Suppression of the inflammatory response in experimental arthritis is mediated via estrogen receptor alpha but not estrogen receptor beta

Introduction: The immune modulatory role of estrogens in inflammation is complex. Both pro- and anti-inflammatory effects of estrogens have been described. Estrogens bind both estrogen receptor (ER)alpha and beta. The contribution of ER alpha and ER beta to ER-mediated immune modulation was studied in delayed type hypersensitivity (DTH) and in experimental arthritis Methods: ER-mediated suppression of rat adjuvant arthritis (AA) was studied using ethinyl-estradiol (EE) and a selective ER beta agonist (ERB-79). Arthritis was followed for 2 weeks. Next, effects of ER agonists (ethinyl-estradiol, an ER alpha selective agonist (ERA-63) and a selective ER beta agonist (ERB-79) on the development of a tetanus toxoid (TT)-specific delayed type hypersensitivity response in wild type (WT) and in ER alpha- or ER beta-deficient mice were investigated. Finally, EE and ERA-63 were tested for their immune modulating potential in established collagen induced arthritis in DBA/1J mice. Arthritis was followed for three weeks. Joint pathology was examined by histology and radiology. Local synovial cytokine production was analyzed using Luminex technology. Sera were assessed for COMP as a biomarker of cartilage destruction. Results: EE was found to suppress clinical signs and symptoms in rat AA. The selective ER beta agonist ERB-79 had no effect on arthritis symptoms in this model. In the TT-specific DTH model, EE and the selective ER alpha agonist ERA-63 suppressed the TT-specific swelling response in WT and ER beta KO mice but not in ER alpha KO mice. As seen in the AA model, the selective ER beta agonist ERB-79 did not suppress inflammation. Treatment with EE or ERA-63 suppressed clinical signs in collagen induced arthritis (CIA) in WT mice. This was associated with reduced inflammatory infiltrates and decreased levels of proinflammatory cytokines in CIA joints. Conclusions: ER alpha, but not ER beta, is key in ER-mediated suppression of experimental arthritis. It remains to be investigated how these findings translate to human autoimmune disease

Org 214007-0: a novel non-steroidal selective glucocorticoid receptor modulator with full anti-inflammatory properties and improved therapeutic index

Contains fulltext : 103595.pdf (publisher's version ) (Open Access)Glucocorticoids (GCs) such as prednisolone are potent immunosuppressive drugs but suffer from severe adverse effects, including the induction of insulin resistance. Therefore, development of so-called Selective Glucocorticoid Receptor Modulators (SGRM) is highly desirable. Here we describe a non-steroidal Glucocorticoid Receptor (GR)-selective compound (Org 214007-0) with a binding affinity to GR similar to that of prednisolone. Structural modelling of the GR-Org 214007-0 binding site shows disturbance of the loop between helix 11 and helix 12 of GR, confirmed by partial recruitment of the TIF2-3 peptide. Using various cell lines and primary human cells, we show here that Org 214007-0 acts as a partial GC agonist, since it repressed inflammatory genes and was less effective in induction of metabolic genes. More importantly, in vivo studies in mice indicated that Org 214007-0 retained full efficacy in acute inflammation models as well as in a chronic collagen-induced arthritis (CIA) model. Gene expression profiling of muscle tissue derived from arthritic mice showed a partial activity of Org 214007-0 at an equi-efficacious dosage of prednisolone, with an increased ratio in repression versus induction of genes. Finally, in mice Org 214007-0 did not induce elevated fasting glucose nor the shift in glucose/glycogen balance in the liver seen with an equi-efficacious dose of prednisolone. All together, our data demonstrate that Org 214007-0 is a novel SGRMs with an improved therapeutic index compared to prednisolone. This class of SGRMs can contribute to effective anti-inflammatory therapy with a lower risk for metabolic side effects

Org 214007-0 does not effect rates of hepatic enzyme fluxes.

<p>Mass Isotopomer Distribution Analysis (MIDA), as described in detail in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0048385#s4" target="_blank">Materials and Methods</a>, was performed in mice treated p.o., once daily, for 7 days with either vehicle, prednisolone (10 mg/kg) or Org 214007-0 (1.5 mg/kg). These doses of each compound are equi-efficacious in suppression of CIA. Neither the glucose-6-phosphatase flux (A) nor the glycogen phosphorylase flux (B) were affected by treatment with prednisolone or Org 214007-0. The glucokinase flux rate (C) was not changed by Org 214007-0, but significantly differed from the effect by prednisolone (##: p = 0.01 <i>vs</i> prednisolone). The glycogen synthase flux rate (D) was significantly decreased by prednisolone (**: p = 0.005 <i>vs</i> vehicle), whereas Org 214007-0 had no significant effect on this flux, but differed significantly from prednisolone (##: p = 0.002 <i>vs</i> prednisolone). Neither Org 214007-0 nor prednisolone, at equi-efficacious dosages, effects the gluconeogenic flux (<i>de novo</i> synthesis of glucose-6-phophate) (E).</p

Org 214007-0 is equally effective as prednisolone in the mouse CIA model.

<p>A) Inhibition of arthritis in the CIA model. Mean clinical score of each group (n = 12), shown as area under the curve (AUC) of the arthritis score monitored every other day during 3 weeks, corrected for baseline, is indicated (± SEM). **  =  significantly different from placebo (p<0.01; ANOVA-test). B) Reduction of bone destruction in the CIA model as measured by X-ray. Mean radiological score (sum of the X-ray scores of left and right hindpaws and knees) of each group of mice (n = 12) at the end of the CIA experiment is indicated (± SEM). **  =  significantly different from placebo (p<0.01; ANOVA-test).</p

Org 214007-0 has a relatively lower impact on induction than on respression of genes.

<p>Fold changes for the top 25 genes either induced (A) or repressed (B) by 1 μM prednisolone and 1 μM Org 214007-0 in THP-1 cells. NB. Scales are in 2log. Example of an induced gene, FK506 binding protein 51 (FKBP51) (C) and a repressed gene, interleukin 6 (IL-6) (D) in comparison to the vehicle control under either non-stimulated or stimulated (IFNγ/TNFα) condition. Fold changes for the top 25 genes either induced (E) or repressed (F) by 1.5 mg/kg prednisolone and 0.3 mg/kg Org 214007-0 in muscle tissue from arthritic mice. NB. Scales are in 2log. Example of an induced gene (Per-2) (G) and a repressed gene (Ccl8) (H) in comparison to vehicle treated arthritic mice and vehicle treated healthy mice.</p

Structure and predicted binding mode of Org 214007-0.

<p>A) The structure of ORG 214007-0. This compound, [(-)-N-(2S,10S,14bS)]-N-(8-cyano-1,2,3,4,10,14b-hexahydro-10-methyl dibenzo<i>[c,f]</i>pyrido[1,2-<i>a</i>]azepin-2-yl)-4-methyl-1,2,3-thiadiazole-5-carboxamide] has a molecular weight of 430 g/mole (C₂₄H₂₃N₅OS) B) The predicted binding mode of Org 214007-0 modeled in complex with the glucocorticoid receptor and demonstrating conservation of interactions typical to steroidal glucocorticoids (Gln564, Asn570, Arg611 and Gln642).</p