An anti-inflammatory selective glucocorticoid receptor modulator preserves osteoblast differentiation.

Glucocorticoids (GCs) are in widespread use to treat inflammatory bone diseases, such as rheumatoid arthritis (RA). Their anti-inflammatory efficacy, however, is accompanied by deleterious effects on bone, leading to GC-induced osteoporosis (GIO). These effects include up-regulation of the receptor activator of NF-κB ligand/osteoprotegerin (RANKL/OPG) ratio to promote bone-resorbing osteoclasts and include inhibition of bone-forming osteoblasts. We previously identified suppression of osteoblast differentiation by the monomer glucocorticoid receptor (GR) via the inhibition of Il11 expression as a crucial mechanism for GIO. Here we show that the GR-modulating substance compound A (CpdA), which does not induce GR dimerization, still suppresses proinflammatory cytokines in fibroblast-like synovial cells from patients with RA and in osteoblasts. In contrast to the full GR agonist dexamethasone, it does not unfavorably alter the RANKL/OPG ratio and does not affect Il11 expression and subsequent STAT3 phosphorylation in these cells. Notably, while dexamethasone inhibits osteoblast differentiation, CpdA does not affect osteoblast differentiation in vitro and in vivo. We describe here for the first time that selective GR modulators can act against inflammation, while not impairing osteoblast differentiation.

Therapeutic implications of the nuclear factor-kappaB/nuclear receptor cross-talk.

More and more evidence reveals that the transcription factor NF-kappaB plays a critical role in tumor development and progression and that it may constitute the missing link between inflammation and cancer. It turned out that many of the well known cancer drugs exert their anti-tumoral effect at least in part through modulating the activity of NF-kappaB. The potential of nuclear receptors to modulate the activity of this widespread transcription factor has repeatedly been reported and illustrates their enormous therapeutic potential. However, the efficacy of these liganded receptors is overshadowed by the occurrence of unwanted effects owing to their broad range of actions. Accordingly, researchers pursue the ambition to improve the specificity of nuclear receptor modulators. In this review we have explored the molecular mechanisms by which nuclear receptors interfere with NF-kappaB signalling and quoted the therapeutic implications of their cross-coupling. Strategies that are explored at the moment and that may hold great potential for the future are extensively reviewed.

Compound A, a selective glucocorticoid receptor modulator, enhances heat shock protein Hsp70 gene promoter activation.

Compound A possesses glucocorticoid receptor (GR)-dependent anti-inflammatory properties. Just like classical GR ligands, Compound A can repress NF-κB-mediated gene expression. However, the monomeric Compound A-activated GR is unable to trigger glucocorticoid response element-regulated gene expression. The heat shock response potently activates heat shock factor 1 (HSF1), upregulates Hsp70, a known GR chaperone, and also modulates various aspects of inflammation. We found that the selective GR modulator Compound A and heat shock trigger similar cellular effects in A549 lung epithelial cells. With regard to their anti-inflammatory mechanism, heat shock and Compound A are both able to reduce TNF-stimulated IκBα degradation and NF-κB p65 nuclear translocation. We established an interaction between Compound A-activated GR and Hsp70, but remarkably, although the presence of the Hsp70 chaperone as such appears pivotal for the Compound A-mediated inflammatory gene repression, subsequent novel Hsp70 protein synthesis is uncoupled from an observed CpdA-induced Hsp70 mRNA upregulation and hence obsolete in mediating CpdA's anti-inflammatory effect. The lack of a Compound A-induced increase in Hsp70 protein levels in A549 cells is not mediated by a rapid proteasomal degradation of Hsp70 or by a Compound A-induced general block on translation. Similar to heat shock, Compound A can upregulate transcription of Hsp70 genes in various cell lines and BALB/c mice. Interestingly, whereas Compound A-dependent Hsp70 promoter activation is GR-dependent but HSF1-independent, heat shock-induced Hsp70 expression alternatively occurs in a GR-independent and HSF1-dependent manner in A549 lung epithelial cells.

Differential mechanism of NF-kappaB inhibition by two glucocorticoid receptor modulators in rheumatoid arthritis synovial fibroblasts.

OBJECTIVE: To investigate and compare the molecular mechanisms by which 2 glucocorticoid receptor (GR)-activating compounds, dexamethasone (DEX) and Compound A (CpdA), interfere with the NF-kappaB activation pathway in rheumatoid arthritis (RA) synovial cells. METHODS: Quantitative polymerase chain reaction was performed to detect the tumor necrosis factor alpha (TNFalpha)-induced cytokine gene expression of interleukin-1beta (IL-1beta) and to investigate the effects of DEX and CpdA in RA fibroblast-like synoviocytes (FLS) transfected with small interfering RNA (siRNA) against GR (siGR) compared with nontransfected cells. Immunofluorescence analysis was used to detect the subcellular distribution of NF-kappaB (p65) under the various treatment conditions, and active DNA-bound p65 was measured using a TransAM assay and by chromatin immunoprecipitation analysis of IL-1beta. Signaling pathways were studied via Western blotting of siGR-transfected cells, compared with nontransfected and nontargeting siRNA-transfected control cells, to detect the regulation of phospho-IKK, IkappaBalpha, phospho-p38, phospho-ERK, and phospho-JNK. RESULTS: Both DEX and CpdA efficiently inhibited IL-1beta gene expression in a GR-dependent manner. In addition, CpdA attenuated the TNFalpha-induced nuclear translocation and DNA binding of p65 in RA FLS, via the attenuation of IKK phosphorylation and subsequent IkappaBalpha degradation. CpdA also displayed profound effects on TNFalpha-induced MAPK activation. The effects of CpdA on TNFalpha-induced kinase activities occurred independently of the presence of GR. In sharp contrast, DEX did not affect TNFalpha-induced IKK phosphorylation, IkappaBalpha degradation, p65 nuclear translocation, or MAPK activation in RA FLS. CONCLUSION: DEX and CpdA display a dissimilar molecular mechanism of interaction with the NF-kappaB activation pathway ex vivo. A dual pathway, partially dependent and partially independent of GR (nongenomic), may explain the gene-inhibitory effects of CpdA in RA FLS.

A plant-derived ligand favoring monomeric glucocorticoid receptor conformation with impaired transactivation potential attenuates collagen-induced arthritis.

The glucocorticoid receptor (GR) is a transcription factor regulating its target genes either positively, through direct binding to the promoter of target genes, or negatively by the interference with the activity of transcription factors involved in proinflammatory gene expression. The well-known adverse effects of glucocorticoids are believed to be mainly caused by their GR-mediated gene-activating properties. Although dimerization of GR is thought to be essential for gene-activating properties, no compound has yet been described which selectively imposes GR monomer formation and interference with other transcription factors. In the present study, we report on a GR-binding, plant-derived compound with marked dissociative properties in rheumatoid arthritis fibroblast-like synoviocytes, which are important effector cells in inflammation and matrix degradation in rheumatoid arthritis. In addition, these findings could be extended in vivo in murine collagen-induced arthritis, in which joint inflammation was markedly inhibited without inducing hyperinsulinemia. Therefore, we conclude that GR monomers are sufficient for inhibition of inflammation in vivo.