Re-highlighting the action of PPARγ in treating metabolic diseases

Introduction

Insulin resistance is the key pathophysiologic abnormality of many metabolic diseases such as type 2 diabetes mellitus, obesity, dyslipidemia, and cardiovascular diseases¹. Therefore, reducing insulin resistance is the most important strategy for improving metabolic deterioration. Thiazolidinediones (TZDs), peroxisome proliferator-activated receptor γ (PPARγ) agonists, have shown many beneficial effects not only by enhancing insulin sensitivity but also by demonstrating anti-inflammatory and antioxidant properties, whose actions are related to anti-atherosclerosis^2,3. Thus, TZDs were considered a magic bullet for the treatment of type 2 diabetes and atherosclerosis. Indeed, TZDs demonstrated a preventive role for recurrent ischemic stroke in several clinical trials⁴ and for restenosis after percutaneous coronary intervention (PCI)^5–7. However, TZDs increased the risk of peripheral edema, bone loss, and congestive heart failure^8–10. A meta-analysis of clinical trials showed that rosiglitazone significantly increased the risk of myocardial infarction¹¹. Although later studies revealed that rosiglitazone did not increase the risk of heart attack and the US Food and Drug Administration (FDA) removed the warning labels from rosiglitazone-containing drugs regarding the issue of increasing heart attack in 2013, rosiglitazone’s cardiovascular safety issue alongside the above-mentioned adverse effects still lead to many physicians hesitating to prescribe TZDs in their clinical practice. Nevertheless, PPARγ is still one of the most important targets for the treatment of insulin resistance and type 2 diabetes, and novel strategies to modulate PPARγ activity to enhance its beneficial effects and reduce unwanted adverse effects are strongly anticipated. Recent studies showed that post-translational modification (PTM) of PPARγ regulates PPARγ activity or stability and may be a novel way to optimize PPARγ activity with reduced adverse effects. In addition, selective PPARγ modulators (sPPARγMs), dual or pan PPAR agonists, have been developed and tested for their metabolic effects in animal studies and in some clinical trials.

PPARγ, a therapeutic target for insulin resistance (Figure 1)

PPARγ is a master regulator of adipocyte differentiation. It is also involved in glucose homeostasis and insulin sensitivity. The expression of PPARγ is most abundant in adipose tissue¹². Evidence has shown that the primary target of TZDs is adipose tissue, where it increases the expression of Glut4 and CAP¹³, and an animal model lacking PPARγ in adipose tissue had a significantly lower response to TZDs^14,15. TZDs inhibit the expression of TNF-α, IL-6, and resistin in adipose tissue, which promote insulin resistance and chronic inflammation^16,17, while TZDs increased the production of adiponectin and fibroblast growth factor 21 (FGF21), which enhance fatty acid oxidation and insulin sensitivity^18,19. TZDs increase lipogenesis by aP2, LPL, CD36, fatty acid transport protein, PEPCK, and the glycerol transporter aquaporin 7² and make adipose tissue store more lipid, while TZDs remove lipid accumulation in other tissues such as muscle and liver²⁰.

Figure 1. Effect of PPARγ activation on various tissues.

ACSL1, acyl-CoA synthetase long chain family member 1; CD36, cluster of differentiation 36; CPT1b, carnitine palmitoyltransferase 1B; FA, fatty acid; FFA, free fatty acid; FGF, fibroblast growth factor; GK, glucokinase; GLUT, glucose transporter; GlyK, glycerol kinase; IRS2, insulin receptor substrate 1; LPL, lipoprotein lipase; NF-κB, nuclear factor kappa-light-chain-enhancer of activated B cells; MCP-1, monocyte chemoattractant protein 1; MMP-9, matrix metalloproteinase 9; NO, nitric oxide; PAI-1, plasminogen activator inhibitor type 1; PCK2, peroxisome proliferator-activated receptor gamma 2 binding site; PEPCK, phosphoenolpyruvate carboxykinase; PI3K, phosphoinositide 3-kinase; PPARγ, peroxisome proliferator-activated receptor γ; PPRE, peroxisome proliferator-activated receptor response element; RXR, retinoid X receptor; STAT6, signal transducer and activator of transcription 6; TG, triglyceride; TNF-α, tumor necrosis factor α; TZD, thiazolidinedione; VCAM-1, vascular cell adhesion molecule-1; VEGF, vascular endothelial growth factor.

From these studies, it seems that improvement of insulin sensitivity in liver and muscle might be secondary to the effects of TZDs in adipose tissue. However, there is also evidence showing that TZDs have an insulin-sensitizing effect on other peripheral organs. It has been demonstrated that ablation of liver PPARγ in mice reduced hepatic steatosis but worsened hyperlipidemia, triglyceride clearance, and muscle insulin resistance²¹. The expression of PPARγ in skeletal muscle is relatively low compared to adipose tissue, and the physiological significance of PPARγ in skeletal muscle has been shown to work indirectly in previous studies²². However, selective activation of PPARγ in skeletal muscle showed significant protection from high-fat diet-induced insulin resistance and associated changes in muscle phenotype, such as decreasing the quantity of lipid in myocytes and increasing the number of oxidative muscle fiber types²³. It suggests that the activation of PPARγ can act directly on muscle tissue to improve insulin sensitivity. Macrophage PPARγ is also implicated in anti-inflammation and lipid metabolism²⁴, and mice lacking macrophage PPARγ are more prone to whole-body insulin resistance^25,26.

PPARγ agonists and their effects on the vascular system: friend or foe?

PPARγ is expressed in the endothelium and vascular smooth muscle in the blood vessel wall^27,28. Despite controversial cardiovascular effects of TZDs in humans, most experimental studies showed beneficial effects on vascular systems. TZDs inhibit the proliferation and migration of vascular smooth muscle cells (VSMCs), with potential favorable effects on atherosclerosis^29,30. Smooth muscle-specific dominant-negative PPARγ transgenic mice showed a loss of nitric oxide responsiveness and high contractility³¹, which resulted in systolic hypertension. In humans, dominant-negative mutations of PPARγ are associated with early hypertension and insulin resistance³². Activation of PPARγ inhibits CCAAT/enhancer-binding protein-δ (C/EBPδ), which is a well-known mediator of the proinflammatory response in vascular cells³³.

TZDs also reduce activation and inflammation in endothelial cells by suppressing the expression of inflammation-associated genes^34–37. On the other hand, TZDs induce vascular endothelial growth factor (VEGF) in endothelial cells and increase endothelial cell proliferation and migration by the Akt-dependent pathway^38–40. In recent data, rosiglitazone significantly increased endothelial cell migration and vascular leakage in an animal study with increased VEGF expression and suppressed tight junction proteins, which caused instability of the endothelial membrane⁴¹. This result could be related to vascular permeability, peripheral edema, and congestive heart failure associated with the use of TZDs, contrary to their beneficial effect on vascular cells. We still need more concrete evidence to understand the role of TZDs in the whole vascular system under various conditions.

Regulation of PPARγ by PTMs to reduce the side effects of TZDs

The PTM of PPARγ involves several pathways, including phosphorylation, SUMOylation, ubiquitination, β-O-linked N-acetylglucosamine modification (O-GlcNAcylation), and acetylation. These PTMs are known to regulate both PPARγ expression and its transcriptional activity⁴² and have been recently suggested as a good modality for reducing the side effects of PPARγ activation by TZDs⁴³ (Figure 2).

Figure 2. Regulation of PPARγ by post translational modification.

Ac, acetyl; ACSL1, acyl-CoA synthetase long chain family member 1; AF1, activation function 1; AF2, activation function 2; CD36, cluster of differentiation 36; Cdk, cyclin-dependent kinase; CPT1b, carnitine palmitoyltransferase 1B; DBD, DNA-binding domain; FABP3, fatty-acid-binding protein 3, muscle and heart; FAO, fatty acid oxidation; FGF, fibroblast growth factor; GlcNAC, N-acetylglucosamine; LBD, ligand-binding domain; Lys, lysine; MAPK, mitogen-activated protein kinase; P, phosphate; PPARγ, peroxisome proliferator-activated receptor γ; PRDM16, PR domain containing 16; SENP2, small ubiquitin-like modifier-specific protease 2; Ser, serine; SIRT1, sirtuin 1; SUMO, small ubiquitin-like modifier; Thr, threonine; Ub, ubiquitin; WAT, white adipose tissue.

Other PPARγ modulators and agonists

Considering that patients with insulin resistance show many conjugated metabolic problems such as atherosclerosis, obesity, fatty liver, etc., there have been many efforts to develop sPPARγMs, dual or pan PPAR agonists, with potent efficacy but less-deleterious side effects.

sPPARγMs bind to the ligand-binding domain of PPARγ in many ways, which leads to different receptor conformations and cofactor functions⁶⁵. INT131, a potent non-TZD sPPARγM now in clinical trials, showed excellent glucose lowering with significantly less weight gain, edema with fluid retention, and cardiomegaly than do current TZDs⁶⁶. Balaglitazone also showed positive effects in the treatment of patients with type 2 diabetes compared to placebo and pioglitazone (phase III clinical study, n = 409), better glycemic control, and less edema compared to pioglitazone 45 mg⁶⁷. CMHX008 was also tested for its effects in in vitro and in vivo models, showing excellent results by far⁶⁸.

PPARα/γ dual activation has been the focus of new targets from many pharmaceutical companies, and several clinical trials have been performed in potential treatments such as muraglitazar, tesaglitazar, and aleglitazar. However, owing to unpredictable side effects, the studies were all stopped for further development. Unfortunately, muraglitazar increased cardiovascular events, tesaglitazar increased renal toxicity, and aleglitazar showed bone fractures, heart failure, and gastrointestinal side effects^69–71. Recently, (E)-N-(4-(3-(5-bromo-4-hydroxy-2-methoxyphenyl)acryloyl)phenyl)-4-tert-butylbenzamide (SN158) showed anti-diabetic effects through PPARα/γ dual activation. SN158 increased adipogenic differentiation of 3T3-L1 preadipocytes, enhanced fatty acid oxidation in hepatocytes, and increased glucose uptake in myotubes. It lowered plasma glucose and lipid levels in ob/ob mice without severe weight gain. Thus, it represents another candidate PPARα/γ agonist to enhance many metabolic profiles in obesity-related diseases⁷².

There are some natural products which activate PPARγ and PPARα simultaneously or activate the PPARγ dimer partner retinoid X receptor. Compared to full TZDs, these natural products usually show fewer side effects and comparable anti-diabetic effects⁷³. Honokiol, amorfrutin 1, amorfrutin B, amorphastilbol, genistein, biochanin A, sargaquinoic acid, sargahydroquinoic acid, resveratrol, etc. were tested for their efficacy in in vitro and in vivo studies⁷³.

The development of PPARα/γ/δ pan agonists as anti-diabetic, anti-obesity, or hypolipidemic drugs is still actively ongoing^74,75. For example, IVA 337 is a potent and well-balanced pan PPAR agonist which showed promising results in vitro and in vivo and is expected to be used to treat patients with metabolic syndrome and non-alcoholic steatohepatitis⁷⁶.

Conclusion

PPARγ is still one of the most important targets for the treatment of insulin resistance and diabetes mellitus, even though current use of TZDs in clinical practice is limited because of undesirable adverse effects. Thus, novel strategies to modulate PPARγ activity to enhance its beneficial effects and reduce unwanted side effects have been strongly anticipated. Recent advances in understanding how PTM of PPARγ modulates PPARγ activity provide novel ways to optimize PPARγ activity with reduced adverse effects. In addition, selective PPARγ modulators, dual or pan PPAR agonists, have been developed and tested for their metabolic effects in animal studies and in some clinical trials.

We hope safer PPARγ agonists or modulators with excellent efficacy and fewer adverse effects will be available for treating metabolic diseases and insulin resistance in the near future.