Antibacterial Activity and Sustained Effectiveness of Calcium Silicate-Based Cement as a Root-End Filling Material against Enterococcus faecalis.

Several calcium silicate cement (CSC) types with improved handling properties have been developed lately for root-end filling applications. While sealing ability is important, a high biocompatibility and antimicrobial effects are critical. This study aimed to conduct a comparative evaluation of the antimicrobial efficacy and sustained antibacterial effectiveness against Enterococcus faecalis (E. faecalis) of commercially available CSCs mixed with distilled water (DW) and chlorhexidine (CHX). Various products, viz., ProRoot mixed with DW (PRW) or with CHX (PRC), Endocem mixed with DW (EW) or with CHX (EC), and Endocem premixed (EP) syringe type, were used. While antibacterial activity against E. faecalis was evaluated using a direct contact method, the specimens were stored in a shaking incubator for 30 d for antibacterial sustainability. The cytotoxicity was evaluated using a cell counting kit-8 assay in human periodontal ligament stem cells. The antibacterial activities of EP, EW, and EC were greater than those of PRC and PRW (p < 0.05). The antibacterial sustainability of EP was the highest without cytotoxicity for up to 30 days (p < 0.05). In conclusion, the pre-mixed injectable type EP was most effective in terms of antibacterial activity and sustained antibacterial effectiveness without cytotoxicity.

Oral Microbiome Using Colocasia antiquorum var. esculenta Extract Varnish in a Mouse Model with Oral Gavage of P. gingivalis ATCC 53978.

Background and Objective: There is increasing interest in preventing periodontitis using natural products. The purpose of this study was to investigate the effect of Colocasia antiquorum var. esculenta (CA) varnish on the oral microbiome and alveolar bone loss in a mouse periodontitis model. Materials and Methods: Antibacterial activity against Porphyromonas gingivalis (P. gingivalis) ATCC 53978 and cell cytotoxicity using CCK-8 on L929 cells were measured. Balb/c mice were assigned into five groups (negative control, positive control, CA in drinking water, varnish, and CA varnish). P. gingivalis was administered to the mice by oral gavage three times. After sacrifice, the oral microbiome and the levels of the inflammatory cytokine IL-1ß and matrix metalloproteinase-9 were analyzed. Alveolar bone loss was measured using micro-computed tomography. Results: CA extract showed an antibacterial effect against P. gingivalis (p < 0.05) and showed no cytotoxicity at that concentration (p > 0.05). Although alpha diversity of the oral microbiome did not statistically differ between the groups (p > 0.05), the relative abundance of dominant bacteria tended to be different between the groups. The inflammatory cytokine IL-1ß was reduced in the CA varnish group (p < 0.05), and no difference was observed in MMP-9 expression and alveolar bone loss (p > 0.05). Conclusions: CA varnish did not affect the overall microflora and exhibited an anti-inflammatory effect, suggesting that it is possibility a suitable candidate for improving periodontitis.

Effects of Colocasia antiquorum var. Esculenta Extract In Vitro and In Vivo against Periodontal Disease.

Background and Objectives: Periodontal disease is a chronic inflammatory disease in which gradual destruction of tissues around teeth is caused by plaque formed by pathogenic bacteria. The purpose of this study was to evaluate the potential of 75% ethanol extract of Colocasia antiquorum var. esculenta (CA) as a prophylactic and improvement agent for periodontal disease in vitro and in vivo. Materials and Methods: The antimicrobial efficacy of CA against Porphyromonas gingivalis (P. gingivalis, ATCC 33277) was evaluated using minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) test, and cytotoxicity was confirmed by CCK-8 assay. For the in vivo study, P. gingivalis was applied by oral gavage to BALB/c mice. Forty-two days after the first inoculation of P. gingivalis, intraoral swabs were taken for microbiome analysis, and the mice were sacrificed to evaluate the alveolar bone loss. Results: The MIC of CA against P. gingivalis was 31.3 µg/mL, the MBC was 62.5 µg/mL, with no cytotoxicity. The diversity of the oral microbiome decreased in the positive control group, while those of the VA (varnish) and VCA (varnish added with CA) groups increased as much as in the negative control group, although the alveolar bone loss was not induced in the mouse model. Conclusions: CA showed antibacterial effects in vitro, and the VA and VCA groups exhibited increased diversity in the oral microbiome, suggesting that CA has potential for improving periodontal disease.

Inhibitory Effect of Asplenium incisum on Bacterial Growth, Inflammation, and Osteoclastogenesis.

Background and Objectives:Asplenium incisum, a natural plant, is known to possess numerous pharmacological and biochemical properties. However, the inhibitory effect of A. incisum against Porphyromonas gingivalis and other factors related to periodontal disease have not yet been demonstrated. This study aimed to investigate the potential of A. incisum extract as a phytotherapeutic candidate for improving periodontal diseases by assessing its antibacterial, anti-inflammatory, and anti-osteoclastogenic activities. Materials and Methods: The inhibition of proliferation of P. gingivalis by A. incisum and the sustainability of its antibacterial activity were evaluated in this study. The production of inflammatory cytokines (tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6)) and nitric oxide (NO) from lipopolysaccharide-stimulated RAW 264.7 cells was assessed using an enzyme-linked immunosorbent assay. To identify the anti-osteoclastogenic activity, tartrate-resistant acid phosphatase (TRAP) staining and TRAP activity analyses were performed on bone marrow macrophages. Results: The proliferation of P. gingivalis was significantly inhibited by A. incisum (p < 0.001), and the antibacterial activity was sustained for up to 3 days. A. incisum showed anti-inflammatory activities by significantly decreasing the release of TNF-α, IL-6 (p < 0.05), and NO (p < 0.01). In addition, A. incisum significantly suppressed TRAP-positive cells and TRAP activity (at 30 µg/mL, p < 0.01) without causing any cytotoxicity (p > 0.05). Conclusions:A. incisum showed antibacterial, anti-inflammatory, and anti-osteoclastogenic activities, suggesting it has strong therapeutic potential against periodontal diseases.

Antibacterial, anti-inflammatory, and anti-osteoclastogenic activities of Colocasia antiquorum var. esculenta: Potential applications in preventing and treating periodontal diseases.

This study aimed to investigate the inhibitory effects of Colocasia antiquorum var. esculenta (CA) on Porphyromonas gingivalis (P. gingivalis) growth, inflammation, and osteoclastogenesis. CA was effective in inhibiting the growth of P. gingivalis when applied together with an experimental fluoride varnish. CA also significantly decreased the release of interleukin-6, tumor necrosis factor-α, and nitric oxide from lipopolysaccharide-induced RAW 264.7 cells. No significant differences in viability were noted between the cells treated with CA and the controls. In addition, CA significantly attenuated osteoclast differentiation on bone marrow macrophages. In conclusion, CA inhibited the growth of P. gingivalis and showed anti-inflammatory and anti-osteoclastogenic effects. Therefore, CA may have the potential to act as a novel natural agent for preventing periodontitis.

Toluene Inhalation Causes Early Anxiety and Delayed Depression with Regulation of Dopamine Turnover, 5-HT1A Receptor, and Adult Neurogenesis in Mice.

Inhaled solvents such as toluene are of particular concern due to their abuse potential that is easily exposed to the environment. The inhalation of toluene causes various behavioral problems, but, the effect of short-term exposure of toluene on changes in emotional behaviors over time after exposure and the accompanying pathological characteristics have not been fully identified. Here, we evaluated the behavioral and neurochemical changes observed over time in mice that inhaled toluene. The mice were exposed to toluene for 30 min at a concentration of either 500 or 2,000 ppm. Toluene did not cause social or motor dysfunction in mice. However, increased anxiety-like behavior was detected in the short-term after exposure, and depression-like behavior appeared as delayed effects. The amount of striatal dopamine metabolites was significantly decreased by toluene, which continued to be seen for up to almost two weeks after inhalation. Additionally, an upregulation of serotonin 1A (5-HT1A) receptor in the hippocampus and the substantia nigra, as well as reduced immunoreactivity of neurogenesis markers in the dentate gyrus, was observed in the mice after two weeks. These results suggest that toluene inhalation, even single exposure, mimics early anxietyand delayed depression-like emotional disturbances, underpinned by pathological changes in the brain.

Mollugin enhances the osteogenic action of BMP-2 via the p38-Smad signaling pathway.

Bone morphogenetic protein 2 (BMP-2) has been used clinically to encourage bone regeneration; although, there can be major side effects with larger doses. Therefore, there is a need to identify new small molecules to potentiate the osteogenic action of BMP-2. In this study, we investigated the effect of mollugin on bone formation in murine bi-potential mesenchymal progenitor C2C12 cells by combination with BMP-2. We found mollugin could enhance the BMP-2-mediated osteoblast differentiation of C2C12 cells. This was accompanied by the induction of other osteogenic BMPs. We also found the enhancing potential of mollugin may involve activation of the p38-Smad1/5/8 signaling axis. Furthermore, mollugin promoted skeletal development in zebrafish. The combination of BMP-2 with small molecules, including mollugin, could minimize its clinical limitations, and these molecules might lead to the development of effective stem cell stimulants for bone regeneration and fracture healing.

Synthesis, characterization and biological evaluation of anti-cancer indolizine derivatives via inhibiting ß-catenin activity and activating p53.

Diversity-oriented construction of new indolizine scaffolds was accomplished by utilizing domino Knoevenagel condensation/intramolecular aldol cyclization. Biological evaluation revealed anticancer activity of these compounds through inhibition of ß-catenin and activation of p53.

Anti-cancer activity of ZnO chips by sustained zinc ion release.

We report anti-cancer activity of ZnO thin-film-coated chips by sustained release of zinc ions. ZnO chips were fabricated by precisely tuning ZnO thickness using atomic layer deposition, and their potential to release zinc ions relative to the number of deposition cycles was evaluated. ZnO chips exhibited selective cytotoxicity in human B lymphocyte Raji cells while having no effect on human peripheral blood mononuclear cells. Of importance, the half-maximal inhibitory concentration of the ZnO chip on the viability of Raji cells was 121.5 cycles, which was comparable to 65.7 nM of daunorubicin, an anti-cancer drug for leukemia. Molecular analysis of cells treated with ZnO chips revealed that zinc ions released from the chips increased cellular levels of reactive oxygen species, including hydrogen peroxide, which led to the down-regulation of anti-apoptotic molecules (such as HIF-1α, survivin, cIAP-2, claspin, p-53, and XIAP) and caspase-dependent apoptosis. Because the anti-cancer activity of ZnO chips and the mode of action were comparable to those of daunorubicin, the development and optimization of ZnO chips that gradually release zinc ions might have clinical anti-cancer potential. A further understanding of the biological action of ZnO-related products is crucial for designing safe biomaterials with applications in disease treatment.

Inflammatory Bone Resorption and Antiosteosarcoma Potentials of Zinc Ion Sustained Release ZnO Chips: Friend or Foe?

Multifunctional zinc oxide (ZnO) has been generated as nanoparticles or nanorods and applied to various medical purposes since it exhibits several biological actions including anticancer activity. Especially, due to antibacterial activity and effects on bone regeneration, ZnO is widely used in implants and scaffolds in the orthopedic and dental fields. However, concerns over side effects have been raised recently in the clinical use of ZnO, and it is necessary to assess the safety of ZnO regarding its inflammatory potential in the bone environment. This made us hypothesize that the inflammatory activity of zinc ions released from ZnO NPs could be harmful to induce bone resorption but that their cytotoxicity would be beneficial to kill osteosarcoma. To clarify this hypothesis, in the present work, the effects of ZnO on bone matrix and abnormal bone environments were investigated quantitatively using ZnO chips, filter paper, or glass slides coated with thin films of ZnO grown via atomic layer deposition (ALD). ALD-grown ZnO thin films exhibit thickness with atomic precision, which enables the quantitative analysis of the effects of ZnO. In vivo application of ZnO chips to mouse calvarial bone induced bone resorption, presumably due to the activation of osteoclasts by zinc ion-induced TNF-α release. However, in vitro application of ZnO chips to osteosarcoma cells induced caspase-dependent apoptosis and oxidative stress. Taken together, the results showed two sides of ZnO as our hypothesis. Therefore, careful design and multiple evaluations for the safety and efficacy of ZnO materials are necessary for its successful clinical application.

In vitro anti-osteoclastogenic activity of p38 inhibitor doramapimod via inhibiting migration of pre-osteoclasts and NFATc1 activity.

The mitogen activated protein kinase p38 plays a role in the receptor activator of NF-ĸB ligand (RANKL)-induced osteoclast differentiation. In this study, we investigated the effect of p38 inhibitor doramapimod on the osteoclast differentiation. Doramapimod significantly inhibited the osteoclastogenesis of bone marrow macrophages (BMMs) via attenuating the activation of p38 induced by M-CSF and RANKL. Importantly, doramapimod blocked the migration and fusion in pre-osteoclasts via the down-regulating NFATc1. The inhibitory effect of doramapimod on the migration/fusion of pre-osteoclasts via inhibiting NFATc1 activity were confirmed by measuring NFATc1 luciferase activity and evaluating the mRNA expression of NFATc1-responsive genes related to the osteoclastic migration/fusion. These results suggested anti-osteoclastogenic activity of doramapimod via inhibiting migration/fusion of pre-osteoclasts and NFATc1 activity.

Micropillar arrays as potential drug screens: Inhibition of micropillar-mediated activation of the FAK-Src-paxillin signaling pathway by the CK2 inhibitor CX-4945.

Here, we demonstrate the possible applications of micropillar arrays in screening anti-metastasis drugs. Human lung adenocarcinoma A549 cells incubated in multiwell plates containing micropillars exhibited markedly different physical/biochemical behavior depending on pillar dimensions. In particular, A549 cells grown in plates containing 2-µm diameter, 16-µm pitched pillar arrays showed epithelial-to-mesenchymal transition (EMT)-like behavior; cell body elongation, and highly increased activation of the focal adhesion kinase (FAK)-Src-paxillin signaling cascade. FAK is the most prominent kinase involved in dynamic regulation of the actin cytoskeleton and cell adhesion, migration, and invasion. Activation of FAK, a hallmark of cancer cell adhesion and migration, is normally induced by various growth factors, such as transforming growth factor-ß (TGF-ß). Here, we found that pillar-mediated activation of signaling molecules mimicked that induced by TGF-ß. Notably, micropillar arrays with specific dimensions accelerated the elongation of cells, an effect linked to the activation of signaling molecules related to EMT. Micropillar-induced FAK activation could be arrested by the casein kinase-2 (CK2) inhibitor CX-4945, a drug candidate with activity against TGF-ß-induced cancer cell metastasis, demonstrating the possibility of using inorganic microstructures for cell-based drug screening. STATEMENT OF SIGNIFICANCE: In this work, we have fabricated flexible substrates with regular arrays of micrometersized pillars, and used them to grow A549 human lung adenocarcinoma cells. Cells exhibit dramatically different behavior depending on the intervals of pillars. Especially, cells grown in certain pillar structures show epithelial-to mesenchmal transition (EMT)-like morphology and related molecules, which is similar to the activation obtained using expensive cytokine TGF-ß. Based on the fact that pillar arrays may activate EMT like transition, screening of anti-cancer drug using pillar arrays have demonstrated as well in our work. Our study confirms that mechanical stimulation may exert similar effects with chemical stimulation, and such mechanical structures could be used as a large-scale drug screening platforms. Cell morphogenesis on engineered substrate is not new, but the present work could be distinguished with its unique fabrication process that can mass produce the structures and it could be applied for high-throughput drug screening. Also, we suggest the formation of focal adhesions on pillar structures and consequent strain as the possible mechanism behind the observed EMT-like transition. Currently, we are working on full-scale profiling of metabolomics and proteomics of cells grown in large-scale pillar arrays as well.

Decursin inhibits osteoclastogenesis by downregulating NFATc1 and blocking fusion of pre-osteoclasts.

Bone sustains its structure through dynamic interaction between osteoblastic cells and osteoclastic cells. But imbalance may lead to osteoporosis caused by overactivated osteoclast cells that have bone-resorbing function. Recently, herbs have been researched as major sources of medicines in many countries. In vitro and in vivo anti-osteoclastogenic activity of Angelica gigas NAKAI have been reported, but the biological activity of decursin, its major component in osteoclast differentiation is still unknown. Therefore, in this study, we explored whether decursin could affect RANKL-mediated osteoclastogenesis. The results showed that decursin efficiently inhibited RANKL-activated osteoclast differentiation by inhibiting transcriptional and translational expression of NFATc1, a major factor in RANKL-mediated osteoclastogenesis. Furthermore, decursin decreased fusion and migration of pre-osteoclasts by downregulating mRNA expression levels of DC-STAMP and ß3 integrin, respectively. In addition, decursin prevents lipopolysaccharide (LPS)-induced bone erosion in vivo. In summary, decursin could prevent osteoclastogenesis and inflammatory bone loss via blockage of NFATc1 activity and fusion and migration of pre-osteoclasts, and it could be developed as a potent phytochemical candidate for treating pathologies of bone diseases.

In vitro and in vivo Bone-Forming Activity of Saururus chinensis Extract.

Bone is maintained by osteoclast-mediated resorption and osteoblast-mediated formation. Recently, anti-osteoporotic activity of Saururus chinensis extract (SCE) and anti-osteoclastogenic activity of its components have been reported, but the effect of SCE on bone formation has not been studied well. Therefore, in this study, we investigated whether Saururus chinensis SCE exhibits in vitro osteogenic and in vivo bone-forming activity. extract strongly enhanced the bone morphogenetic protein (BMP)-2-stimulated induction of alkaline phosphatase, an early phase biomarker of osteoblast differentiation, in bi-potential mesenchymal progenitor C2C12 cells. In vitro osteogenic activity of SCE was accompanied by enhanced expression of BMP-2, BMP-4, BMP-7 and BMP-9 mRNA. In addition, a pharmacological inhibition study suggested the involvement of p38 activation in the osteogenic action of SCE. Moreover, the BMP dependency and the involvement of p38 activation in the osteogenic action of SCE were confirmed by the treatment of noggin, an antagonist of BMP. Saururus chinensis extract also exhibited to induce runt-related transcription factor 2 activation at the high concentration. Furthermore, the in vivo osteogenic activity of SCE was confirmed in zebrafish and mouse calvarial bone formation models, suggesting the possibility of its use for bone formation. In conclusion, we suggested that in vivo anti-osteoporotic activity of SCE could be because of its dual action in bone, anti-osteoclastogenic and anabolic activity.

Mollugin from Rubea cordifolia suppresses receptor activator of nuclear factor-κB ligand-induced osteoclastogenesis and bone resorbing activity in vitro and prevents lipopolysaccharide-induced bone loss in vivo.

Osteopenic diseases, such as osteoporosis, are characterized by progressive and excessive bone resorption mediated by enhanced receptor activator of nuclear factor-κB ligand (RANKL) signaling. Therefore, downregulation of RANKL downstream signals may be a valuable approach for the treatment of bone loss-associated disorders. In this study, we investigated the effects of the naphthohydroquinone mollugin on osteoclastogenesis and its function in vitro and in vivo. Mollugin efficiently suppressed RANKL-induced osteoclast differentiation of bone marrow macrophages (BMMs) and bone resorbing activity of mature osteoclasts by inhibiting RANKL-induced c-Fos and NFATc1 expression. Mollugin reduced the phosphorylation of signaling pathways activated in the early stages of osteoclast differentiation, including the MAP kinase, Akt, and GSK3ß and inhibited the expression of different genes associated with osteoclastogenesis, such as OSCAR, TRAP, DC-STAMP, OC-STAMP, integrin αν, integrin ß3, cathepsin K, and ICAM-1. Furthermore, mice treated with mollugin showed significant restoration of lipopolysaccharide (LPS)-induced bone loss as indicated by micro-CT and histological analysis of femurs. Consequently, these results suggested that mollugin could be a novel therapeutic candidate for bone loss-associated disorders including osteoporosis, rheumatoid arthritis, and periodontitis.

Repositioning Potential of PAK4 to Osteoclastic Bone Resorption.

Drug repositioning is a rational approach for expanding the use of existing drugs or candidate drugs to treat additional disorders. Here we investigated the possibility of using the anticancer p21-activated kinase 4 (PAK4)-targeted inhibitor PF-3758309 to treat osteoclast-mediated disorders. PAK4 was highly expressed in bone marrow cells and was phosphorylated during their differentiation into osteoclasts, and osteoclast differentiation was significantly inhibited by the dominant negative form of PAK4 and by PF-3758309. Specifically, PF-3758309 significantly inhibited the fusion of preosteoclasts, the podosome formation, and the migration of preosteoclasts. PF-3758309 also had in vivo antiresorptive activity in a lipopolysaccharide-induced bone erosion model and in vitro antiosteoclastogenic activity in the differentiation of human bone marrow-derived cells and peripheral blood mononuclear cells into osteoclasts. These data demonstrate the relevance of PAK4 in osteoclast differentiation and the potential of PAK4 inhibitors for treating osteoclast-related disorders.

A novel aminothiazole KY-05009 with potential to inhibit Traf2- and Nck-interacting kinase (TNIK) attenuates TGF-ß1-mediated epithelial-to-mesenchymal transition in human lung adenocarcinoma A549 cells.

Transforming growth factor (TGF)-ß triggers the epithelial-to-mesenchymal transition (EMT) of cancer cells via well-orchestrated crosstalk between Smad and non-Smad signaling pathways, including Wnt/ß-catenin. Since EMT-induced motility and invasion play a critical role in cancer metastasis, EMT-related molecules are emerging as novel targets of anti-cancer therapies. Traf2- and Nck-interacting kinase (TNIK) has recently been considered as a first-in-class anti-cancer target molecule to regulate Wnt signaling pathway, but pharmacologic inhibition of its EMT activity has not yet been studied. Here, using 5-(4-methylbenzamido)-2-(phenylamino)thiazole-4-carboxamide (KY-05009) with TNIK-inhibitory activity, its efficacy to inhibit EMT in cancer cells was validated. The molecular docking/binding study revealed the binding of KY-05009 in the hinge region of TNIK, and the inhibitory activity of KY-05009 against TNIK was confirmed by an ATP competition assay (Ki, 100 nM). In A549 cells, KY-05009 significantly and strongly inhibited the TGF-ß-activated EMT through the attenuation of Smad and non-Smad signaling pathways, including the Wnt, NF-κB, FAK-Src-paxillin-related focal adhesion, and MAP kinases (ERK and JNK) signaling pathways. Continuing efforts to identify and validate potential therapeutic targets associated with EMT, such as TNIK, provide new and improved therapies for treating and/or preventing EMT-based disorders, such as cancer metastasis and fibrosis.

Anti-osteoclastogenic activity of praeruptorin A via inhibition of p38/Akt-c-Fos-NFATc1 signaling and PLCγ-independent Ca2+ oscillation.

BACKGROUND: A decrease of bone mass is a major risk factor for fracture. Several natural products have traditionally been used as herbal medicines to prevent and/or treat bone disorders including osteoporosis. Praeruptorin A is isolated from the dry root extract of Peucedanum praeruptorum Dunn and has several biological activities, but its anti-osteoporotic activity has not been studied yet. MATERIALS AND METHODS: The effect of praeruptorin A on the differentiation of bone marrow-derived macrophages into osteoclasts was examined by phenotype assay and confirmed by real-time PCR and immunoblotting. The involvement of NFATc1 in the anti-osteoclastogenic action of praeruptorin A was evaluated by its lentiviral ectopic expression. Intracellular Ca(2+) levels were also measured. RESULTS: Praeruptorin A inhibited the RANKL-stimulated osteoclast differentiation accompanied by inhibition of p38 and Akt signaling, which could be the reason for praeruptorin A-downregulated expression levels of c-Fos and NFATc1, transcription factors that regulate osteoclast-specific genes, as well as osteoclast fusion-related molecules. The anti-osteoclastogenic effect of praeruptorin A was rescued by overexpression of NFATc1. Praeruptorin A strongly prevented the RANKL-induced Ca(2+) oscillation without any changes in the phosphorylation of PLCγ. CONCLUSION: Praeruptorin A could exhibit its anti-osteoclastogenic activity by inhibiting p38/Akt-c-Fos-NFATc1 signaling and PLCγ-independent Ca(2+) oscillation.

The protein kinase 2 inhibitor CX-4945 regulates osteoclast and osteoblast differentiation in vitro.

Drug repositioning can identify new therapeutic applications for existing drugs, thus mitigating high R&D costs. The Protein kinase 2 (CK2) inhibitor CX-4945 regulates human cancer cell survival and angiogenesis. Here we found that CX-4945 significantly inhibited the RANKL-induced osteoclast differentiation, but enhanced the BMP2-induced osteoblast differentiation in a cell culture model. CX-4945 inhibited the RANKL-induced activation of TRAP and NFATc1 expression accompanied with suppression of Akt phosphorylation, but in contrast, it enhanced the BMP2-mediated ALP induction and MAPK ERK1/2 phosphorylation. CX-4945 is thus a novel drug candidate for bone-related disorders such as osteoporosis.

Osteoclast differentiation inhibitors: a patent review (2008 - 2012).

INTRODUCTION: Mononuclear macrophage/monocyte-lineage hematopoietic precursors differentiate into multinucleated osteoclasts. Abnormally increased numbers and/or overactivation of osteoclasts can lead to bone loss. Therefore, pharmaceutical inhibition of osteoclast differentiation is one therapeutic strategy for mitigating the occurrence of bone loss-associated disorders and related fractures. AREAS COVERED: This review surveys the patents and patent applications from 2008 to 2012 that are related to inventions of therapeutics and/or methods for inhibiting osteoclast differentiation. EXPERT OPINION: Over the past 20 years, the identification and validation of signaling molecules involved in osteoclast differentiation has led to a better understanding of the molecular mechanism, and to the development of new therapeutic agents for treating bone loss-associated disorders. Since 2008, 34 WO patents or patent applications have been filed that relate to inventions of therapeutics and/or methods for chemical-based, natural product-based, or biological-based inhibitors of osteoclast differentiation. Here, analysis of these patents and patent applications is presented, and summarize the disclosed osteoclast differentiation-inhibiting target molecules. This report can support further advances in the development of anti-osteoclastogenic therapeutics for bone loss-associated disorders, including osteoporosis, rheumatoid arthritis, Paget's disease, periodontal disease, osteosarcoma, and cancer bone metastasis.