RANKL acts directly on RANK-expressing prostate tumor cells and mediates migration and expression of tumor metastasis genes

Department of Hematology, Amgen Inc., Seattle, Washington 98119, USA.
The Prostate (Impact Factor: 3.57). 01/2008; 68(1):92-104. DOI: 10.1002/pros.20678
Source: PubMed


Metastases to bone are a frequent complication of human prostate cancer and result in the development of osteoblastic lesions that include an underlying osteoclastic component. Previous studies in rodent models of breast and prostate cancer have established that receptor activator of NF-kappaB ligand (RANKL) inhibition decreases bone lesion development and tumor growth in bone. RANK is essential for osteoclast differentiation, activation, and survival via its expression on osteoclasts and their precursors. RANK expression has also been observed in some tumor cell types such as breast and colon, suggesting that RANKL may play a direct role on tumor cells.
Male CB17 severe combined immunodeficient (SCID) mice were injected with PC3 cells intratibially and treated with either PBS or human osteprotegerin (OPG)-Fc, a RANKL antagonist. The formation of osteolytic lesions was analyzed by X-ray, and local and systemic levels of RANKL and OPG were analyzed. RANK mRNA and protein expression were assessed on multiple prostate cancer cell lines, and events downstream of RANK activation were studied in PC3 cells in vitro.
OPG-Fc treatment of PC3 tumor-bearing mice decreased lesion formation and tumor burden. Systemic and local levels of RANKL expression were increased in PC3 tumor bearing mice. PC3 cells responded to RANKL by activating multiple signaling pathways which resulted in significant changes in expression of genes involved in osteolysis and migration. RANK activation via RANKL resulted in increased invasion of PC3 cells through a collagen matrix.
These data demonstrate that host stromal RANKL is induced systemically and locally as a result of PC3 prostate tumor growth within the skeleton. RANK is expressed on prostate cancer cells and promotes invasion in a RANKL-dependent manner.

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Available from: Allison Jacob, Jun 17, 2014
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    • "(B) The same cells used for the above tumor xenografts were grown in vitro and processed for flow cytometry. For anti-RANK staining, the pattern of staining by both test antibodies N-1H8 and N-2B10 was compared with a previously identified mAb (M331) useful for flow cytometry applications (Armstrong [31]). Solid grey line: Unstained; Red line: Secondary control¼Goat antimouse APC; Blue line: Isotype control 4D2 (anti-AGP3 muIgG1), 1 μg/mL; Purple line: M331 (anti-huRANK muIgG1), 1 μg/mL; Green line: N-1H8 (anti-huRANK muIgG1), 1 μg/mL; Black line: N-2B10 (anti-huRANK muIgG1), 1 μg/mL. "
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    ABSTRACT: Receptor activator of nuclear factor kappa-B ligand (RANKL) is an essential mediator of osteoclast formation, function and survival. In patients with solid tumor metastasis to the bone, targeting the bone microenvironment by inhibition of RANKL using denosumab, a fully human monoclonal antibody (mAb) specific to RANKL, has been demonstrated to prevent tumor-induced osteolysis and subsequent skeletal complications. Recently, a prominent functional role for the RANKL pathway has emerged in the primary bone tumor giant cell tumor of bone (GCTB). Expression of both RANKL and RANK is extremely high in GCTB tumors and denosumab treatment was associated with tumor regression and reduced tumor-associated bone lysis in GCTB patients. In order to address the potential role of the RANKL pathway in another primary bone tumor, this study assessed human RANKL and RANK expression in human primary osteosarcoma (OS) using specific mAbs, validated and optimized for immunohistochemistry (IHC) or flow cytometry.
    Full-text · Article · Jul 2015 · Journal of Bone Oncology
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    • "These data implicate the function of RANK/ERK-mediated signaling in the timing of normal osteoblasts chemotactic migration during bone remodeling which is no longer necessary during osteosarcoma tumorigenesis, either by loss of RANK function or loss of signaling events. RANK signaling has been implicated in the migration of human prostate cancer cells (Jones et al., 2006), breast cancer cells and melanoma cells (Armstrong et al., 2008; Canon et al., 2008). In osteosarcoma, two groups found contradictory results (Mori et al., 2007; Akiyama et al., 2010). "
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    ABSTRACT: Bone remodeling requires osteoclast activation, resorption and reversal, prior to osteoblast migration into the bone pit. The Receptor Activator of NF-κB (RANK) signaling pathway plays an important role in bone remodeling. Two components of the RANK signaling pathway, RANK Ligand (RANKL) and the decoy receptor Osteoprotegerin (OPG), are expressed predominantly on the surface of osteoblasts, while RANK is principally expressed on the surface of osteoclasts. However, RANK has also been reported to be expressed on the surface of osteoblasts and osteosarcoma tumor cells. Treatment with soluble RANKL (sRANKL) of both normal osteoblasts and osteosarcoma tumor cells activated phosphorylation of ERK, p38(MAPK) , Akt and p65(NF-κB) . However, modified Boyden chamber assays and wound repair assays showed differential response to sRANKL-induced chemotactic migration in normal osteoblasts and osteosarcoma tumor cells. In contrast to previously published results, both normal osteoblasts and osteosarcoma tumor cells responded to sRANKL-induced chemotactic migration but the normal osteoblasts did so only in the presence of an ERK pathway inhibitor. For both normal and tumor cells, the chemotactic response could be blocked by inhibiting the PI3K/Akt or p65(NF-κB) pathway. Response to sRANKL in normal and tumor cells suggests a role for RANK/ERK-mediated signaling in normal osteoblasts chemotactic migration during bone remodeling that is altered or lost during osteosarcoma tumorigenesis. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.
    Full-text · Article · Apr 2015 · Journal of Cellular Physiology
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    • "RANK also played a key role in tumor cell migration and invasion [20], [21]. Moreover, it has been demonstrated that RANKL promotes migration, and invasion of several types of human tumor cells expressing its receptor RANK [22], [23], [24]. However, the role of RANKL-RANK axis in modulating the behaviors of HCC cells is mostly unknown. "
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    ABSTRACT: Background Metastasis accounts for the most deaths in patients with hepatocellular carcinoma (HCC). Receptor activator of nuclear factor kappa B ligand (RANKL) is associated with cancer metastasis, while its role in HCC remains largely unknown. Methods Immunohistochemistry was performed to determine the expression of RANK in HCC tissue (n = 398). Quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot were used to examine the expression of RANK, E-cadherin, N-cadherin, vimentin, Snail, Slug, Twist and MMPs in HCC cells. Wound healing and Transwell assays were used to evaluate cell migration and invasion ability. Results We found that expression of RANK, the receptor of RANKL, was significantly higher in HCC tumor tissues than in peritumor liver tissues (p<0.001). Constitutive expression of RANK was detected in HCC cell lines, which can be up-regulated when HCC cells were stimulated with RANKL. Notably, in vitro experiments showed that activation of RANKL-RANK axis significantly promoted migration and invasion ability of HCC cells. In addition, RANKL stimulation increased the expression levels of N-cadherin, Snail, and Twist, while decreased the expression of E-cadherin, with concomitant activation of NF-κB signaling pathway. Moreover, administration of the NF-κB inhibitor attenuated RANKL-induced migration, invasion and epithelial-mesenchymal transition of HCC cells. Conclusions RANKL could potentiate migration and invasion ability of RANK-positive HCC cells through NF-κB pathway-mediated epithelial-mesenchymal transition, which means that RANKL-RANK axis could be a potential target for HCC therapy.
    Full-text · Article · Sep 2014 · PLoS ONE
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