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IKKβ has previously been implicated in breast cancer bone metastasis and bone remodelling. However, the contribution of IKKβ expressed by bone cells of the tumour microenvironment to breast cancer-induced osteolysis has yet to be investigated. Here, we studied the effects of the verified selective IKKβ inhibitors IKKβIII or IKKβV on osteoclast formation and osteoblast differentiation in vitro and in vivo, human and mouse breast cancer cells' support for osteoclast formation and signalling in vitro and osteolysis ex vivo and in immunocompetent mice after supracalvarial injection of human MDA-MB-231 conditioned medium or intra-cardiac injection of syngeneic 4T1 breast cancer cells. Pre-treatment with IKKβIII or IKKβV prior to exposure to tumour-derived factors from human and mouse breast cancer cell lines protected against breast cancer-induced osteolysis in two independent immunocompetent mouse models of osteolysis and the ex vivo calvarial bone organ system. Detailed functional and mechanistic studies showed that direct inhibition of IKKβ kinase activity in osteoblasts and osteoclasts was associated with significant reduction of osteoclast formation, enhanced osteoclast apoptosis and reduced the ability of osteoblasts to support osteoclastogenesis in vitro. When combined with previous findings that suggest NFκB inhibition reduces breast cancer tumorigenesis and metastasis our present findings have an important clinical implication on raising the possibility that IKKβ inhibitors, as bone anabolics, osteoclast inhibitors as well as anti-metastatic agents, may have advantages over anti-osteoclasts agents in the treatment of both skeletal and non-skeletal complications associated with metastatic breast cancer. [ABSTRACT FROM AUTHOR]

Tumour necrosis factor receptor-associated factor 6 (TRAF6) has been implicated in breast cancer and osteoclastic bone destruction. Here, we report that 6877002, a verified small-molecule inhibitor of TRAF6, reduced metastasis, osteolysis and osteoclastogenesis in models of osteotropic human and mouse breast cancer. First, we observed that TRAF6 is highly expressed in osteotropic breast cancer cells and its level of expression was higher in patients with bone metastasis. Pre-exposure of osteoclasts and osteoblasts to non-cytotoxic concentrations of 6877002 inhibited cytokine-induced NFκB activation and osteoclastogenesis, and reduced the ability of osteotropic human MDA-MB-231 and mouse 4T1 breast cancer cells to support bone cell activity. 6877002 inhibited human MDA-MB-231-induced osteolysis in the mouse calvaria organ system, and reduced soft tissue and bone metastases in immuno-competent mice following intra-cardiac injection of mouse 4T1-Luc2 cells. Of clinical relevance, combined administration of 6877002 with Docetaxel reduced metastasis and inhibited osteolytic bone damage in mice bearing 4T1-Luc2 cells. Thus, TRAF6 inhibitors such as 6877002 - alone or in combination with conventional chemotherapy - show promise for the treatment of metastatic breast cancer. [ABSTRACT FROM AUTHOR]

Activating receptor activator of NF-kappaB (RANK) and TNF receptor (TNFR) promote osteoclast differentiation. A critical ligand contact site on the TNFR is partly conserved in RANK. Surface plasmon resonance studies showed that a peptide (WP9QY) that mimics this TNFR contact site and inhibits TNF-alpha-induced activity bound to RANK ligand (RANKL). Changing a single residue predicted to play an important role in the interaction reduced the binding significantly. WP9QY, but not the altered control peptide, inhibited the RANKL-induced activation of RANK-dependent signaling in RAW 264.7 cells but had no effect on M-CSF-induced activation of some of the same signaling events. WP9QY but not the control peptide also prevented RANKL-induced bone resorption and osteoclastogenesis, even when TNFRs were absent or blocked. In vivo, where both RANKL and TNF-alpha promote osteoclastogenesis, osteoclast activity, and bone loss, WP9QY prevented the increased osteoclastogenesis and bone loss induced in mice by ovariectomy or low dietary calcium, in the latter case in both wild-type and TNFR double-knockout mice. These results suggest that a peptide that mimics a TNFR ligand contact site blocks bone resorption by interfering with recruitment and activation of osteoclasts by both RANKL and TNF. [ABSTRACT FROM AUTHOR]

The NFκB signaling pathway is implicated in breast cancer and bone metastasis. However, the bone-autonomous contribution of NFκB to breast cancer-induced osteolysis is poorly understood. Here, we report that pretreatment of osteoblasts with the sesquiterpene lactone Parthenolide (PTN), a verified NFκB inhibitor, prior to exposure to conditioned medium from human and mouse breast cancer cell lines enhanced osteoblast differentiation and reduced osteoblast ability to stimulate osteoclastogenesis. PTN prevented breast cancer-induced osteoclast formation and reduced the ability of breast cancer cells to prolong osteoclast survival and to inhibit osteoclast apoptosis. In vivo, administration of PTN in immuno-competent mice reduced osteolytic bone loss and skeletal tumour growth following injection of the syngeneic 4T1-BT1 cells and reduced local osteolysis caused by conditioned medium from human and mouse osteotropic breast cancer cell lines. Mechanistic studies revealed that NFκB inhibition by PTN in osteoblasts and osteoclasts was accompanied by a significant increase in β-catenin activation and expression. Collectively, these results raise the possibility that combined targeting of NFκB and β-catenin signalling in the tumour microenvironment may be of value in the treatment of breast cancer related osteolysis. [ABSTRACT FROM AUTHOR]