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Review Articles |
1 Endocrine Sciences Research Group, Room 3-903, Stopford Building, University of Manchester, Manchester M13 9PT, UK
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(Received 24 October 2006;
accepted after revision 16 November 2006; first published online 30 November 2006)
Corresponding author D. W. Ray: Endocrine Sciences Research Group, Room 3-903, Stopford Building, University of Manchester, Manchester M13 9PT, UK. Email: david.w.ray{at}man.ac.uk
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Introduction |
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Glucocorticoid production by the adrenal cortex is tightly controlled by the brain, via activation of hypothalamic neurones, which secrete corticotrophin releasing factor (CRF) into the hypophyseal portal system, and that in turn induces secretion of ACTH from pituitary corticotroph cells. ACTH is the final common pathway, resulting in both adrenal hyperplasia and also augmented hormone production, principally cortisol in humans, corticosterone in rodents. This axis is under glucocorticoid negative feedback at the pituitary, hypothalamic and higher centre levels. Clinical manifestations of altered glucocorticoid sensitivity typically arise because either isolated tissues or organs acquire a differential sensitivity threshold to the central glucocorticoid sensors or, in the face of generalized resistance to glucocorticoid action, the increased tone of the hypothalamic–pituitary–adrenal (HPA) axis gives rise to production of abnormal quantities of other adrenal steroids with a discrete pattern of effects, such as is seen in congenital glucocorticoid resistance.
Glucocorticoids are widely used to treat a wide variety of allergic and inflammatory diseases, including rheumatoid arthritis, inflammatory bowel disease, leukaemia and Hodgkin's disease. Various synthetic glucocorticoid molecules are available for therapeutic use. Although the chemical structures of these steroids are based on natural corticosteroids, changes have been made in order to optimize therapeutic potential and minimize adverse reaction. Understanding the mechanism of action of glucocortcoids has allowed therapeutics to be designed with specific pharmacokinetics and pharmacodynamics (absorption factor, half-life, volume of distribution). However, long-term use of glucocorticoids can lead to pleiotropic side-effects, including fat redistribution, obesity and osteoporosis. This review highlights some of the novel glucocorticoid receptor ligands that are being developed in order to achieve minimal side-effects while maintaining maximal, typically anti-inflammatory, efficacy.
Structure of GR
The GR is member of the nuclear receptor superfamily that includes mineralocorticoid, thyroid hormone, retinoic acid and vitamin D receptors. The GR is located at chromosome 5q31–32 and consists of nine exons, which are highly conserved across species (Stolte et al. 2006). Like all steroid receptors, the GR consists of a variable N-terminal domain, a DNA binding domain with two zinc finger motifs, a hinge region, and a C-terminal hormone binding domain (Fig. 1).
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, there is a C-terminal variant, GRß, which does not bind ligand, and may have a dominant negative action on GR. A further C-terminal truncated GR variant, GR
or GR-P, is principally expressed in cancer, and a constitutive splice variant, GR
, has an additional arginine in the DNA binding domain. The function of these last two variants is still uncertain (Yudt et al. 2003; Lu & Cidlowski, 2004). Recently, it was reported that multiple proteins are translated from the GR transcript, giving rise to further diversity in protein expression (Lu & Cidlowski, 2005). These protein variants have been shown to have differential activity on gene regulation, and also are expressed at different abundance in different cell types. The glucocorticoid receptor in its inactive state is predominantly found in the cytoplasm of target cells. It forms a complex consisting of the receptor polypeptide, two molecules of HSP90, one molecule of HSP70, and one molecule of HSP56, which is an immunophilin of the cyclosporin-, FK506- and rapamycin-binding classes. This receptor complex is stabilized by protein–protein interaction and maintains high affinity of the receptor for its ligand (Galigniana et al. 1999; Galigniana et al. 2001; Guo et al. 2001).
Action of GR
Glucocorticoids, both natural (cortisol in humans, corticosterone in rodents) and synthetic (e.g. prednisolone and dexamethasone), are lipophilic and gain access to cells by diffusion across the plasma membrane. Within target cells, glucocorticoids are subject to metabolism by 11ß-hydroxysteroid dehydrogenase (Seckl, 2004; Tomlinson et al. 2004). This enzyme exists in two principal isoforms. The type 1 enzyme acts predominantly to generate the active glucocorticoid cortisol from inactive cortisone. This enzyme is predominantly expressed in liver and adipose tissue and so acts not only to increase the circulating concentration of active glucocorticoid but can also act in a tissue-specific manner to amplify glucocorticoid action (Stewart, 2003; Tomlinson et al. 2004; Seckl, 2004; Kershaw et al. 2005). The type 2 enzyme predominantly acts in the opposite direction. This results in inactivation of cortisol by oxidation to the inactive cortisone. The tissue distribution of the type 2 enzyme is restricted to mineralocorticoid target tissues, notably the renal tubule (Seckl et al. 2002; Paterson et al. 2005; Holmes et al. 2006).
The GR molecule binds glucocorticoids and transactivates or transrepresses glucocorticoid-responsive promoters (Fig. 2). After binding glucocorticoid, the receptor–ligand complex undergoes a conformational change, thus releasing the HSP complex and homodimerizing with another activated GR molecule. The activated GR interacts with the importin system and translocates via the nuclear pore into the nucleus, to regulate gene expression (Hager, 2002; Elbi et al. 2004; Nagaich et al. 2004). In the nucleus, GR binds to glucocorticoid response elements (GREs) and subsequently recruits coactivators to the DNA in order for gene transcription to occur. The first GREs analysed were associated with enhanced transcription; however, there are several examples of ‘negative’ GREs (nGREs), as described in the pro-opiomelanocortin (Drouin et al. 1993), osteocalcin (Meyer et al. 1997) and prolactin promoters (Sakai et al. 1988), which are associated with repression of transcription. The GR molecule acting as a monomer, in contrast, modulates the transcription rates of non-GRE-containing genes by interacting with nuclear transcription factors, including activator protein 1 (AP1), nuclear factor 
B (NFB) and signal transducer and activator of transcription 5 (STAT5; Rogatsky et al. 2002, 2003; Wang et al. 2004; Fig. 2).
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, GRß does not bind to glucocorticoids and functions as a weak dominant negative inhibitor of GR(Yudt et al. 2003). This action is mediated by GRE binding, since no protein–protein interaction has been described. The physiological relevance of GRß is still under investigation (Chrousos et al. 1996; Leung et al. 1997; Hamid et al. 1999; Shahidi et al. 1999; Vottero & Chrousos, 1999; Hauk et al. 2000; Leung & Chrousos, 2000; Strickland et al. 2001). Glucocorticoid receptors and chromatin
When glucocorticoid receptors associate with target chromatin, a series of co-ordinated, timed protein recruitment events are promoted (Baumann et al. 2001; Stavreva et al. 2004). These result in assembly of macromolecular complexes on chromatin, which remodel the chromatin in such a way that its structure becomes more open and promotes further complex assembly leading to recruitment of RNA polymerase 2 and initiation of gene transcription (Abruzzese et al. 2000; Baumann et al. 2001; Han et al. 2006; Nawaz & O'Malley, 2004; O'Malley, 2005; Jung et al. 2005; Wu et al. 2005a). A large number of these comodulator proteins and complexes have now been defined, and it has been shown that altered expression of such comodulators can alter sensitivity to steroid hormones (Baumann et al. 2001; Cheskis et al. 2003; Stevens et al. 2003; Lonard et al. 2004). There is currently considerable interest in understanding how altered expression of the comodulators can titrate hormone sensitivity, but interestingly, it appears that the crucial regulatory step determining the final intracellular concentration of these factors is post-translational rather than transcriptional (O'Malley, 2004; Wu et al. 2005b; Ying et al. 2005).
Glucocorticoid receptor cross-talk with other signalling pathways
Glucocorticoids do not typically act in isolation. A number of lines of evidence support important combinatorial signalling events involving the glucocorticoid receptor and the intracellular signalling cascades initiated by a number of transmembrane receptors. In particular, activation of the stress-responsive Jun N-terminal kinase (JNK) results in phosphorylation of the glucocorticoid receptor N-terminal domain (Krstic et al. 1997; Wang et al. 2002; Wang & Garabedian, 2003; Ismaili & Garabedian, 2004; Szatmary et al. 2004). When the receptor is so phosphorylated, it impairs nuclear translocation, and the ability of the receptor to increase target gene transcription is reduced. The full implications of this cross-talk between JNK and glucocorticoid sensitivity have not been explored in a physiological or pathophysiological context (Fig. 2).
A further important example of cross-talk between glucocorticoid receptor function and other signalling cascades is direct physical interaction between the glucocorticoid receptor and other transcription factors (Fig. 2). The best characterized examples of these are NFB and AP1. Nuclear factor B is a heterodimeric complex of transcription factors that share the Rel homology domain. It has been shown that the p65 component, otherwise known as Rel A, is capable of directly binding to the glucocorticoid receptor. This physical interaction results in mutual inhibition of transcriptional activation of the two factors. For example, the glucocorticoid receptor will inhibit p65-dependent gene transcription and equally p65 overexpression inhibits gene activation by the glucocorticoid receptor. Since NFB is activated downstream of the proinflammatory cytokines tumour necrosis factor (TNF) and interleukin-1 (IL-1) and forms an important part of the innate immune response, its activation may act to prevent the anti-inflammatory activities of glucocorticoids within foci of active inflammation (Nissen & Yamamoto, 2000; Rogatsky et al. 2002; Garside et al. 2004).
A further example of transcription factor cross-talk is seen between the glucocorticoid receptor and AP1 (Fig. 2). The AP1 transcription factor is activated as a consequence of mitogen-activated protein (MAP) kinase pathways, and was the first documented example of transcription factor cross-talk involving the glucocorticoid receptor. The mode of action is complex, involving inhibition of GR and AP1 activity. In the case of the well-studied proliferin gene, a rather complex interaction occurs, with some heterodimeric complexes of AP1 being potentiated by the glucocorticoid receptor and others being inhibited (Reichardt et al. 1998, 2001; Tuckermann et al. 1999).
It has been recognized that interleukin-2 (IL-2) resulted in a glucocorticoid-resistant state within its target cells. More recently, the basis for this interaction has been defined to be due to interaction between glucocorticoid receptor and STAT5 transcription factor, which is activated downstream of IL-2 receptor activation. STAT5 is either capable of productive synergy with the glucocorticoid receptor on some target genes, particularly milk protein genes and gene expression in the liver where it is activated by prolactin or growth hormone (Groner, 2002), but in a cell type- and target gene-specific way is also capable of inhibiting glucocorticoid actions, as demonstrated within cells of the lymphoid lineage in response to IL-2 (Biola et al. 2001; Goleva et al. 2002). In addition, Il-4 is also capable of inducing glucocorticoid resistance in target cells by activation of the related transcription factor STAT6 (Biola et al. 2000; Nelson et al. 2003).
Physiological effects of glucocorticoids
Glucocorticoid receptors seem to be present in all cell types, resulting in the steroid hormones having a diverse range of effects on physiological systems, including endocrine, renal, immune and neural systems. The pleiotropic hormone helps to maintain homeostasis during challenges such as haemorrhage, metabolic disturbances, infection and anxiety. The diverse effects of glucocorticoids on various systems are demonstrated with excessive glucocorticoid levels resulting from drug administration, which cause iatrogenic Cushing's syndrome or hyperadrenocorticism (Brown et al. 1977; Flower et al. 1986; Gallant & Kenny, 1986; Laue et al. 1988; Belvisi et al. 2001a; Crim et al. 2001; Scribano & Prantera, 2003; Purdy & Wiley, 2004). The best known and studied effects of glucocorticoids are on carbohydrate metabolism and immune function.
Effects on inflammation and immune function. Glucocorticoids have potent anti-inflammatory and immunosuppressive properties. Increased cytokine activity present during inflammation not only activates components of the inflammatory system but also triggers the HPA axis to initiate glucocorticoid production. This produces a negative feedback loop, with the glucocorticoids reducing inflammatory processes, via the mechanisms previously outlined, of a range of cell types including T-cells, macrophages and neutrophils (Guyre et al. 1984; Cupps et al. 1985; Cohan et al. 1989). Therapeutically, glucocorticoids are used in a range of inflammatory diseases and physiologically they are crucially important in normal immune responses.
Effects on metabolism. The name glucocorticoid is derived from initial observations that glucose metabolism is affected by the hormones. During fasting, cortisol stimulates several processes that initiate mechanisms to increase and maintain normal concentrations of glucose in blood, as follows.
Stimulation of gluconeogenesis. This pathway results in the synthesis of glucose from other non-hexose organic molecules, such as pyruvate, lactate, glycerol and amino acids. Glucocorticoids increase the expression of several key enzymes in gluconeogenesis and increase the availability of amino acids essential for the process (Pilkis & Granner, 1992).
Glucose conservation. Glucose is conserved for neural tissues and uptake is inhibited for muscle and adipose tissues (McMahon et al. 1988).
Lipolysis in adipose tissue. The fatty acids released by lipolysis are used for production of energy in tissues such as muscle, and the released glycerol provides another substrate for gluconeogenesis. Glucocorticoids also inhibit the action of leptin, an important hormone in body weight regulation and reproductive function (Zakrzewska et al. 1997).
Other effects of glucocorticoids. Glucocorticoids influence bone and cartilage formation by changing expression release of several mediators, including insulin-like growth factor 1 (IGF-1), IGF-binding protein, growth hormones, and thyroid hormones. Excessive glucocorticoid levels result in osteoporosis and impair skeletal growth and bone formation by inhibiting osteoblasts and collagen synthesis (Robson et al. 2002).
A variety of glucocorticoid effects are evident in the cardiovascular system, including the regulation of blood pressure and vascular reactivity. Glucocorticoids play an important role in the regulation of blood pressure by increasing the vascular reactivity to vasoactive substances such as angiotensin II and noradrenaline. The importance of this regulation can be seen in adrenalectomized animals and patients with glucocorticoid deficiency, where hypotension and reduced sensitivity to vasoconstrictors is evident.
Cognitive function is also known to be influenced by glucocorticoids and effect glucocorticoid secretion. Several reports demonstrate the importance of the hormone in the formation of fear memory. Studies have shown that glucocorticoid levels after conditioning were correlated with fear conditioning levels, and adrenalectomy can reduce the unconditioned freezing behaviour of newborn mice (Cordero et al. 1998; Cordero & Sandi, 1998).
Glucocorticoid receptor structure and function
The structures of numerous nuclear receptor ligand binding domains have been published and these have helped rational drug design. For example, the crystal structure of the oestrogen receptor bound to both agonist and antagonist ligands has indicated the critical importance of the final protein conformation for determining the ultimate activity of the receptor. The crystal structures of all of the steroid receptors solved to date indicates a conserved 12 -helical structure. It appears that the hydrophobic ligand is buried within a hydrophobic core and that the structure of the ligand alters how the -helices compact around the ligand and, in particular, affects the final position of helix 12. The position of helix 12 in the final ligand-bound conformation is very different in the presence of an agonist compared with an antagonist. It is thought that the final position of helix 12 alters the specificity of comodulator binding to the ligand binding domain of the steroid receptor. In this way, when an agonist is bound, coactivator proteins are attracted which ultimately result in induction of gene transcription. In contrast, when an antagonist is bound, a repressor protein, for example nuclear receptor corepressor 1 (NcoR), is recruited and this results in inhibition of gene transcription. Although the glucocorticoid receptor was the first member of the nuclear receptor superfamily to be cloned and the first human transcription factor to be cloned, its crystal structure was not solved until much more recently (Bledsoe et al. 2002). Initially, structural models were built based on the high degree of amino acid homology between the progesterone receptor ligand binding domain and the glucocorticoid receptor. These models were highly informative and were largely supported by the subsequent crystallization studies (Ray et al. 1999). However, it is very hard to use models based on crystallization of an agonist-bound receptor to predict the final conformation of the receptor when bound to an antagonist. Therefore, it was very important to examine the crystal structure in the presence of an antagonist, and this was achieved more recently (Kauppi et al. 2003). This crystal structure showed that, as predicted, the final position of helix 12 was very different compared with that seen with a full agonist. In fact, part of helix 12 finally lay in a position which partly covered the binding sites for the coactivator protein, thereby explaining why the antagonist-bound receptor was unable to activate target gene transcription. Further functional analysis identified that not only did the antagonist-bound glucocorticoid receptor fail to recruit coactivators, as would be predicted from the crystal structure, but in fact antagonist-bound glucocorticoid receptor actively recruited the repressor protein NCoR. By homology studies, it was possible to identify amino acids within the ligand binding domain of the glucocorticoid receptor which would be capable of generating a charge clamp that would stabilize the nuclear receptor interacting -helix present on the corepressor protein NcoR (Garside et al. 2004).
The structural and functional studies of the glucocorticoid receptor ligand binding domain performed in the presence of agonist or antagonist ligands has shed new light on how the receptor interprets its ligands. However, it is interesting that a purely synthetic ligand such as RU486 can result in recruitment of a corepressor protein such as NCoR. It remains to be seen whether there is in fact an endogenous ligand of the glucocorticoid receptor which is capable of promoting this interaction. Indeed, studies from other receptors, notably the oestrogen receptor, have shown that antagonist ligands not only result in blockade (albeit partial) of agonist gene transcription events but also upregulate a subset of antagonist specific genes. This is really quite an unexpected finding and indicates the importance of considering cell response to a ligand as an integrated whole rather than picking individual genes and extrapolating from them.
Novel glucocorticoid receptor ligands
Identification of compounds that select transactivation or transrepression. Glucocorticoids are potent anti-inflammatory agents but also demonstrate a wide range of effects on other tissues. Recently, there has been interest in trying to identify compounds which retain the beneficial effects but do not have any of the undesirable side-effects. Efforts to date have identified compounds which can be grouped into a number of different classes (Rosen & Miner, 2005).
Selective glucocorticoid receptor modulator. A selective glucocorticoid receptor modulator describes a compound which retains useful anti-inflammatory activity but with impaired activity to influence bone metabolism or glucose and lipid metabolism. These compounds are therefore predicted to have an improved therapeutic index when used in vivo.
Gene-selective compound. This refers to molecules that act on the receptor to influence gene expression in a gene-specific fashion. These compounds would result in a different profile on the expressed transcriptome compared with conventional glucocorticoids.
Dissociated compounds. This refers to a class of molecules that completely dissociate transactivation from transrepression by the glucocorticoid receptor. Such compounds should fail to transactivate gene expression globally within target cells but retain the ability to globally repress transcription on the glucocorticoid repressible genes.
Soft steroids. These are a class of corticosteroids that have a particular focus of activity but which are very rapidly inactivated by enzymes, thereby reducing their ability to exert effects as a result of carriage in the systematic circulation. Such agents would typically be used in a topical or inhaled fashion.
Dissociated glucocorticoids.
The first reports of a truly dissociating glucocorticoid receptor ligand used compounds based on a steroid core made by the Roussel Uclaf organization. The group were able to show significant differences between a test compound and conventional glucocorticoids using cell-based in vitro assays. The initial data suggested efficient inhibition of both AP1- and NFB-mediated gene induction but negligible transactivation activity on several genes (Vayssiere et al. 1997). However, these initial promising findings were not replicated by other investigators (Garside et al. 2004). In a number of assays, transactivation was similar to conventional full agonist ligands such as hydrocortisone. Furthermore, in vivo they were found to have the same side-effect as steroids (Belvisi et al. 2001b). There was no therapeutic advantage of using this group of compound in comparison with a conventional glucocorticoid. It may well be that significant effects were genuinely being generated by the novel compounds, since even now reports are being published suggesting that this same range of molecules is capable of a clearly different spectrum of activities compared with conventional glucocorticoids in specific cell types (Tanigawa et al. 2002; Humphrey et al. 2006).
It is important to note that compounds with the opposite profile, that is strong transactivation but weak repression, have also been described. These compounds have no anti-inflammatory activity. This in vivo and in vitro pharmacology was supplemented by the important observations stemming from genetic study. Using a sophisticated knock-out knock-in approach, Schutz was able to show that a dimerization-deficient glucocorticoid receptor, which was unable to transactivate a reporter gene but did not affect repression, responded to conventional glucocorticoids when anti-inflammatory end-points were studied (Reichardt et al. 1998, 2001). This result strongly supports the idea that repression alone may be sufficient for anti-inflammatory activity.
The variable results found in some of the initially analysed compounds suggest that some of them were in fact gene selective and perhaps also subject to cell type-specific factors and therefore not truly dissociated. There are further examples of how selective steroid receptor ligands can function in this way, and a partial explanation for the mechanism has been identified as an expression level of critical comodulator proteins, including p300 and cAMP response element-binding (CREB) binding protein. This observation does make the task of developing and characterizing truly dissociated compound much more difficult. It is clearly insufficient to base the analysis on the regulation of one or two index genes, but rather the entire effect on the transcriptome needs to be compared between compounds. It is likely that none of the compounds currently in development would fulfil this criterion, but it is possible with iterative drug design that such compound structures will be identified in the future.
It has been known for some time that the dose dependence of different gene regulatory events varies. For example, the concentration of dexamethasone required to repress co-inflammatory cytokine expression in cells in culture is typically an order of magnitude less than that required to activate target genes. Therefore, on this basis, even a conventional glucocorticoid such as dexamethasone could be regarded as having partially dissociated activity. It is important that effects on transactivation and transrepression are directly compared to ensure that a global loss of potency of the steroid does not mislead one into believing that true selectivity has been obtained. The problem that such incomplete analysis can cause is best exemplified by the steroid deflazacort. This was initially marketed clinically as having beneficial anti-inflammatory activity with decreased impact on bone and glucose metabolism. Indeed, at the manufacturers' suggested dose equivalents it did appear to be relatively free of side-effects compared with prednisolone. However, the dose equivalents were not adequately supported by experimental data. Thus, when large-scale trials using biologically equivalent doses of deflazacort were conducted, the advantages of deflazacort over prednisolone were essentially lost (Markham & Bryson, 1995).
Novel dissociating ligands. A number of compounds have been identified as a result of drug discovery activities both by biotechnology companies and large pharmaceutical enterprises. Some of these compounds have utility as tool compounds but none as yet has reached the clinic.
Ligand pharmaceuticals have developed a non-steroidal series of molecules based on the ability to promote DNA binding of the glucocorticoid receptor. One compound, AL082D06, was further characterized. Using in vitro cell culture systems, this compound was an antagonist of dexamethasone on transactivation end-points and was found to bind the glucocorticoid receptor with specificity but not particularly high affinity. The molecule was also found to antagonize dexamethasone inhibition of both TNF- and IL-1ß-induced selectin expression. Therefore, the molecule was a full antagonist, unlike the partial antagonist RU486, with potential utility in blocking the global effects of glucocorticoids resulting from, for example, Cushing's syndrome (Rosen & Miner, 2005).
Further work on the series of compounds identified a potentially selective, non-steroidal glucocorticoid receptor modulator, AL-438. This compound was characterized in vitro using cell line models and was found to have anti-inflammatory activity on rat carrageenan-induced paw oedema. AL-438 also had identical binding affinity for the glucocorticoid receptor compared with prednisolone but only had weak agonist activity on a classical transactivation assay. Importantly, in an in vivo study, AL-438 did not result in impaired glucose tolerance, in contrast to bioequivalent anti-inflammatory doses of prednisolone. Therefore, AL-438 appeared to retain the desirable anti-inflammatory activities of prednisolone but to be free of the hyperglycaemic side-effects of full glucocorticoid receptor agonists. Since glucocorticoid induced osteoporosis is a major drawback to long-term treatment in humans, the effects of AL-438 on bone metabolism were also studied. In this assay, mineralizing bone formation was measured in vivo. Prednisolone was found to reduce bone mineralization rate but, in contrast, AL-438 had no suppressive effect. Therefore, the compound had a promising lack of side-effects measured not only with glucose metabolism but also with bone (Coghlan et al. 2003).
A further promising molecular structure was identified by a Belgian group from the Namibian shrub Salsola tubercultiformis. This molecular structure, termed compound A, lacks a steroid core but was found to efficiently downregulate NFB-driven genes. This effect was dependent on its binding to the glucocorticoid receptor. In contrast, and perhaps most interestingly, however, it completely failed to stimulate transactivation of target genes, suggesting that it was indeed a dissociating compound. Both compound A and dexamethasone induced nuclear translocation of the glucocorticoid receptor. The mechanism of action of compound A appears to be by promoting the glucocorticoid receptor to interfere with the transactivation potential of the p65 component of NFB. There appears to be a dual mechanism of action involving both a reduction in in vivo DNA binding capacity as well as a reduction in the transactivation potential of NFB. In vivo evidence supports the role of compound A as an effective anti-inflammatory agent with equivalent potency to dexamethasone. However, importantly, there were no hyperglycaemic side-effects seen with compound A. Therefore, this compound also holds promise as a lead molecule for further development of therapeutics (De et al. 2005).
A further series of compounds based on an arylpyrazole structure have recently been published. These compounds bind the glucocorticoid receptor with specificity and relatively high affinity, similar to that seen with conventional agonists such as prednisolone or dexamethasone. These compounds differed in their relative activity to inhibit cell-based assays of proliferation, adipocyte differentiation or osteoblast differentiation. Furthermore, transcriptome profiles of their relative activities on a set of index glucocorticoid-regulated genes were assessed. The different molecular structures had differential effects on individual target genes. It was striking that very small changes in structure of the ligand caused markedly distinct glucocorticoid receptor regulatory effects in more than one cell line. Further analysis suggested that the different compounds altered the relative affinity of the glucocorticoid receptor for specific DNA sequences. Therefore, when chromatin immunoprecipitation studies were performed to look at GR occupancy, in vivo differences were seen. The induced structure of the ligand binding domain of the glucocorticoid receptor appears to influence the interaction with DNA sequence and thereby specify a distinct profile of gene regulatory events (Wang et al. 2006). This study overall supports the idea that pursuit of the perfect dissociating glucocorticoid ligand may well be complicated, but it is certainly possible that even an imperfectly dissociating compound may be more than sufficient to offer an improved therapeutic index and thereby unleash the full anti-inflammatory potency of glucocorticoids without the side-effect profile.
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References |
|---|
|
TopAbstractIntroductionReferences |
|---|
Baumann CT, Ma H, Wolford R, Reyes JC, Maruvada P, Lim C, Yen PM, Stallcup MR & Hager GL (2001). The glucocorticoid receptor interacting protein 1 (GRIP1) localizes in discrete nuclear foci that associate with ND10 bodies and are enriched in components of the 26S proteasome. Mol Endocrinol 15, 485–500.
Belvisi MG, Brown TJ, Wicks S & Foster ML (2001a). New Glucocorticosteroids with an improved therapeutic ratio? Pulm Pharmacol Ther 14, 221–227.[CrossRef][Medline]
Belvisi MG, Wicks SL, Battram CH, Bottoms SE, Redford JE, Woodman P, Brown TJ, Webber SE & Foster ML (2001b). Therapeutic benefit of a dissociated glucocorticoid and the relevance of in vitro separation of transrepression from transactivation activity. J Immunol 166, 1975–1982.
Biola A, Andreau K, David M, Sturm M, Haake M, Bertoglio J & Pallardy M (2000). The glucocorticoid receptor and STAT6 physically and functionally interact in T lymphocytes. FEBS Lett 487, 229–233.[CrossRef][Medline]
Biola A, Lefebvre P, Perrin-Wolff M, Sturm M, Bertoglio J & Pallardy M (2001). Interleukin-2 inhibits glucocorticoid receptor transcriptional activity through a mechanism involving STAT5 (signal transducer and activator of transcription 5) but not AP-1. Mol Endocrinol 15, 1062–1076.
Bledsoe RK, Montana VG, Stanley TB, Delves CJ, Apolito CJ, McKee DD, Consler TG, Parks DJ, Stewart EL, Willson TM, Lambert MH, Moore JT, Pearce KH & Xu HE (2002). Crystal structure of the glucocorticoid receptor ligand binding domain reveals a novel mode of receptor dimerization and coactivator recognition. Cell 110, 93–105.[CrossRef][Medline]
Brown HM, Storey G & Jackson FA (1977). Beclomethasone dipropionate aerosol in treatment of perennial and seasonal rhinitis: a review of five years' experience. Br J Clin Pharmacol 4 (Suppl. 3), 283S–286S.
Cheskis BJ, McKenna NJ, Wong CW, Wong J, Komm B, Lyttle CR & O'Malley BW (2003). Hierarchical affinities and a bipartite interaction model for estrogen receptor isoforms and full-length steroid receptor coactivator (SRC/p160) family members. J Biol Chem 278, 13271–13277.
Chrousos GP, Castro M, Leung DY, Webster E, Kino T, Bamberger C, Elliot S, Stratakis C & Karl M (1996). Molecular mechanisms of glucocorticoid resistance/hypersensitivity. Potential clinical implications. Am J Respir Crit Care Med 154, S39–S43.[Medline]
Coghlan MJ, Jacobson PB, Lane B, Nakane M, Lin CW, Elmore SW, Kym PR, Luly JR, Carter GW, Turner R, Tyree CM, Hu J, Elgort M, Rosen J & Miner JN (2003). A novel antiinflammatory maintains glucocorticoid efficacy with reduced side effects. Mol Endocrinol 17, 860–869.
Cohan VL, Undem BJ, Fox CC, Adkinson NF Jr, Lichtenstein LM & Schleimer RP (1989). Dexamethasone does not inhibit the release of mediators from human mast cells residing in airway, intestine, or skin. Am Rev Respir Dis 140, 951–954.[Medline]
Cordero MI, Merino JJ & Sandi C (1998). Correlational relationship between shock intensity and corticosterone secretion on the establishment and subsequent expression of contextual fear conditioning. Behav Neurosci 112, 885–891.[CrossRef][Medline]
Cordero MI & Sandi C (1998). A role for brain glucocorticoid receptors in contextual fear conditioning: dependence upon training intensity. Brain Res 786, 11–17.[CrossRef][Medline]
Crim C, Pierre LN & Daley-Yates PT (2001). A review of the pharmacology and pharmacokinetics of inhaled fluticasone propionate and mometasone furoate. Clin Ther 23, 1339–1354.[CrossRef][Medline]
Cupps TR, Gerrard TL, Falkoff RJ, Whalen G & Fauci AS (1985). Effects of in vitro corticosteroids on B cell activation, proliferation, and differentiation. J Clin Invest 75, 754–761.[Medline]
De BK, Vanden BW, Van Beck IMMW, Hennuyer N, Hapgood J, Libert C, Staels B, Louw A & Haegeman G (2005). A fully dissociated compound of plant origin for inflammatory gene repression. Proc Natl Acad Sci U S A 102, 15827–15832.
Drouin J, Sun YL, Chamberland M, De Gauthier YLA, Nemer M & Schmidt TJ (1993). Novel glucocorticoid receptor complex with DNA element of the hormone-repressed POMC gene. EMBO J 12, 145–156.[Medline]
Elbi C, Walker DA, Romero G, Sullivan WP, Toft DO, Hager GL & DeFranco DB (2004). Molecular chaperones function as steroid receptor nuclear mobility factors. Proc Natl Acad Sci U S A 101, 2876–2881.
Flower RJ, Parente L, Persico P & Salmon JA (1986). A comparison of the acute inflammatory response in adrenalectomised and sham-operated rats. Br J Pharmacol 87, 57–62.
Galigniana MD, Housley PR, DeFranco DB & Pratt WB (1999). Inhibition of glucocorticoid receptor nucleocytoplasmic shuttling by okadaic acid requires intact cytoskeleton. J Biol Chem 274, 16222–16227.
Galigniana MD, Radanyi C, Renoir JM, Housley PR & Pratt WB (2001). Evidence that the peptidylprolyl isomerase domain of the hsp90-binding immunophilin FKBP52 is involved in both dynein interaction and glucocorticoid receptor movement to the nucleus. J Biol Chem 276, 14884–14889.
Gallant C & Kenny P (1986). Oral glucocorticoids and their complications. A review. J Am Acad Dermatol 14, 161–177.[Medline]
Garside H, Stevens A, Farrow S, Normand C, Houle B, Berry A, Maschera B & Ray D (2004). Glucocorticoid ligands specify different interactions with NF-B by allosteric effects on the glucocorticoid receptor DNA binding domain. J Biol Chem 279, 50050–50059.
Goleva E, Kisich KO & Leung DY (2002). A role for STAT5 in the pathogenesis of IL-2-induced glucocorticoid resistance. J Immunol 169, 5934–5940.
Groner B (2002). Transcription factor regulation in mammary epithelial cells. Domest Anim Endocrinol 23, 25–32.[CrossRef][Medline]
Guo Y, Guettouche T, Fenna M, Boellmann F, Pratt WB, Toft DO, Smith DF & Voellmy R (2001). Evidence for a mechanism of repression of heat shock factor 1 transcriptional activity by a multichaperone complex. J Biol Chem 276, 45791–45799.
Guyre PM, Bodwell JE & Munck A (1984). Glucocorticoid actions on lymphoid tissue and the immune system: physiologic and therapeutic implications. Prog Clin Biol Res 142, 181–194.[Medline]
Hager GL (2002). The dynamics of intranuclear movement and chromatin remodeling by the glucocorticoid receptor. Ernst Schering Res Found Workshop 40, 111–129.
Hamid QA, Wenzel SE, Hauk PJ, Tsicopoulos A, Wallaert B, Lafitte JJ, Chrousos GP, Szefler SJ & Leung DY (1999). Increased glucocorticoid receptor ß in airway cells of glucocorticoid-insensitive asthma. Am J Respir Crit Care Med 159, 1600–1604.
Han SJ, Demayo FJ, Xu J, Tsai SY, Tsai MJ & O'Malley BW (2006). Steroid receptor coactivator (SRC)-1 and SRC-3 differentially modulate tissue-specific activation functions of the progesterone receptor. Mol Endocrinol 20, 45–55.
Hauk PJ, Hamid QA, Chrousos GP & Leung DY (2000). Induction of corticosteroid insensitivity in human PBMCs by microbial superantigens. J Allergy Clin Immunol 105, 782–787.[CrossRef][Medline]
Holmes MC, Sangra M, French KL, Whittle IR, Paterson J, Mullins JJ & Seckl JR (2006). 11ß-Hydroxysteroid dehydrogenase type 2 protects the neonatal cerebellum from deleterious effects of glucocorticoids. Neuroscience 137, 865–873.[CrossRef][Medline]
Humphrey EL, Williams JH, Davie MW & Marshall MJ (2006). Effects of dissociated glucocorticoids on OPG and RANKL in osteoblastic cells. Bone 38, 652–661.[Medline]
Ismaili N & Garabedian MJ (2004). Modulation of glucocorticoid receptor function via phosphorylation. Ann N Y Acad Sci 1024, 86–101.
Jung SY, Malovannaya A, Wei J, O'Malley BW & Qin J (2005). Proteomic analysis of steady-state nuclear hormone receptor coactivator complexes. Mol Endocrinol 19, 2451–2465.
Kauppi B, Jakob C, Farnegardh M, Yang J, Ahola H, Alarcon M, Calles K, Engstrom O, Harlan J, Muchmore S, Ramqvist AK, Thorell S, Ohman L, Greer J, Gustafsson JA, Carlstedt-Duke J & Carlquist M (2003). The three-dimensional structures of antagonistic and agonistic forms of the glucocorticoid receptor ligand-binding domain: RU-486 induces a transconformation that leads to active antagonism. J Biol Chem 278, 22748–22754.
Kershaw EE, Morton NM, Dhillon H, Ramage L, Seckl JR & Flier JS (2005). Adipocyte-specific glucocorticoid inactivation protects against diet-induced obesity. Diabetes 54, 1023–1031.
Krstic MD, Rogatsky I, Yamamoto KR & Garabedian MJ (1997). Mitogen-activated and cyclin-dependent protein kinases selectively and differentially modulate transcriptional enhancement by the glucocorticoid receptor. Mol Cell Biol 17, 3947–3954.[Abstract]
Laue L, Kawai S, Brandon DD, Brightwell D, Barnes K, Knazek RA, Loriaux DL & Chrousos GP (1988). Receptor-mediated effects of glucocorticoids on inflammation: enhancement of the inflammatory response with a glucocorticoid antagonist. J Steroid Biochem 29, 591–598.[CrossRef][Medline]
Leung DY & Chrousos GP (2000). Is there a role for glucocorticoid receptor ß in glucocorticoid-dependent asthmatics? Am J Respir Crit Care Med 162, 1–3.
Leung DY, Hamid Q, Vottero A, Szefler SJ, Surs W, Minshall E, Chrousos GP & Klemm DJ (1997). Association of glucocorticoid insensitivity with increased expression of glucocorticoid receptor ß. J Exp Med 186, 1567–1574.
Lonard DM, Tsai SY & O'Malley BW (2004). Selective estrogen receptor modulators 4-hydroxytamoxifen and raloxifene impact the stability and function of SRC-1 and SRC-3 coactivator proteins. Mol Cell Biol 24, 14–24.
Lu NZ & Cidlowski JA (2004). The origin and functions of multiple human glucocorticoid receptor isoforms. Ann N Y Acad Sci 1024, 102–123.
Lu NZ & Cidlowski JA (2005). Translational regulatory mechanisms generate N-terminal glucocorticoid receptor isoforms with unique transcriptional target genes. Mol Cell 18, 331–342.[CrossRef][Medline]
McMahon M, Gerich J & Rizza R (1988). Effects of glucocorticoids on carbohydrate metabolism. Diabetes Metab Rev 4, 17–30.[Medline]
Markham A & Bryson HM (1995). Deflazacort. A review of its pharmacological properties and therapeutic efficacy. Drugs 50, 317–333.[Medline]
Meyer T, Gustafsson JA & Carlstedt-Duke J (1997). Glucocorticoid-dependent transcriptional repression of the osteocalcin gene by competitive binding at the TATA box. DNA Cell Biol 16, 919–927.[Medline]
Nagaich AK, Walker DA, Wolford R & Hager GL (2004). Rapid periodic binding and displacement of the glucocorticoid receptor during chromatin remodeling. Mol Cell 14, 163–174.[CrossRef][Medline]
Nawaz Z & O'Malley BW (2004). Urban renewal in the nucleus: is protein turnover by proteasomes absolutely required for nuclear receptor-regulated transcription? Mol Endocrinol 18, 493–499.
Nelson G, Wilde GJC, Spiller DG, Kennedy SM, Ray DW, Sullivan E, Unitt JF & White MRH (2003). NF-B signalling is inhibited by glucocorticoid receptor and STAT6 via distinct mechanisms. J Cell Sci 116, 2495–2503.
Nissen RM & Yamamoto KR (2000). The glucocorticoid receptor inhibits NFB by interfering with serine-2 phosphorylation of the RNA polymerase II carboxy-terminal domain. Genes Dev 14, 2314–2329.
O'Malley BW (2004). Results of a search for the mechanisms of steroid receptor regulation of gene expression. Ann N Y Acad Sci 1038, 80–87.
O'Malley BW (2005). A life-long search for the molecular pathways of steroid hormone action. Mol Endocrinol 19, 1402–1411.
Paterson JM, Seckl JR & Mullins JJ (2005). Genetic manipulation of 11ß-hydroxysteroid dehydrogenases in mice. Am J Physiol Regul Integr Comp Physiol 289, R642–R652.
Pilkis SJ & Granner DK (1992). Molecular physiology of the regulation of hepatic gluconeogenesis and glycolysis. Annu Rev Physiol 54, 885–909.[CrossRef][Medline]
Purdy IB & Wiley DJ (2004). Perinatal corticosteroids: A review of research. Part I: Antenatal administration. Neonatal Netw 23, 15–30.[Medline]
Ray DW, Suen CS, Brass A, Soden J & White A (1999). Structure/function of the human glucocorticoid receptor: tyrosine 735 is important for transactivation. Mol Endocrinol 13, 1855–1863.
Reichardt HM, Kaestner KH, Tuckermann J, Kretz O, Wessely O, Bock R, Gass P, Schmid W, Herrlich P, Angel P & Schutz G (1998). DNA binding of the glucocorticoid receptor is not essential for survival. Cell 93, 531–541.[CrossRef][Medline]
Reichardt HM, Tuckermann JP, Gottlicher M, Vujic M, Weih F, Angel P, Herrlich P & Schutz G (2001). Repression of inflammatory responses in the absence of DNA binding by the glucocorticoid receptor. EMBO J 20, 7168–7173.[CrossRef][Medline]
Robson H, Siebler T, Shalet SM & Williams GR (2002). Interactions between GH, IGF-I, glucocorticoids, and thyroid hormones during skeletal growth. Pediatr Res 52, 137–147.[CrossRef][Medline]
Rogatsky I, Luecke HF, Leitman DC & Yamamoto KR (2002). Alternate surfaces of transcriptional coregulator GRIP1 function in different glucocorticoid receptor activation and repression contexts. Proc Natl Acad. Sci U S A 99, 16701–16706.
Rogatsky I, Wang JC, Derynck MK, Nonaka DF, Khodabakhsh DB, Haqq CM, Darimont BD, Garabedian MJ & Yamamoto KR (2003). Target-specific utilization of transcriptional regulatory surfaces by the glucocorticoid receptor. Proc Natl Acad Sci U S A 100, 13845–13850.
Rosen J & Miner JN (2005). The search for safer glucocorticoid receptor ligands. Endocr Rev 26, 452–464.
Sakai DD, Helms S, Carlstedt-Duke J, Gustafsson JA, Rottman FM & Yamamoto KR (1988). Hormone-mediated repression: a negative glucocorticoid response element from the bovine prolactin gene. Genes Dev 2, 1144–1154.
Scribano M & Prantera C (2003). Review article: medical treatment of moderate to severe Crohn's disease. Aliment Pharmacol Ther 17 (Suppl. 2), 23–30.
Seckl JR (2004). 11ß-Hydroxysteroid dehydrogenases: changing glucocorticoid action. Curr Opin Pharmacol 4, 597–602.[CrossRef][Medline]
Seckl JR, Yau J & Holmes M (2002). 11ß-Hydroxysteroid dehydrogenases: a novel control of glucocorticoid action in the brain. Endocr Res 28, 701–707.[CrossRef][Medline]
Shahidi H, Vottero A, Stratakis CA, Taymans SE, Karl M, Longui CA, Chrousos GP, Daughaday WH, Gregory SA & Plate JM (1999). Imbalanced expression of the glucocorticoid receptor isoforms in cultured lymphocytes from a patient with systemic glucocorticoid resistance and chronic lymphocytic leukemia. Biochem Biophys Res Commun 254, 559–565.[CrossRef][Medline]
Stavreva DA, Muller WG, Hager GL, Smith CL & McNally JG (2004). Rapid glucocorticoid receptor exchange at a promoter is coupled to transcription and regulated by chaperones and proteasomes. Mol Cell Biol 24, 2682–2697.
Stevens A, Garside H, Berry A, Waters C, White A & Ray D (2003). Dissociation of steroid receptor coactivator 1 and nuclear receptor corepressor recruitment to the human glucocorticoid receptor by modification of the ligand-receptor interface: the role of tyrosine 735. Mol Endocrinol 17, 845–859.
Stewart PM (2003). Tissue-specific Cushing's syndrome, 11ß-hydroxysteroid dehydrogenases and the redefinition of corticosteroid hormone action. Eur J Endocrinol 149, 163–168.[Abstract]
Stolte EH, van Kemenade BM, Savelkoul HF & Flik G (2006). Evolution of glucocorticoid receptors with different glucocorticoid sensitivity. J Endocrinol 190, 17–28.
Strickland I, Kisich K, Hauk PJ, Vottero A, Chrousos GP, Klemm DJ & Leung DY (2001). High constitutive glucocorticoid receptor ß in human neutrophils enables them to reduce their spontaneous rate of cell death in response to corticosteroids. J Exp Med 193, 585–593.
Szatmary Z, Garabedian MJ & Vilcek J (2004). Inhibition of glucocorticoid receptor-mediated transcriptional activation by p38 mitogen-activated protein (MAP) kinase. J Biol Chem 279, 43708–43715.
Tanigawa K, Tanaka K, Nagase H, Miyake H, Kiniwa M & Ikizawa K (2002). Cell type-dependent divergence of transactivation by glucocorticoid receptor ligand. Biol Pharm Bull 25, 1619–1622.[CrossRef][Medline]
Tomlinson JW, Walker EA, Bujalska IJ, Draper N, Lavery GG, Cooper MS, Hewison M & Stewart PM (2004). 11ß-Hydroxysteroid dehydrogenase type 1: a tissue-specific regulator of glucocorticoid response. Endocr Rev 25, 831–866.
Tuckermann JP, Reichardt HM, Arribas R, Richter KH, Schutz G & Angel P (1999). The DNA binding-independent function of the glucocorticoid receptor mediates repression of AP-1-dependent genes in skin. J Cell Biol 147, 1365–1370.
Vayssiere BM, Dupont S, Choquart A, Petit F, Garcia T, Marchandeau C, Gronemeyer H & Resche-Rigon M (1997). Synthetic glucocorticoids that dissociate transactivation and AP-1 transrepression exhibit antiinflammatory activity in vivo. Mol Endocrinol 11, 1245–1255.
Vottero A & Chrousos GP (1999). Glucocorticoid receptor ß: View I. Trends Endocrinol Metab 10, 333–338.[CrossRef][Medline]
Wang JC, Derynck MK, Nonaka DF, Khodabakhsh DB, Haqq C & Yamamoto KR (2004). Chromatin immunoprecipitation (ChIP) scanning identifies primary glucocorticoid receptor target genes. Proc Natl Acad Sci U S A 101, 15603–15608.
Wang Z, Frederick J & Garabedian MJ (2002). Deciphering the phosphorylation "code" of the glucocorticoid receptor in vivo. J Biol Chem 277, 26573–26580.
Wang Z & Garabedian MJ (2003). Modulation of glucocorticoid receptor transcriptional activation, phosphorylation, and growth inhibition by p27Kip1. J Biol Chem 278, 50897–50901.
Wang JC, Shah N, Pantoja C, Meijsing SH, Ho JD, Scanlan TS & Yamamoto KR (2006). Novel arylpyrazole compounds selectively modulate glucocorticoid receptor regulatory activity. Genes Dev 20, 689–699.
Wu HY, Hamamori Y, Xu J, Chang SC, Saluna T, Chang MF, O'Malley BW & Kedes L (2005a). Nuclear hormone receptor coregulator GRIP1 suppresses, whereas SRC1A and p/CIP coactivate, by domain-specific binding of MyoD. J Biol Chem 280, 3129–3137.
Wu RC, Smith CL & O'Malley BW (2005b). Transcriptional regulation by steroid receptor coactivator phosphorylation. Endocr Rev 26, 393–399.
Ying H, Furuya F, Willingham MC, Xu J, O'Malley BW & Cheng SY (2005). Dual functions of the steroid hormone receptor coactivator 3 in modulating resistance to thyroid hormone. Mol Cell Biol 25, 7687–7695.
Yudt MR, Jewell CM, Bienstock RJ & Cidlowski JA (2003). Molecular origins for the dominant negative function of human glucocorticoid receptor ß. Mol Cell Biol 23, 4319–4330.
Zakrzewska KE, Cusin I, Sainsbury A, Rohner-Jeanrenaud F & Jeanrenaud B (1997). Glucocorticoids as counterregulatory hormones of leptin: toward an understanding of leptin resistance. Diabetes 46, 717–719.[Abstract]
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