Fluorine-substituted ligands for the peroxisome proliferator-activated receptor gamma (PPARgamma): potential imaging agents for metastatic tumors

The peroxisome proliferator-activated receptor gamma (PPARgamma), a primary regulator of lipid metabolism, is present in many tumor cell lines and animal tumor systems and, in some cases, can mediate effective antitumor therapy with potent synthetic ligands. In an approach to image tumors with positron-emission tomography (PET) based on their content of PPARgamma, we have synthesized two fluorine-substituted analogues of a high affinity ligand from the phenylpropanoic acid class. The analogue having the highest affinity for PPARgamma was labeled with the positron-emitting radionuclide fluorine-18. In tissue distribution studies in normal rats and in SCID mice bearing human breast tumor xenografts, this compound did not show evidence of receptor-mediated uptake. The prospects for using PPARgamma as a target for imaging tumors may be limited by the low receptor concentrations in tumors and by the pharmacokinetic behavior of this class of ligands, which appears to be more favorable for therapy than for imaging.     

Strategies for the labeling of halogen-substituted peroxisome proliferator-activated receptor gamma ligands: potential positron emission tomography and single photon emission computed tomography imaging agents

Well-known as an important regulator of lipid metabolism and adipocyte differentiation, the peroxisome proliferator-activated receptor gamma (PPARgamma) also has potential use as a target for antitumor therapy in certain cancers. To develop agents for radionuclide imaging PPARgamma in vivo, we synthesized fluorine, bromine, and iodine-substituted analogs (1-3) of a high-affinity benzophenone-tyrosine PPARgamma ligand; all three analogs retain very high affinity for the PPARgamma receptor. In preparation for the synthesis of these PPARgamma ligands in radiolabeled form, we have synthesized two types of precursors: (a) an aryltributylstannane (9), from which the bromine and iodine-substituted analogs (2 and 3) can readily be prepared by electrophilic destannylation, and (b) three diaryliodonium tosylate derivatives (12a-c), precursors for nucleophilic aromatic fluorination using fluoride ion. Conditions were developed whereby the thiophenyliodonium tosylate (12c) underwent nucleophilic aromatic substitution with fluoride ion, efficiently and in short reaction times, to produce the desired fluorine-substituted target compound 1. These reactions laid the groundwork for producing these three PPARgamma ligands in radiolabeled form; in addition, our use of diaryliodonium ion precursors for aromatic fluorination in this series provides an example that should encourage application of this approach for radiofluorination of more complicated radiopharmaceuticals.     

PPARadigms and PPARadoxes: expanding roles for PPARgamma in the control of lipid metabolism

The nuclear receptor PPARgamma is a central regulator of adipose tissue development and an important modulator of gene expression in a number of specialized cell types including adipocytes, epithelial cells, and macrophages. PPARgamma signaling pathways impact both cellular and systemic lipid metabolism and have links to obesity, diabetes, and cardiovascular disease. The ability to activate this receptor with small molecule ligands has made PPARgamma an attractive target for intervention in human metabolic disease. As our understanding of PPARgamma biology has expanded, so has the therapeutic potential of PPARgamma ligands. Recent studies have provided insight into the paradoxical relationship between PPARgamma and metabolic disease and established new paradigms for the control of lipid metabolism. This review focuses on recent advances in PPARgamma biology in the areas of adipocyte differentiation, insulin resistance, and atherosclerosis.     

Regulated production of a peroxisome proliferator-activated receptor-gamma ligand during an early phase of adipocyte differentiation in 3T3-L1 adipocytes

Peroxisome proliferator-activated receptor-gamma (PPARgamma) is a nuclear hormone receptor that is critical for adipogenesis and insulin sensitivity. Ligands for PPARgamma include some polyunsaturated fatty acids and prostanoids and the synthetic high affinity antidiabetic agents thiazolidinediones. However, the identity of a biologically relevant endogenous PPARgamma ligand is unknown, and limited insight exists into the factors that may regulate production of endogenous PPARgamma ligands during adipocyte development. To address this question, we created a line of 3T3-L1 preadipocytes that carry a beta-galactosidase-based PPARgamma ligand-sensing vector system. In this system, induction of adipogenesis resulted in elevated beta-galactosidase activity that signifies activation of PPARgamma via its ligand-binding domain (LBD) and suggests generation and/or accumulation of a ligand moiety. The putative endogenous ligand appeared early in adipogenesis in response to increases in cAMP, accumulated in the medium, and dissipated later in adipogenesis. Organically extracted and high pressure liquid chromatography-fractionated conditioned media from differentiating cells, but not from mature adipocytes, were enriched in this activity. One or more components within the organic extract activated PPARgamma through interaction with its LBD, induced lipid accumulation in 3T3-L1 cells as efficiently as the differentiation mixture, and competed for binding of rosiglitazone to the LBD of PPARgamma. The active species appears to be different from other PPARgamma ligands identified previously. Our findings suggest that a novel biologically relevant PPARgamma ligand is transiently produced in 3T3-L1 cells during adipogenesis.     

Novel peroxisome proliferator-activated receptor (PPAR) gamma and PPARdelta ligands produce distinct biological effects

The peroxisome proliferator-activated receptors (PPARs) include three receptor subtypes encoded by separate genes: PPARalpha, PPARdelta, and PPARgamma. PPARgamma has been implicated as a mediator of adipocyte differentiation and the mechanism by which thiazolidinedione drugs exert in vivo insulin sensitization. Here we characterized novel, non-thiazolidinedione agonists for PPARgamma and PPARdelta that were identified by radioligand binding assays. In transient transactivation assays these ligands were agonists of the receptors to which they bind. Protease protection studies showed that ligand binding produced specific alterations in receptor conformation. Both PPARgamma and PPARdelta directly interacted with a nuclear receptor co-activator (CREB-binding protein) in an agonist-dependent manner. Only the PPARgamma agonists were able to promote differentiation of 3T3-L1 preadipocytes. In diabetic db/db mice all PPARgamma agonists were orally active insulin-sensitizing agents producing reductions of elevated plasma glucose and triglyceride concentrations. In contrast, selective in vivo activation of PPARdelta did not significantly affect these parameters. In vivo PPARalpha activation with WY-14653 resulted in reductions in elevated triglyceride levels with minimal effect on hyperglycemia. We conclude that: 1) synthetic non-thiazolidinediones can serve as ligands of PPARgamma and PPARdelta; 2) ligand-dependent activation of PPARdelta involves an apparent conformational change and association of the receptor ligand binding domain with CREB-binding protein; 3) PPARgamma activation (but not PPARdelta or PPARalpha activation) is sufficient to potentiate preadipocyte differentiation; 4) non-thiazolidinedione PPARgamma agonists improve hyperglycemia and hypertriglyceridemia in vivo; 5) although PPARalpha activation is sufficient to affect triglyceride metabolism, PPARdelta activation does not appear to modulate glucose or triglyceride levels.     

PPAR(alpha) and PPAR(gamma) activators suppress the monocyte-macrophage apoB-48 receptor

Certain triglyceride-rich lipoproteins (TRLs), specifically chylomicrons, dyslipemic VLDLs, and their remnants, are atherogenic and can induce monocyte-macrophage foam cell formation in vitro via the apolipoprotein B-48 receptor (apoB-48R). Human atherosclerotic lesion foam cells express the apoB-48R, as determined immunohistochemically, suggesting it can play a role in the conversion of macrophages into foam cells in vivo. The regulation of the apoB-48R in monocyte-macrophages is not fully understood, albeit previous studies indicated that cellular sterol levels and state of differentiation do not affect apoB-48R expression. Since peroxisome proliferator-activated receptors (PPARs) regulate some aspects of cellular lipid metabolism and may be protective in atherogenesis by up-regulation of liver X-activated receptor alpha and ATP-binding cassette transporter A1, we examined the regulation of apoB-48R by PPAR ligands in human monocyte-macrophages. Using real-time PCR, Northern, Western, and functional cellular lipid accumulation assays, we show that PPARalpha and PPARgamma activators significantly suppress the expression of apoB-48R mRNA in human THP-1 and blood-borne monocyte-macrophages. Moreover, PPAR activators inhibit the expression of the apoB-48R protein and, notably, the apoB-48R-mediated lipid accumulation of TRL by THP-1 monocytes in vitro. If PPAR activators also suppress the apoB-48R pathway in vivo, diminished apoB-48R-mediated monocyte-macrophage lipid accumulation may be yet another antiatherogenic effect of the action of PPAR ligands.     

Peroxisome proliferator-activated receptor (PPAR)gamma is expressed by human spermatozoa: its potential role on the sperm physiology

The peroxisome proliferation-activated receptor gamma (PPARgamma) is mainly expressed in the adipose tissue and integrates the control of energy, lipid, and glucose homeostasis. The present study, by means of RT-PCR, Western blot, and immunofluorescence techniques, demonstrates that human sperm express the PPARgamma. The functionality of the receptor was evidenced by 15-deoxy-12,14-prostaglandin J(2) (PGJ2) and rosiglitazone (BRL) PPARgamma-agonists that were tested on capacitation, acrosome reaction, and motility. Both treatments also increase AKT phosphorylations and influence glucose and lipid metabolism in sperm. The specificity of PGJ2 and BRL effects through PPARgamma on human sperm was confirmed by an irreversible PPARgamma antagonist, GW9662. Our findings provide evidence that human sperm express a functional PPARgamma whose activation influences sperm physiology. In conclusion, the presence of PPARgamma in male gamete broadens the field of action of this nuclear receptor, bringing us to look towards sperm as an endocrine mobile unit independent of the systemic regulation.     

Evaluation of 5-(2-(4-pyridinyl)vinyl)-6-chloro-3-(1-methyl-2-(S)-pyrrolidinylmethoxy)pyridine and its analogues as PET radioligands for imaging nicotinic acetylcholine receptors

A novel series of compounds derived from the high-affinity nicotinic acetylcholine receptor (nAChR) ligand, 5-(2-(4-pyridinyl)vinyl)-6-chloro-3-((1-methyl-2-(S)-pyrrolidinyl)methoxy)pyridine (Me-p-PVC), originally developed by Abbott Laboratories, was characterized in vitro in nAChR binding assays at 37 degrees C to show K(i) values in the range of 9-611 pm. Several compounds of this series were radiolabeled with (11)C and evaluated in vivo in mice and monkeys as potential candidates for PET imaging of nAChRs. [(11)C]Me-p-PVC (K(i) =56 pm at 37 degrees C; logD = 1.6) was identified as a radioligand suitable for the in vivo imaging of the alpha 4 beta 2* nAChR subtype. Compared with 2-[(18)F]FA, a PET radioligand that has been successfully used in humans and is characterized by a slow kinetic of brain distribution, [(11)C]Me-p-PVC is more lipophilic. As a result, [(11)C]Me-p-PVC accumulated in the brain more rapidly than 2-[(18)F]FA. Pharmacological evaluation of Me-p-PVC in mice demonstrated that the toxicity of this compound was comparable with or lower than that of 2-FA. Taken together, these results suggest that [(11)C]Me-p-PVC is a promising PET radioligand for studying nAChR occupancy by endogenous and exogenous ligands in the brain in vivo.     

Peroxisome proliferator-activated receptor-gamma gene polymorphisms are not associated with osteonecrosis of the femoral head in the Korean population

Osteonecrosis of the femoral head (ONFH) is a multifactorial disease to which certain individuals are more at risk. Altered lipid metabolism is one of the major risk factors for osteonecrosis, especially corticosteroid therapy and alcoholism. Peroxisome Proliferator-Activated Receptor-gamma (PPARgamma) plays a crucial role in differentiation of mesenchymal cells to adipocytes, lipid homeostasis, and bone metabolism. To investigate the possible association between PPARgamma gene variants and susceptibility to ONFH, we genotyped three common polymorphisms (-796A > G, +34C > G[Pro12Ala], and +82466C > T[His477His]) in 448 ONFH patients and 336 control subjects. Genotypes, allele frequencies, and haplotypes of the polymorphisms in the complete set of patients as well as in subgroups by sex or etiology were not significantly different from those in the control group. This suggests that the examined polymorphisms and haplotypes of the PPARgamma gene are unlikely to be associated with susceptibility to ONFH.