Discovery of selective glucocorticoid receptor modulator MK-5932
Department of Medicinal Chemistry, Merck Research Laboratories, 770 Sumneytown Pike, West Point, PA 19486, USA.Bioorganic & medicinal chemistry (Impact Factor: 2.79). 10/2011; 19(24):7374-86. DOI: 10.1016/j.bmc.2011.10.054
A series of partial agonists of the Glucocorticoid Receptor were prepared targeting reduced transactivation activity, while maintaining significant transrepression activity. Incorporation of an ortho-aryl amide produced compounds with the desired in vitro profile. Bioreactors consisting of Suspension cultures of Sf21 cells co expressing a CYP3A4 and NADPH-cytochrome P450 oxireductase were used to prepare the major metabolites of these compounds and revealed that oxidative N-dealkylation provided a pathway for formation of metabolites that were more agonistic than the parent partial agonists. Oxidative N-dealkylation was blocked in a new series of compounds, however oxidation alone was capable of producing full agonist metabolites. Incorporation of an ortho-primary amide and utilization of fluorine to modulate agonism afforded partial agonist MK-5932. Synthesis of the major metabolites of MK-5932 using bioreactor technology revealed that no significant GR-active metabolites were formed. Orally administered MK-5932 displayed anti-inflammatory efficacy in a Rat Oxazolone-induced chronic dermatitis model, while sparing plasma insulin.
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ABSTRACT: Glucocorticoids are used widely in the treatment of inflammatory diseases, but use is accompanied by a significant burden of adverse effects. It has been hypothesized that gene- and cell-specific regulation of the glucocorticoid receptor by small molecule ligands could be translated into a therapeutic with an improved risk-benefit profile. MK-5932 is a highly selective glucocorticoid receptor modulator that is anti-inflammatory in vivo with an improved profile on glucose metabolism: Bungard (2011) Bioorg. Med. Chem. 19, 7374-86. Here we describe the full biological profile of MK-5932. Cytokine production following lipopolysaccharide (LPS) challenge was blocked by MK-5932 in both rat and human whole blood. Oral administration reduced inflammatory cytokine levels in the serum of rats challenged with LPS. MK-5932 was anti-inflammatory in a rat contact dermatitis model, but was differentiated from 6-methylprednisolone by a lack of elevation of fasting insulin or glucose levels after 7 days of dosing, even at high exposure levels. In fact, animals in the vehicle group were consistently hyperglycemic at the end of the study, and MK-5932 normalized glucose levels in a dose-dependent manner. MK-5932 was also anti-inflammatory in the rat collagen-induced arthritis and adjuvant-induced arthritis models. In healthy dogs, oral administration of MK-5932 exerted acute pharmacodynamic effects with potency comparable to prednisone, but with important differences on neutrophil counts, again suggestive of a dissociated profile. Important gaps in our understanding of mechanism of action remain, but MK-5932 will be a useful tool in dissecting the mechanisms of glucose dysregulation by therapeutic glucocortiocids.
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ABSTRACT: TNF expression is elevated in asthma and other inflammatory airway diseases that are commonly treated with glucocorticoid-based therapies, but the impact of glucocorticoids on negative feedback control of TNF is not well understood. We analyzed the effect of dexamethasone, a potent synthetic glucocorticoid, on TNF-regulated gene expression in cultured airway epithelial cells. Although dexamethasone-mediated activation of the glucocorticoid receptor (GR) potently repressed expression of IL1β, IL8, and several other pro-inflammatory TNF targets, the expression of anti-inflammatory TNF targets such as TNFAIP3 (A20) and NFKBIA was selectively spared or augmented by dexamethasone treatment. Despite divergent effects on gene expression, GR and NF-κB occupancy at the TNFAIP3 locus and GR-repressed targets was similar. A co-occupied intronic TNFAIP3 regulatory element mediated cooperative enhancement of transcription by GR and NF-κB that required the presence of a functional GR binding site (GBS). GBS exchanges between reporters for TNFAIP3 and FKBP5, a canonical GR-induced target, revealed substantial latitude in the GBS sequence requirements for GR/NF-κB cooperation, suggesting that the TNFAIP3 GBS acts primarily as a docking site in this context. Supporting this notion, a selective GR ligand with only weak agonist activity for induction of FKBP5 enabled robust GR/NF-κB cooperative induction of a mutant TNFAIP3 reporter harboring the FKBP5 GBS. Taken together, our data support a model in which the expression of anti-inflammatory targets of TNF is maintained during treatment with glucocorticoids through context-dependent cooperation between GR and NF-κB.
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ABSTRACT: The molecular mechanism of boswellic acids (BAs) responsible for their anti-inflammatory features might be attributable to interference with the human glucocorticoid receptor (GR). Due to obvious structural similarities with glucocorticoids (GCs), we conducted pharmacophore studies as well as molecular docking simulations of BAs as putative ligands at the ligand binding site (LBS) of the GR in distinct functional states. In order to verify receptor binding and functional activation of the GR by BAs, radiometric binding assays as well as GR response element-dependent luciferase reporter assay were performed with dexamethasone (DEX) as a functional positive control. With respect to the observed position of GCs in GR crystal complexes in the active antagonist state, BAs docked in a flipped orientation with estimated binding constants reflecting nanomolar affinities. For validation, DEX and other steroids were successfully re-docked into their crystal poses in similar ranges as reported in the literature. In line with the pharmacophore and docking models the BAs were strong GR binders (radiometric binding assay), albeit none of the BAs activated the GR in the reporter gene assay, when compared to the GC agonist DEX. The flipped scaffolds of all BAs dislodge the known C-11 function from its receiving amino acid (Asn564) which may explain the silencing effects of receptor-bound BAs in the reporter gene assay. Together, our results constitute a compelling example of rigid keys acting in an adaptable lock qualifying as a reversed induced fit mechanism, thereby extending the hitherto published knowledge about molecular target interactions of BAs.
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