 mRNA, a gene whose expression is unaffected by adipogenesis, serves as the control in the assay. Two assay formats have been developed, a single analyte assay in which aP2 and 36B4 mRNA expression are assayed in separate lysate aliquots and a dual analyte assay which can measure aP2 and 36B4 mRNA simultaneously. Both forms of the assay have been used to quantify attomole levels of aP2 and 36B4 mRNAs in differentiating 3T3-L1 preadipocytes treated with PPARγ agonists. The potencies of PPARγ agonists determined by this novel methodology showed good correlation with those derived from aP2 mRNA slot-blot analysis and PPARγ transactivation assays. We conclude that the aP2 single and dual analyte assays both provide specific and sensitive measurements of endogenous aP2 mRNA levels that can be used to assess the activity of PPARγ ligands in 3T3-L1 cells. Since the assay obviates the need for RNA isolation and is performed in an automatable multiwell format, it can serve as a high-throughput, cell-based screen for the identification and characterization of PPARγ modulators.

Trans10, cis12-conjugated linoleic acid prevents triacylglycerol accumulation in adipocytes by acting as a PPARγ modulator

A group of polyunsaturated fatty acids called conjugated linoleic acids (CLAs) are found in ruminant products, where the most common isomers are cis9, trans11 (c 9,t11) and trans10, cis12 (t10,c12) CLA. A crude mixture of these isomers has been shown in animal studies to alter body composition by a reduction in body fat mass as well as an increase in lean body mass, with the t10,c12 isomer having the most pronounced effect. The objective of this study was to establish the molecular mechanisms by which t10,c12 CLA affects lipid accumulation in adipocytes. We have shown that t10,c12 CLA prevents lipid accumulation in human and mouse adipocytes at concentrations as low as 5 microM and 25 microM, respectively. t10,c12 CLA fails to activate peroxisome proliferator-activated receptor gamma (PPARgamma) but selectively inhibits thiazolidinedione-induced PPARgamma activation in 3T3-L1 adipocytes. Treatment of mature adipocytes with t10,c12 CLA alone or in combination with Darglitazone down-regulates the mRNA expression of PPARgamma as well as its target genes, fatty acid binding protein (aP2) and liver X receptor alpha (LXRalpha). Taken together, our results suggest that the trans10, cis12 CLA isomer prevents lipid accumulation in adipocytes by acting as a PPARgamma modulator.

3WS16-6 Identification of angiotensin receptor antagonists with PPARγ modulating activity: Implications for treatment of the metabolic syndrome

3WS16-6 Identification of angiotensin receptor antagonists with PPARγ modulating activity: Implications for treatment of the metabolic syndrome

Broadening TRAIL's Targets

Identifying agents that destroy tumor cells but leave normal cells unharmed is a major goal in developing effective cancer treatments. The cytokine TRAIL, a member of the tumor necrosis family, has shown such potential, as it preferentially induces apoptosis of cancer cells. Now, Kim et al. report a potential mechanism to expand the range of TRAIL cancer targets. Agonists for a nuclear receptor called peroxisome proliferator-activated receptor-gamma (PPARγ) sensitized tumor cells that are normally not responsive to TRAIL such that TRAIL did induce apoptosis. Although neither the PPARγ modulators nor TRAIL alone killed these tumor cells, combined treatment did. Furthermore, the combination did not kill normal cells. The mechanism appears not to involve PPARγ, because it persisted even in the presence of dominant-negative form of PPARγ. It seems to be mediated by the formation of a TRAIL receptor-associated signaling complex that activates the cell death program through caspase activation. A cytoplasmic protein called FLIP normally inhibits assembly of this complex, but PPARγ modulators somehow stimulate the degradation of FLIP through the ubiquitin-proteosome pathway. The cellular target of the PPARγ modulators that stimulates this proapoptotic effect has yet to be determined. Y. Kim, N. Suh, M. Sporn, J. C. Reed, An inducible pathway for degradation of FLIP protein sensitizes tumor cells to TRAIL-induced apoptosis. J. Biol. Chem. 277 , 22320-22329 (2002). [Abstract] [Full Text]

A Unique PPARγ Ligand with Potent Insulin-Sensitizing yet Weak Adipogenic Activity

FMOC-L-Leucine (F-L-Leu) is a chemically distinct PPARγ ligand. Two molecules of F-L-Leu bind to the ligand binding domain of a single PPARγ molecule, making its mode of receptor interaction distinct from that of other nuclear receptor ligands. F-L-Leu induces a particular allosteric configuration of PPARγ, resulting in differential cofactor recruitment and translating in distinct pharmacological properties. F-L-Leu activates PPARγ with a lower potency, but a similar maximal efficacy, than rosiglitazone. The particular PPARγ configuration induced by F-L-Leu leads to a modified pattern of target gene activation. F-L-Leu improves insulin sensitivity in normal, dietinduced glucose-intolerant, and in diabetic db/db mice, yet it has a lower adipogenic activity. These biological effects suggest that F-L-Leu is a selective PPARγ modulator that activates some (insulin sensitization), but not all (adipogenesis), PPARγ-signaling pathways.