Inhibition of RANKL-induced osteoclastogenesis by (-)-DHMEQ, a novel NF-kappaB inhibitor, through downregulation of NFATc1.
(-)-DHMEQ, a newly designed NF-kappaB inhibitor, inhibited RANKL-induced osteoclast differentiation in mouse BMMs through downregulation of the induction of NFATc1, an essential transcription factor of osteoclastogenesis.


BACKGROUND
Bone destruction is often observed in advanced case of rheumatoid arthritis and neoplastic diseases, including multiple myeloma. Effective and nontoxic chemotherapeutic agents are expected for the suppression of these bone destructions. RANKL induces activation of NF-kappaB and osteoclastogenesis in bone marrow-derived monocyte/macrophage precursor cells (BMMs). Targeted disruption or pharmacological suppression of NF-kappaB result in impaired osteoclastogenesis, but how NF-kappaB is involved in the regulation of osteoclastogenesis is not known.


METHODS
The effect of (-)-dehydroxymethylepoxyquinomicin [(-)-DHMEQ] on osteoclast differentiation was studied using a culture system of mouse BMMs stimulated with RANKL and macrophage colony-stimulating factor. The mechanism of the inhibition was studied by biochemical analysis such as immunoblotting and retroviral transfer experiments.


RESULTS
(-)-DHMEQ strongly inhibited RANKL-induced NF-kappaB activation in BMMs and inhibited RANKL-induced formation of TRACP(+) multinucleated cells. Interestingly, (-)-DHMEQ specifically inhibited the RANKL-induced expression of NFATc1 but not the expressions of TRAF6 or c-fos. Inhibition of osteoclast differentiation by (-)-DHMEQ was rescued by overexpression of NFATc1, suggesting that the inhibition is not caused by a toxic effect. Moreover, pit formation assays showed that (-)-DHMEQ also inhibited the bone-resorbing activity of mature osteoclasts.


CONCLUSIONS
The inhibition of NF-kappaB suppresses osteoclastogenesis by downregulation of NFATc1, suggesting that NFATc1 expression is regulated by NF-kappaB in RANKL-induced osteoclastogenesis. Our results also indicate the possibility of (-)-DHMEQ becoming a new therapeutic strategy against bone erosion.
Takatsuna
Hiroshi
H
Department of Applied Chemistry, Faculty of Science and Technology, Keio University, Yokohama, Japan.
Asagiri
Masataka
M
Kubota
Takeshi
T
Oka
Kotaro
K
Osada
Toshihiro
T
Sugiyama
Chie
C
Saito
Hiroaki
H
Aoki
Kazuhiro
K
Ohya
Keiichi
K
Takayanagi
Hiroshi
H
Umezawa
Kazuo
K
D000818:Animals; D001549:Benzamides / Q000737:chemistry / Q000494:pharmacology*; D001854:Bone Marrow Cells / Q000187:drug effects / Q000378:metabolism; D001862:Bone Resorption; D002352:Carrier Proteins / Q000037:antagonists & inhibitors* / Q000494:pharmacology; D002454:Cell Differentiation / Q000187:drug effects; D003512:Cyclohexanones / Q000737:chemistry / Q000494:pharmacology*; D015536:Down-Regulation; D008264:Macrophages / Q000187:drug effects / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000037:antagonists & inhibitors* / Q000494:pharmacology; D051379:Mice; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D015394:Molecular Structure; D016328:NF-kappa B / Q000037:antagonists & inhibitors*; D050778:NFATC Transcription Factors / Q000378:metabolism*; D010010:Osteoclasts / Q000187:drug effects* / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D053245:RANK Ligand; D053246:Receptor Activator of Nuclear Factor-kappa B
D001549:Benzamides; D002352:Carrier Proteins; D003512:Cyclohexanones; D008562:Membrane Glycoproteins; D016328:NF-kappa B; D050778:NFATC Transcription Factors; C494005:Nfatc1 protein, mouse; D053245:RANK Ligand; D053246:Receptor Activator of Nuclear Factor-kappa B; C505687:Tnfrsf11a protein, mouse; C505689:Tnfsf11 protein, mouse; C464444:dehydroxymethylepoxyquinomicin; D020928:Mitogen-Activated Protein Kinases