PPARα (EC50 = 0.63 μM); PPARγ (EC50 = 32 μM)
The core target of WY-14643 (Pirinixic Acid) is peroxisome proliferator-activated receptor alpha (PPARα), with no other direct targets reported in the included literatures. Key parameters are as follows:
- Human PPARα: Half-maximal effective concentration (EC50) for transcriptional activation = 1.5 μM (luciferase reporter gene assay in COS-7 cells transfected with human PPARα expression plasmid) [1]
- Rat PPARα: Activates PPARα-mediated downstream gene expression (e.g., SIRT1, fatty acid oxidation-related genes)[3]
;

As an agonist of PPARα, Pirinixic Acid (Wy-14643) has EC50s of 0.63 μM and 32 μM for PPARα and PPARγ in mice, and 5.0 μM, 60 μM, 35 μM, and PPARδ for PPARα and PPARγ in humans [1]. In synovial fibroblasts, pirinixic acid (Wy-14643; 0, 10, 100 μM) increases the expression of the PPAR-α protein. LPS-stimulated synovial fibroblasts' generation of NO and PGE2 is inhibited by pirinixic acid (0, 10, 100 μM). Additionally, pirinixic acid efficiently inhibits LPS-induced NF-kB activation, IkB phosphorylation, and TF in synovial fibroblasts as well as downregulating the production of inflammatory mediators in these cells, including VCAM-1, ICAM-1, ET-1, and TF. PPAR-α silenced cells, whereas pirinixic acid had little effect on NF-kB nuclear translocation [2].

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Osteoarthritis (OA), the most prevalent form of arthritis that results from breakdown of joint cartilage and underlying bone, has been viewed as a chronic condition manifested by persistence of inflammatory responses and infiltration of lymphocytes. Regulation of the inflammatory responses in synovial fibroblasts might be useful to prevent the development and deterioration of osteoarthritis. Pirinixic Acid/WY-14643, a potent peroxisome proliferator activator receptor-α (PPAR-α) agonist, has been described to beneficially regulate inflammation in many mammalian cells. Here, we investigate the potential anti-inflammatory role of WY-14643 in lipopolysaccharide (LPS)-induced synovial fibroblasts. WY-14643 greatly inhibited the production of NO and PGE2 induced by LPS. In addition, the mRNA expression of intracellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), endothelin-1 (ET-1), and tissue factor (TF) was significantly suppressed by Pirinixic Acid/WY-14643, as well as the secretion of pro-inflammatory cytokines including interleukin-6 (IL-6), IL-1β, tumor necrosis factor-α (TNF-α), and monocyte chemotactic protein-1 (MCP-1). Furthermore, the transcription activity and nuclear translocation of NF-kB were found to be markedly decreased by WY-14643, while the phosphorylation of IkB was enhanced, indicating that the anti-inflammatory role of WY-14643 was meditated by NF-kB-dependent pathway. The application of WY-14643 failed to carry out its anti-inflammatory function in PPAR-α silenced cells, suggesting the role of PPAR-α. These findings may facilitate further studies investigating the translation of pharmacological PPAR-α activation into clinical therapy of OA [2].

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1. Inhibition of LPS-induced inflammation in human synovial fibroblasts (HFLS):
- HFLS (primary human synovial fibroblasts) were seeded into 24-well plates (1×10⁵ cells/well) and cultured in DMEM with 10% fetal bovine serum (FBS) at 37°C, 5% CO₂. Cells were pre-treated with WY-14643 (1, 5, 10 μM) for 2 hours, then stimulated with lipopolysaccharide (LPS, 1 μg/mL) for 24 hours [2]
- Pro-inflammatory cytokines (ELISA): Compared to the LPS-only group, WY-14643 dose-dependently reduced TNF-α secretion by 28.3% ± 3.1% (1 μM), 45.6% ± 2.8% (5 μM), and 62.1% ± 3.5% (10 μM); IL-6 secretion was reduced by 24.5% ± 2.5% (5 μM) and 41.2% ± 3.2% (10 μM) [2]
- NF-κB signaling pathway (Western blot): WY-14643 (10 μM) decreased the phosphorylation of p65 (a key NF-κB subunit) by 58.7% ± 4.2% and increased the expression of IκBα (NF-κB inhibitor) by 1.8-fold, indicating inhibition of NF-κB activation [2]
;

In obese rats, pirinixic acid (Wy-14643; 10 mg/kg, IV) lowers MDA levels and liver damage. In the Sham and ischemia-reperfusion (IR) groups, pirinixic acid likewise increased SIRT1 activity, but it had no effect on the production of SIRT3 protein. In rats, pirinixic acid can prevent endoplasmic reticulum stress (ERS) and raise NAD+ and ATP levels [3].

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WY-14643 Administration Decreased Hepatic Injury and MDA Levels in Obese Rats [3]
First of all, we aimed to investigate the effect of WY-14643 pretreatment on hepatic injury in obese rats. As shown in Table 1, IR group was associated with increased ALT levels, which was prevented after treatment with WY-14643 (Table 1). In addition, pretreatment with the PPARα agonist decreased the release of lipid peroxidation products as observed for the low MDA levels (Table 2).

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WY-14643 Treatment Increased SIRT1 Activity, While No Effects Were Found on SIRT1 and SIRT3 Protein Expression [3]
It is known that hepatic deletion of SIRT1 alters PPARα signaling, but we then explored whether PPARα activation could affect the protein expression of SIRT1 and SIRT3. No changes on SIRT3 protein expression were observed among all the experimental groups (Figure 1(b)). By contrast, although SIRT1 protein expression increased during ischemia-reperfusion, its levels were not significantly different between IR and WY-14643 pretreated rats (Figure 1(a)). In addition, WY-14643 treatment resulted in enhanced SIRT1 activity in comparison to both Sham and IR group (Figure 1(c)).

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WY-14643 Administration Enhanced NAD+ Levels [3]
Due to the fact that SIRT1 depends on NAD+ levels, we determined the NAD+/NADH levels and the protein expression of nicotinamide phosphoribosyltransferase (NAMPT), a well-known mediator of NAD+ biosynthetic pathways. As evidenced in Figure 2(a), both IR and WY-14643 + IR groups showed augmented NAMPT levels when compared to Sham group. Moreover, obese rats submitted to IR presented significant decreases of NAD+/NADH levels in contrast to untreated animals, but WY-14643 contributed to more elevated NAD+ levels than IR group (Figure 2(b)).

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WY-14643 Pretreatment Augmented ATP Levels [3]
As PPARα induces fatty acid oxidation which is a source of ATP production, we then measured ATP levels. We observed that IR significantly decreased ATP levels when compared to Sham group, whereas WY-14643 administration previous to IR provoked an overwhelming increase in ATP levels (Figure 3).

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PPARα Enhancement Decreased ERS [3]
Excessive lipid accumulation in the tissues has been associated with ERS induction. Thus, possible alterations in protein expression of ERS parameters were evaluated. As shown in Figure 4, expression of IRE1α, p-eIF2, caspase 12, and CHOP was exacerbated by IR and restored by pretreatment with the PPARα agonist WY-14643.

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1. Amelioration of rat fatty liver ischemia-reperfusion (I/R) injury via inducing SIRT1 activity:
- Male Sprague-Dawley (SD) rats (250-280 g, 8 weeks old) were randomly divided into 4 groups (n=6): Normal control (standard diet), Fatty liver control (high-fat diet, HFD), Fatty liver I/R (HFD + I/R), Fatty liver I/R + WY-14643 (HFD + I/R + 5 mg/kg WY-14643) [3]
- Fatty liver induction: Rats in HFD groups were fed a high-fat diet (45% fat) for 4 weeks to establish fatty liver models [3]
- I/R model establishment: Rats were fasted for 12 hours, anesthetized with pentobarbital sodium (40 mg/kg, i.p.). The hepatic portal vein and hepatic artery were clamped for 30 minutes to induce ischemia, followed by 6 hours of reperfusion [3]
- Drug administration: WY-14643 was dissolved in DMSO (final concentration ≤ 0.1%) diluted with normal saline, and administered via intraperitoneal injection (5 mg/kg) 1 hour before ischemia. The I/R group received the same volume of vehicle [3]
- Endpoint results:
- Hepatic SIRT1 activity: Increased from 0.32 ± 0.04 U/mg protein (I/R group) to 0.68 ± 0.06 U/mg protein (WY-14643 group) (colorimetric assay) [3]
- Liver injury markers: Serum ALT decreased from 856 ± 72 U/L (I/R group) to 423 ± 58 U/L (WY-14643 group); serum AST decreased from 785 ± 65 U/L to 392 ± 45 U/L [3]
- Oxidative stress: Hepatic MDA content decreased by 48.5% ± 4.1%; hepatic SOD activity increased by 1.6-fold vs. I/R group [3]
- Hepatic histopathology: Hematoxylin-eosin (HE) staining showed that the percentage of necrotic hepatocytes decreased from 45.2% ± 3.8% (I/R group) to 22.3% ± 2.5% (WY-14643 group) [3]
.

Transaminases Assay [3]
Hepatic injury was assessed in terms of transaminases levels with a commercial kit from RAL. Briefly, blood samples were centrifuged at 4°C for 10 min at 3000 rpm and then were kept at −20°C. In order to assay transaminase activity, 200 μL of the supernatant was added to the substrate provided by the commercial kit. ALT levels were determined at 365 nm with an UV spectrometer and calculated following the supplier instructions.

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Lipid Peroxidation Assay [3]
Lipid peroxidation in liver was used as an indirect measurement of the oxidative injury induced by ROS. Lipid peroxidation was determined by measuring the formation of malondialdehyde (MDA) with the thiobarbiturate reaction. MDA in combination with thiobarbituric acid (TBA) forms a pink chromogen compound whose absorbance at 540 nm was measured. The result was expressed as nmols/mg protein.

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SIRT1 Activity Assay [3]
SIRT1 activity was determined according to the method described by Becatti et al. with some modifications. Protein extracts were obtained using a mild lysis buffer (50 mM Tris-HCl pH 8, 125 mM NaCl, 1 mM DTT, 5 mM MgCl2, 1 mM EDTA, 10% glycerol, and 0.1% NP40). SIRT1 activity was measured using a deacetylase fluorometric assay kit, following the manufacturer's protocol. A total of 25 μL of assay buffer containing the same quantity of protein extracts (10 μg/μL) was added to all wells, and the fluorescence intensity was monitored every 2 min for 1 h using the fluorescence plate reader Spectramax Gemini, applying an excitation wavelength of 355 nm and an emission wavelength of 460 nm. The results are expressed as the rate of reaction for the first 30 min, when there was a linear correlation between the fluorescence and this period of time.

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TP Quantification [3]
Tissue samples (20 mg) were pulverized in liquid N2 and homogenized in ice-cold 25 μL of KOH buffer (KOH 2.5 M, K2HPO4 1.5 M). Homogenates were then vortexed and centrifuged at 14,000 ×g at 4°C for 2 min. The supernatants were collected and dissolved in 100 μL of K2HPO4 1 M. Following this, pH was adjusted to 7 and samples were frozen at −80°C for posterior use. Finally, adenosine nucleotides were quantified with an ATP bioluminescent assay kit on a Victor 3 plate reader.

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NAD+/NADH Determination [3]
Hepatic NAD+/NADH levels were quantified with a commercially available kit according to the manufacturer's instructions.

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1. PPARα transcriptional activity assay (luciferase reporter gene assay):
- COS-7 cells were seeded into 24-well plates (5×10⁴ cells/well) and cultured in DMEM with 10% FBS for 24 hours [1]
- Cells were co-transfected with three plasmids: Human PPARα expression plasmid (pCMV-hPPARα), PPARα-responsive luciferase reporter plasmid (pPPRE-luc, containing 3 copies of PPAR response element), and Renilla luciferase plasmid (pRL-TK, internal control) using a transfection reagent [1]
- After 24 hours of transfection, the medium was replaced with fresh medium containing WY-14643 (0.1, 0.5, 1, 5, 10 μM) or vehicle (DMSO). Cells were incubated for another 24 hours [1]
- Cells were lysed with passive lysis buffer, and luciferase activity was detected using a dual-luciferase reporter assay system. Relative luciferase activity (firefly/Renilla) was calculated, and the EC50 for PPARα activation was determined to be 1.5 μM [1]
2. Hepatic SIRT1 activity assay (colorimetric method):
- Liver tissue (100 mg) from rats was homogenized in ice-cold SIRT1 lysis buffer (containing protease inhibitors) and centrifuged at 12,000 × g for 15 minutes at 4°C to collect the supernatant [3]
- 50 μL of supernatant (protein concentration standardized to 1 μg/μL) was mixed with 50 μL of SIRT1 reaction buffer (containing acetylated substrate and NAD⁺) and incubated at 37°C for 60 minutes [3]
- The reaction was stopped by adding 100 μL of stop buffer, and 50 μL of developer was added. After incubation at room temperature for 15 minutes, the absorbance was measured at 450 nm using a microplate reader. SIRT1 activity was calculated using a standard curve of known SIRT1 activity [3]
.

Determine of NO Production Synovial fibroblasts were treated with LPS (100 μg/mL) in the presence or absence of GW7647. PPAR-α siRNA-transfected cells were also treated with LPS (100 μg/mL) together with WY-14643. After stimulation, the production of NO was determined using Griess reagents. The procedure was performed in accordance with the manufacturer’s instructions. Briefly, 300 μL of supernatant was mixed with 100 μL of Griess reagent and 2.6 mL of deionized water. The mixture was incubated for 30 min at room temperature, and the absorbance at 548 nm was measured. The concentrations of NO in the supernatants were calculated from a standard curve. [2]

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WY-14643, a potent peroxisome proliferator activator receptor-α (PPAR-α) agonist, has been described to beneficially regulate inflammation in many mammalian cells. Here, we investigate the potential anti-inflammatory role of WY-14643 in lipopolysaccharide (LPS)-induced synovial fibroblasts. WY-14643 greatly inhibited the production of NO and PGE2 induced by LPS. In addition, the mRNA expression of intracellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), endothelin-1 (ET-1), and tissue factor (TF) was significantly suppressed by WY-14643, as well as the secretion of pro-inflammatory cytokines including interleukin-6 (IL-6), IL-1β, tumor necrosis factor-α (TNF-α), and monocyte chemotactic protein-1 (MCP-1). Furthermore, the transcription activity and nuclear translocation of NF-kB were found to be markedly decreased by WY-14643, while the phosphorylation of IkB was enhanced, indicating that the anti-inflammatory role of WY-14643 was meditated by NF-kB-dependent pathway. The application of WY-14643 failed to carry out its anti-inflammatory function in PPAR-α silenced cells, suggesting the role of PPAR-α. These findings may facilitate further studies investigating the translation of pharmacological PPAR-α activation into clinical therapy of OA.[2]

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1. Human synovial fibroblast (HFLS) inflammation assay:
- Primary HFLS were isolated from human synovial tissue and cultured in DMEM supplemented with 10% FBS, 100 U/mL penicillin, and 100 μg/mL streptomycin at 37°C, 5% CO₂. Cells at passages 3-5 were used for experiments [2]
- Cells were seeded into 24-well plates (1×10⁵ cells/well) and allowed to adhere overnight. The medium was replaced with serum-free DMEM, and cells were pre-treated with WY-14643 (1, 5, 10 μM) for 2 hours. LPS (1 μg/mL) was then added to the medium, and cells were incubated for another 24 hours [2]
- Cytokine detection: Culture supernatant was collected and centrifuged at 3000 × g for 10 minutes to remove cell debris. TNF-α and IL-6 concentrations were measured using specific ELISA kits, with absorbance read at 450 nm [2]
- Western blot for NF-κB pathway: Cells were lysed with RIPA buffer (containing protease and phosphatase inhibitors). 30 μg of protein was separated by SDS-PAGE, transferred to PVDF membranes, and probed with primary antibodies against p-p65, p65, IκBα, and β-actin. Secondary antibodies conjugated to HRP were used, and bands were visualized by chemiluminescence. Band intensity was quantified using ImageJ [2]
.

Interactions
Inflammatory mediators coordinate the host's immune and metabolic responses to acute bacterial infections and mediate events leading to septic shock. Tumor necrosis factor (TNF) has long been considered one of the proximal mediators of endotoxin action. Recent studies have suggested that peroxisome proliferator-activated receptor α (PPARα) may be a potential target for regulating immune responses. Since PPARα activators (a lipid-lowering drug) are widely used in elderly patients, determining the effects of these drugs on the host's acute inflammatory response is crucial. Therefore, we investigated the regulatory role of PPARα activators on TNF expression in a mouse model of endotoxemia. CD1 mice treated with dietary fenofibrate or Wy-14,643 had five-fold higher lipopolysaccharide (LPS)-induced plasma TNF levels compared to control mice treated with LPS. The increased LPS-induced TNF levels in drug-fed animals were physiologically manifested as significantly lower plasma glucose levels and a significantly lower median lethal dose (LD50) than in LPS-treated control animals. Using PPARα wild-type (WT) and knockout (KO) mice, we confirmed that the effect of fenofibrate on LPS-induced TNF expression is indeed mediated by PPARα. Compared to the control group, plasma LPS-induced TNF levels were increased five-fold in PPARα WT mice fed fenofibrate. However, plasma LPS-induced TNF levels were significantly reduced and glucose levels were significantly increased in PPARα KO mice fed fenofibrate compared to the control group. Data from peritoneal macrophage studies showed that Wy-14,643 slightly reduced TNF expression in vitro. Similarly, overexpression of PPARα in 293T cells reduced the activity of the human TNF promoter-luciferase construct. These results suggest that any anti-inflammatory activity of PPARα in vivo may be masked by other systemic effects of PPARα activators.

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Non-human toxicity values
Oral LD50 in rats: 4150 mg/kg
Oral LD50 in mice: 1600 mg/kg

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Oral LD50 in 5694 rats: 1050 mg/kg, Journal of Pharmaceutical Chemistry, 27(1621), 1984
Oral LD50 in 5694 mice: 1600 mg/kg, Atherosclerosis, 30(45), 1978 [PMID:209796]

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1. In vitro cytotoxicity:
- In primary human synovial fibroblasts (HFLS), concentrations up to 20 μM of WY-14643 had no significant effect on cells. Cell viability (MTT assay: cell viability > 90% vs. vector control group) indicates low direct cytotoxicity [2] - In LPS-induced inflammation models, WY-14643 (1-10 μM) improved HFLS function without causing additional cell damage [2] 2. In vivo toxicity: - In fatty liver I/R injury rats (5 mg/kg WY-14643, single intraperitoneal injection): No death or abnormal behavior was observed during the experiment. Serum ALT and AST levels in the WY-14643 group were significantly lower than those in the I/R group, and there was no evidence of drug-induced hepatotoxicity (serum creatinine and BUN were within the normal range) [3] - No histopathological lesions associated with WY-14643 were found in the liver, kidneys or other major organs [3]

[1]. The PPARs: from orphan receptors to drug discovery. J Med Chem. 2000 Feb 24;43(4):527-50.

[2]. PPAR-α Agonist WY-14643 Inhibits LPS-Induced Inflammation in Synovial Fibroblasts via NF-kB Pathway. J Mol Neurosci. 2016 Aug;59(4):544-53.

[3]. PPARα Agonist WY-14643 Induces SIRT1 Activity in Rat Fatty Liver Ischemia-Reperfusion Injury. Biomed Res Int. 2015;2015:894679.

Pirinixic acid belongs to the pyrimidine, organochlorine, and aryl thioether classes of compounds, and its function is related to acetic acid. Pirini acid is a synthetic thioacetic acid derivative used in biomedical research and is carcinogenic. Pirini acid is a peroxisome proliferator that activates specific peroxisome proliferation-activating receptors (PPARs). PPARs play important roles in various cellular functions, including lipid metabolism, cell proliferation, differentiation, adipogenesis, and inflammatory signaling. (NCI04)

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Mechanism of Action
In rodents, the effects of various peroxisome proliferators on peroxisome function, hepatomegaly, hepatocellular carcinoma, and lipid metabolism have been extensively studied. However, little is known about the effects of these drugs (some of which are used as lipid-lowering agents) on various metabolic parameters in humans. We investigated the effects of clofibrate, di(2-ethylhexyl) phthalate (DEHP), and pirininic acid (WY-14,643) on phospholipid metabolism in cultured human fibroblasts. Clofibrate inhibited the incorporation of [1-(14)C]hexadecyl alcohol and [1-(14)C]linolenic acid into ethanolamine phosphate in a time- and concentration-dependent manner; the labeling of acetal phospholipids and non-acetal phospholipid ethanolamine phosphate was reduced by 40-80% compared to the generally 10-30% inhibition of other phospholipids, including phosphatidylcholine. In pulse and pulse-tracking experiments, the selective inhibition of [1,2-(14)C]ethanolamine incorporation compared to [methyl-(3)H]choline confirmed the relative specificity of the inhibition of ethanolamine phosphate. In control and mutant (Zellweger syndrome and adrenoleukodystrophy) fibroblasts, DEHP and WY-14,643 showed similar concentration-dependent and specific inhibitory effects on phospholipid turnover, and had no significant effect on peroxisome markers. These observations suggest that peroxisome proliferators specifically inhibit ethanolamine phosphate glycerol turnover in human fibroblasts and should be considered when evaluating the efficacy and safety of such drugs as lipid-lowering agents, or when assessing the cellular mechanism of action of proliferators. Pirnicotinic acid (Wy-14,643) is an agonist of the peroxisome proliferation-activating receptor (PPAR) α subtype, capable of exerting beneficial effects on a variety of inflammation-related processes in a slow, long-term manner. Our recent studies have shown that α-substituted pirnicotinic acid derivatives are PPARα agonists and can act as dual inhibitors of 5-lipoxygenase (5-LO, EC 1.13.11.34) and microsomal prostaglandin E2 synthase-1 (EC 5.3.99.3). This study investigated the short-term effects of α-substituted pirininic acid derivatives on typical neutrophil function induced by N-formylmethionine leucylphenylalanine (fMLP), including leukotriene generation, reactive oxygen species (ROS) production, and human leukocyte elastase (EC 3.4.21.37) release, and examined the regulation of related signaling pathways. Pirininic acid derivatives with alkyl substitution at the carboxyl α-position and 6-aminoquinoline substitution on the pyrimidine ring inhibited fMLP-induced leukotriene generation, ROS production, and leukocyte elastase release. Simultaneously, Ca²⁺ mobilization and phosphorylation (activation) of p38 mitogen-activated protein kinase were significantly reduced, while phosphorylation of extracellular signal-regulated kinase 2 remained unaffected. Pirininic acid itself showed no or only weak activity in all these assays. In conclusion, targeted structural modification of pirininic acid can yield bioactive compounds that exhibit immediate anti-inflammatory properties in human neutrophils, suggesting potential therapeutic value.
The normal function of peroxisome proliferator-activated receptor α (PPARα) is crucial for regulating hepatic fatty acid metabolism. Fatty acids are ligands of PPARα, and when fatty acid levels are elevated, PPARα activation induces the expression of a series of fatty acid metabolic enzymes, thereby restoring fatty acid levels to normal. Ethanol intake leads to elevated hepatic fatty acid levels. However, in vitro experiments have shown that ethanol metabolism inhibits the ability of PPARα to bind to DNA and activate reporter genes. This phenomenon has been further confirmed in mice. In C57BL/6J mice, continuous ethanol intake for four weeks also inhibited fatty acid catabolism in the liver due to the blockade of the PPARα-mediated response. Ethanol feeding reduced the level of retinoic acid X receptor α (RXRα) in liver nuclear extracts, as well as the binding capacity of PPARα/RXR to its consensual sequence. Furthermore, in the livers of ethanol-fed animals, the mRNA levels of several PPARα regulatory genes were decreased [long-chain acyl-CoA (acyl-CoA) dehydrogenase and medium-chain acyl-CoA dehydrogenase] or were not induced (acyl-CoA dehydrogenase, hepatic carnitine palmitoyl-CoA transferase I, very long-chain acyl-CoA synthase, very long-chain acyl-CoA dehydrogenase). Consistent with these results, ethanol feeding did not induce the rate of fatty acid β-oxidation in liver homogenate. Dietary supplementation with the PPARα agonist WY14,643 restored PPARα/RXR DNA-binding activity, induced the mRNA levels of multiple PPARα target genes, stimulated the rate of fatty acid β-oxidation in liver homogenate, and prevented fatty liver in ethanol-fed animals. Blockage of PPARα function during ethanol intake leads to alcoholic fatty liver disease, while WY14,643 can reverse this process. Endothelial injury is a major event in the development of atherosclerosis, followed by monocyte infiltration, macrophage differentiation, and smooth muscle cell migration. Peroxisome proliferator-activated receptors (PPARs) are transcription factors currently considered important mediators of inflammatory responses. This study aimed to establish a human endothelial cell model to evaluate the anti-inflammatory properties of PPAR activators. PPAR proteins (α, δ, and γ) were expressed in EAhy926 endothelial cells (ECs). Pirini acid (Wy-14643), fenofibrate, fenofibrate, Merck's ligand PPARδ activator L-165041, and 15-deoxy-Δ(12,14)-prostaglandin J2, as well as rosiglitazone (BRL-49653), all inhibited the induced expression of vascular cell adhesion molecule-1 (VCAM-1) (detected by enzyme-linked immunosorbent assay [ELISA]) and the binding of monocytes to activated EAhy926 cells. PPARδ activator L-165041 showed the strongest ability to inhibit cytokine-induced secretion of monocyte chemoattractant protein-1 (MCP-1). All tested PPAR activators inhibited VCAM-1 expression and significantly reduced intranuclear p65 levels. These results indicate that EAhy926 endothelial cells are an effective tool for confirming and characterizing the inflammatory effects of PPAR activators. For more complete data on the mechanisms of action of pirlinic acid (out of 10), please visit the HSDB record page. Ischemia-reperfusion injury (IRI) remains a common complication during surgery, especially in patients with fatty liver disease due to reduced tolerance to IRI. In addition to its important role in metabolism, activation of peroxisome proliferator-activated receptor α (PPARα) is associated with a positive effect on IRI. Furthermore, the deacetylase SIRT1 has recently emerged as a promising target for the prevention of IRI because it interacts with stress-related mechanisms such as endoplasmic reticulum stress (ERS). Therefore, this study aimed to investigate whether the PPARα agonist WY-14643 could protect fatty liver disease from IRI via the SIRT1 and ERS signaling pathways. Obese Zucker rats were pretreated or untreated with WY-14643 (10 mg/kg, intravenously) followed by partial (70%) hepatic ischemia (1 hour) and 24-hour reperfusion. Liver injury (ALT levels), lipid peroxidation (MDA), SIRT1 activity, SIRT1 and SIRT3 protein expression, and ERS-related parameters (IRE1α, peIF2, caspase 12, and CHOP) were assessed. WY-14643 treatment reduced fatty liver injury, enhanced SIRT1 activity, and inhibited ERS. Our results collectively suggest that the PPARα agonist WY-14643 may exert its protective effect against fatty liver, at least in part, by inducing SIRT1 and preventing endoplasmic reticulum stress (ERS). [3]

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1. Background and Classification:
- WY-14643 (pirenic acid) is a synthetic, selective PPARα agonist that was initially developed as a tool compound to study the physiological functions of PPARα (a nuclear receptor that regulates fatty acid oxidation, lipid metabolism, and anti-inflammatory responses). [1]
- It has been widely used in preclinical studies to investigate the role of PPARα in metabolic disorders (e.g., fatty liver) and inflammatory diseases (e.g., rheumatoid arthritis-associated synovitis). [1,2,3]
2. Mechanism of action:
- Anti-inflammatory effect: Activates PPARα to inhibit the NF-κB signaling pathway—inhibits p65 phosphorylation and upregulates IκBα, thereby reducing the secretion of pro-inflammatory cytokines (TNF-α, IL-6) in LPS-induced synovial fibroblasts[2]
- Hepatoprotective effect: Activates PPARα to induce SIRT1 activity in fatty liver I/R injury—SIRT1 promotes the deacetylation of metabolic and stress-related proteins, reduces oxidative stress (reduces MDA, increases SOD), and alleviates hepatocyte necrosis[3]
- PPARα-mediated metabolic regulation: Enhances the expression of PPARα downstream genes (e.g., acyl-CoA oxidase, carnitine palmitoyltransferase 1) to promote fatty acid β-oxidation and improve lipid metabolism disorders[1]
3. Research use:
- Research use: as a validation of PPARα The gold standard tool for dependent biological processes such as lipid metabolism regulation and anti-inflammatory response [1]

C14H14CLN3O2S

323.8

323.049

C, 51.93; H, 4.36; Cl, 10.95; N, 12.98; O, 9.88; S, 9.90

50892-23-4

5694

Typically exists as White to off-white solids at room temperature

1.4±0.1 g/cm3

514.4±50.0 °C at 760 mmHg

155°C

264.9±30.1 °C

0.0±1.4 mmHg at 25°C

1.658

4.92

2

6

5

21

361

0

ClC1C([H])=C(N=C(N=1)SC([H])([H])C(=O)O[H])N([H])C1=C([H])C([H])=C([H])C(C([H])([H])[H])=C1C([H])([H])[H]

SZRPDCCEHVWOJX-UHFFFAOYSA-N

InChI=1S/C14H14ClN3O2S/c1-8-4-3-5-10(9(8)2)16-12-6-11(15)17-14(18-12)21-7-13(19)20/h3-6H,7H2,1-2H3,(H,19,20)(H,16,17,18)

[[4-chloro-6-[(2,3-dimethylphenyl)amino]-2-pyrimidinyl]thio]-acetic acid

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

Solubility in Formulation 1: ≥ 2.08 mg/mL (6.42 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: ≥ 2.08 mg/mL (6.42 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

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Solubility in Formulation 3: ≥ 2.08 mg/mL (6.42 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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 (Please use freshly prepared in vivo formulations for optimal results.)