Research IndicatorsGraph generated 11 March 2017 using data from PubMed using criteria.
Mouse over the terms for more detail; many indicate links which you can click for dedicated pages about the topic. Tag cloud generated 11 March, 2017 using data from PubMed, MeSH and CancerIndex
Specific Cancers (6)
Data table showing topics related to specific cancers and associated disorders. Scope includes mutations and abnormal protein expression.
Note: list is not exhaustive. Number of papers are based on searches of PubMed (click on topic title for arbitrary criteria used).
OMIM, Johns Hopkin University
Referenced article focusing on the relationship between phenotype and genotype.
International Cancer Genome Consortium.
Summary of gene and mutations by cancer type from ICGC
Cancer Genome Anatomy Project, NCI
COSMIC, Sanger Institute
Somatic mutation information and related details
GEO Profiles, NCBI
Search the gene expression profiles from curated DataSets in the Gene Expression Omnibus (GEO) repository.
Latest Publications: TNFSF11 (cancer-related)
Zhu W, Hu X, Xu J, et al.Effect of PI3K/Akt Signaling Pathway on the Process of Prostate Cancer Metastasis to Bone.
Cell Biochem Biophys. 2015; 72(1):171-7 [PubMed
] Related Publications
We sought to study the effects of PI3K/Akt pathway and its downstream substrate NF-κB on prostate cancer bone metastatic process. Expression level of active p-Akt in PC3 cells was upregulated by transient expression with constitutively active plasmid CA-Akt or, alternatively, suppressed by dominant negative construct DN-Akt. NF-κB activity was determined by luciferase reporter assays. mRNA and protein expressions of receptor activator of NF-κB ligand (RANKL), parathyroid hormone-related protein (PTHrP), and bone morphogenetic protein 2 (BMP-2) were evaluated using RT-PCR and Western blotting. The effect of cross-talk between PC3 and SaOS2 cells on cell proliferation was analyzed using a co-culture system. Stimulation of p-Akt promoted NF-κB activity, and led to an increase in mRNA and protein expressions of RANKL, PTHrP, and BMP-2 in PC3 PCa cells through NF-κB. Co-culturing PC3 and SaOS2 cells significantly increased the expression of p-Akt and the activity of NF-κB, and promoted proliferation of both PC3 and SaOS2 cells. Increasing expression levels of p-Akt by transfection with CA-Akt led to further increase in cells proliferation, whereas NF-κB inhibitor PDTC partially blocked this effect. PI3K/Akt pathway stimulates the expressions of RANKL, PTHrP, and BMP-2 partly through NF-κB, suggesting its importance for bone metastasis of prostate carcinoma. Interaction of prostate cancer cells with bone cells has a stimulatory effect on cell proliferation.
Cellular interactions with the extracellular matrix play critical roles in tumor progression. We previously reported that receptor activator of NF-κB ligand (RANKL) specifically facilitates head and neck squamous cell carcinoma (HNSCC) progression in vivo. Here, we report a novel role for RANKL in the regulation of cell adhesion. Among the major type I collagen receptors, integrin α2 was significantly upregulated in RANKL-expressing cells, and its knockdown suppressed cell adhesion. The mRNA abundance of integrin α2 positively correlated with that of RANKL in human HNSCC tissues. We also revealed that RANK-NF-κB signaling mediated integrin α2 expression in an autocrine/paracrine manner. Interestingly, the amount of active integrin β1 on the cell surface was increased in RANKL-expressing cells through the upregulation of integrin α2 and endocytosis. Moreover, the RANK-integrin α2 pathway contributed to RANKL-dependent enhanced survival in a collagen gel and inhibited apoptosis in a xenograft model, demonstrating an important role for RANKL-mediated cell adhesion in three-dimensional environments.
Xie T, Ren HY, Lin HQ, et al.Sinomenine prevents metastasis of human osteosarcoma cells via S phase arrest and suppression of tumor-related neovascularization and osteolysis through the CXCR4-STAT3 pathway.
Int J Oncol. 2016; 48(5):2098-112 [PubMed
] Related Publications
Osteosarcoma is the most common primary malignant tumor of the bone. The long-term survivals continue to be unsatisfactory for patients with metastatic and recurrent disease. Metastasis is still a severe challenge in osteosarcoma treatment. Sinomenine, an alkaloid from traditional Chinese medicine, has been proved to possess potent antitumor and anti-invasion effect on various cancers. However, the effect of sinomenine on human osteosarcoma and the underlying mechanisms remains unknown. We report here that sinomenine inhibited proliferation by inducing S phase arrest and suppressing the clone formation. Significant inhibitory effects were found in invasion and metastasis in osteosarcoma, but little cytotoxicity was observed in tested concentrations. Exposure to sinomenine resulted in suppression of invasion and migration in osteosarcoma cells as well as tube formation ability in the human umbilical vein endothelial cells (HUVEC) and U2OS cells. Furthermore, it demonstrated that CXCR4 played a key role contributing to invasion in osteosarcoma which is considered to be a core target site in sinomenine treatment. Sinomenine inhibited invasion by suppressing CXCR4 and STAT3 phosphorylation then downregulating the expression of MMP-2, MMP-9, RANKL, VEGF downstream. In addition, then RANKL-mediated bone destruction stimulated by osteoclastogenesis and VEGF-related neovascularization were restrained. Importantly, in vivo, sinomenine suppressed proliferation, osteoclastogenesis and bone destruction. Through these various comprehensive means, sinomenine inhibits metastasis in osteosarcoma. Taken together, our results revealed that sinomenine caused S phase arrest, inhibited invasion and metastasis via suppressing the CXCR4-STAT3 pathway and then osteoclastogenesis-mediated bone destruction and neovascularization in osteosarcoma. Sinomenine is therefore a promising adjuvant agent for metastasis control in osteosarcoma.
Vyzoukaki R, Tsirakis G, Pappa CA, et al.The Impact of Mast Cell Density on the Progression of Bone Disease in Multiple Myeloma Patients.
Int Arch Allergy Immunol. 2015; 168(4):263-8 [PubMed
] Related Publications
BACKGROUND: Osteolytic bone disease is a major hallmark in multiple myeloma (MM) progression and affects many patients. Several inflammatory cells are involved in MM progression. Among them, mast cells (MCs) accumulated in the bone marrow (BM) microenvironment are known to play an important role in the mechanism of neovascularization.
METHODS: In 52 newly diagnosed active MM patients, we measured BM MC density (MCD) using an immunohistochemical stain for tryptase, serum levels of matrix metalloproteinase-9 (MMP-9) and receptor activator of nuclear factor x03BA;B ligand (RANKL) by a solid-phase sandwich enzyme-linked immunosorbent assay, along with urine levels of N-terminal propeptide of procollagen type I (Ntx) by a competitive inhibition enzyme-linked immunosorbent assay, in various clinical stages and skeletal grades.
RESULTS: MCD, RANKL and Ntx were higher in MM patients. All values increased in association with both the clinical stage and skeletal grade. Furthermore, MCD correlated positively with MMP-9, RANKL and Ntx.
CONCLUSIONS: Our data suggest that MCs may contribute to osteolytic processes during MM progression. Although the major role of MCs in tumor progression is to enhance angiogenesis, it seems that they may affect MM bone disease and may secrete a plethora of mediators that may directly and indirectly have an impact on osteolysis.
Su T, Li J, Meng M, et al.Bone marrow stromal cells induced activation of nuclear factor κB signaling protects non-Hodgkin's B lymphoma cells from apoptosis.
Tumour Biol. 2016; 37(8):10745-52 [PubMed
] Related Publications
The microenvironment encompassing a variety of non-malignant cells in close proximity with malignant tumor cells has been well known to significantly affect the behavior of tumor cells. In this study, we therefore studied the mechanism of bone marrow stromal cells in protection of lymphoma cells from spontaneous apoptosis. We demonstrated that adhesion of the freshly isolated lymphoma B cells to bone marrow stromal cells or freshly isolated lymphoma stromal cells inhibited B cell spontaneous apoptosis in culture. This inhibition of apoptosis correlated with decreased cleavage of caspase-3/8 and increased activation of canonical and non-canonical NF-κB signaling pathway. In addition to BAFF signaling which has been reported as a functional determinant for B lymphoma cell survival in the bone marrow environment, we demonstrated RANKL from BMSCs works synergistically with BAFF to activate NF-κB signaling pathway and thus protects lymphoma B cells from spontaneous apoptosis.
Giant cell tumors of bone (GCTB) are locally aggressive osteolytic bone tumors. Recently, some clinical trials have shown that denosumab is a novel and effective therapeutic option for aggressive and recurrent GCTB. This study was performed to investigate the molecular mechanism underlying the therapeutic effect of denosumab. Comparative proteomic analyses were performed using GCTB samples which were taken before and after denosumab treatment. Each expression profile was analyzed using the software program to further understand the affected biological network. One of identified proteins was further evaluated by gelatin zymography and an immunohistochemical analysis. We identified 13 consistently upregulated proteins and 19 consistently downregulated proteins in the pre- and post-denosumab samples. Using these profiles, the software program identified molecular interactions between the differentially expressed proteins that were indirectly involved in the RANK/RANKL pathway and in several non-canonical subpathways including the Matrix metalloproteinase pathway. The data analysis also suggested that the identified proteins play a critical functional role in the osteolytic process of GCTB. Among the most downregulated proteins, the activity of MMP-9 was significantly decreased in the denosumab-treated samples, although the residual stromal cells were found to express MMP-9 by an immunohistochemical analysis. The expression level of MMP-9 in the primary GCTB samples was not correlated with any clinicopathological factors, including patient outcomes. Although the replacement of tumors by fibro-osseous tissue or the diminishment of osteoclast-like giant cells have been shown as therapeutic effects of denosumab, the residual tumor after denosumab treatment, which is composed of only stromal cells, might be capable of causing bone destruction; thus the therapeutic application of denosumab would be still necessary for these lesions. We believe that the protein expression patterns and the results of the network analysis will provide a better understanding of the effects of denosumab administration in patients with GCTB.
Yoshimoto S, Morita H, Matsubara R, et al.Surface vacuolar ATPase in ameloblastoma contributes to tumor invasion of the jaw bone.
Int J Oncol. 2016; 48(3):1258-70 [PubMed
] Related Publications
Ameloblastoma is the most common benign odontogenic tumor in Japan. It is believed that it expands in the jaw bone through peritumoral activation of osteoclasts by receptor activator of nuclear factor kappa-B ligand (RANKL) released from the ameloblastoma, as in bone metastases of cancer cells. However, the clinical features of ameloblastoma, including its growth rate and patterns of invasion, are quite different from those of bone metastasis of cancer cells, suggesting that different underlying mechanisms are involved. Therefore, in the present study, we examined the possible mechanisms underlying the invasive expansion of ameloblastoma in the jaw bone. Expression levels of RANKL assessed by western blotting were markedly lower in ameloblastoma (AM-1) cells than in highly metastatic oral squamous cell carcinoma (HSC-3) cells. Experiments coculturing mouse macrophages (RAW264.7) with AM-1 demonstrated low osteoclastogenic activity, as assessed by tartrate-resistant acid phosphatase (TRAP)-positive multinuclear cell formation, probably because of low release of RANKL, whereas cocultures of RAW264.7 with HSC-3 cells exhibited very high osteoclastogenic activity. Thus, RANKL release from AM-1 appeared to be too low to generate osteoclasts. However, AM-1 cultured directly on calcium phosphate-coated plates formed resorption pits, and this was inhibited by application of bafilomycin A1. Furthermore, vacuolar-type H+-ATPase (V-ATPase) and H+/Cl- exchange transporter 7 (CLC-7) were detected on the surface of AM-1 cells by plasma membrane biotinylation and immunofluorescence analysis. Immunohistochemical analysis of clinical samples of ameloblastoma also showed plasma membrane-localized V-ATPase and CLC-7 in the epithelium of plexiform, follicular and basal cell types. The demineralization activity of AM-1 was only 1.7% of osteoclasts demineralization activity, and the growth rate was 20% of human normal skin keratinocytes and HSC-3 cells. These results suggest that the slow expansion of several typical types of ameloblastomas in jaw bone is attributable to its slow growth and low demineralization ability.
Owen S, Sanders AJ, Mason MD, Jiang WGImportance of osteoprotegrin and receptor activator of nuclear factor κB in breast cancer response to hepatocyte growth factor and the bone microenvironment in vitro.
Int J Oncol. 2016; 48(3):919-28 [PubMed
] Free Access to Full Article Related Publications
Osteoprotegrin (OPG), receptor activator of nuclear factor κB (RANK) and RANK ligand (RANKL) are signal transducers which have pleiotropic actions. Each tumour necrosis factor receptor superfamily member has unique structural attributes which directly couples them to signalling pathways involved in cell proliferation, differentiation and survival. Previous studies have clinically linked OPG, RANK and RANKL to increasing tumour burden, metastatic bone involvement and estrogen status. This study aimed to establish the potential implications of targeting endogenously produced OPG and RANK in the osteotropic breast cancer cell line MDA-MB‑231 in vitro. Subsequently this study also aimed to explore the potential links between these molecules with regards to hepatocyte growth factor (HGF) signalling and extracted bone proteins (BME). OPG and RANK expression was successfully suppressed using hammerhead ribozyme technology. Subsequently effects were explored in MDA-MB‑231 cell proliferation, matrix adhesion, migration and invasion in vitro function assays. Reduced OPG expression resulted in increased breast cancer cell migration and invasion. These increases, particularly invasion, appeared to however be reduced under the influence of the exogenous stimuli (HGF and BME). In contrast, suppression of RANK in MDA-MB‑231 breast cancer cells resulted in decreased cancer cell proliferation, matrix-adhesion, motility and invasion with little cumulative effect being noted after the addition of exogenous stimuli. The complexity of the bone environment underpins the vast number of soluble factors and signalling pathways which can influence osteotropic cancer behaviour and progression. Further work into elucidating all the pathways affected could potentially lead to better identification of those patients most at risk.
Tsubaki M, Takeda T, Yoshizumi M, et al.RANK-RANKL interactions are involved in cell adhesion-mediated drug resistance in multiple myeloma cell lines.
Tumour Biol. 2016; 37(7):9099-110 [PubMed
] Related Publications
Interaction between multiple myeloma (MM) cells and the bone marrow microenvironment plays a critical role in MM pathogenesis and the development of drug resistance. Recently, it has been reported that MM cells express the receptor activator of nuclear factor-κB (NF-κB) (RANK). However, the role of the RANK/RANK ligand (RANKL) system in drug resistance remains unclear. In this study, we demonstrated a novel function of the RANK/RANKL system in promoting drug resistance in MM. We found that RANKL treatment induced drug resistance in RANK-expressing but not RANK-negative cell lines. RANKL stimulation of RANK-expressing cells increased multidrug resistance protein 1 (MDR1), breast cancer resistance protein (BCRP), and lung resistance protein 1 (LRP1) expression and decreased Bim expression through various signaling molecules. RNA silencing of Bim expression induced drug resistance, but the RANKL-mediated drug resistance could not be overcome through the RNA silencing of MDR1, BCRP, and LRP1 expression. These results indicate that the RANK/RANKL system induces chemoresistance through the activation of multiple signal transduction pathways and by decreasing Bim expression in RANK-positive MM cells. These findings may prove to be useful in the development of cell adhesion-mediated drug resistance inhibitors in RANK-positive MM cells.
RANK/RANKL plays a key role in metastasis of certain malignant tumors, which makes it a promising target for developing novel therapeutic strategies for cancer. However, the prognostic value and pro-metastatic activity of RANK in endometrial cancer (EC) remain to be determined. Thus, the present study investigated the effect of RANK on the prognosis of EC patients, as well as the pro-metastatic activity of EC cells. The results indicated that those with high expression of RANK showed decreased overall survival and progression-free survival. Statistical analysis revealed the positive correlations between RANK/RANKL expression and metastasis-related factors. Additionally, RANK/RANKL significantly promoted cell migration/invasion via activating AKT/β-catenin/Snail pathway in vitro. However, RANK/RANKL-induced AKT activation could be suppressed after osteoprotegerin (OPG) treatment. Furthermore, the combination of medroxyprogesterone acetate (MPA) and RANKL could in turn attenuate the effect of RANKL alone. Similarly, MPA could partially inhibit the RANK-induced metastasis in an orthotopic mouse model via suppressing AKT/β-catenin/Snail pathway. Therefore, therapeutic inhibition of MPA in RANK/RANKL-induced metastasis was mediated by AKT/β-catenin/Snail pathway both in vitro and in vivo, suggesting a potential target of RANK for gene-based therapy for EC.
Sánchez CA, Andahur EI, Valenzuela R, et al.Exosomes from bulk and stem cells from human prostate cancer have a differential microRNA content that contributes cooperatively over local and pre-metastatic niche.
Oncotarget. 2016; 7(4):3993-4008 [PubMed
] Free Access to Full Article Related Publications
The different prostate cancer (PCa) cell populations (bulk and cancer stem cells, CSCs) release exosomes that contain miRNAs that could modify the local or premetastatic niche. The analysis of the differential expression of miRNAs in exosomes allows evaluating the differential biological effect of both populations on the niche, and the identification of potential biomarkers and therapeutic targets. Five PCa primary cell cultures were established to originate bulk and CSCs cultures. From them, exosomes were purified by precipitation for miRNAs extraction to perform a comparative profile of miRNAs by next generation sequencing in an Illumina platform. 1839 miRNAs were identified in the exosomes. Of these 990 were known miRNAs, from which only 19 were significantly differentially expressed: 6 were overexpressed in CSCs and 13 in bulk cells exosomes. miR-100-5p and miR-21-5p were the most abundant miRNAs. Bioinformatics analysis indicated that differentially expressed miRNAs are highly related with PCa carcinogenesis, fibroblast proliferation, differentiation and migration, and angiogenesis. Besides, miRNAs from bulk cells affects osteoblast differentiation. Later, their effect was evaluated in normal prostate fibroblasts (WPMY-1) where transfection with miR-100-5p, miR-21-5p and miR-139-5p increased the expression of metalloproteinases (MMPs) -2, -9 and -13 and RANKL and fibroblast migration. The higher effect was achieved with miR21 transfection. As conclusion, miRNAs have a differential pattern between PCa bulk and CSCs exosomes that act collaboratively in PCa progression and metastasis. The most abundant miRNAs in PCa exosomes are interesting potential biomarkers and therapeutic targets.
Chen Y, Di Grappa MA, Molyneux SD, et al.RANKL blockade prevents and treats aggressive osteosarcomas.
Sci Transl Med. 2015; 7(317):317ra197 [PubMed
] Related Publications
Osteosarcoma (OS) is the most common primary bone cancer, which occurs primarily in children and adolescents, severely affecting survivors' quality of life. Despite its chemosensitivity and treatment advances, long-term survival rates for OS patients have stagnated over the last 20 years. Thus, it is necessary to develop new molecularly targeted therapies for this metastatic bone cancer. Mutations in TP53 and RB are linked to OS predisposition and to the evolution of spontaneous OS. We established receptor activator of nuclear factor κB ligand (RANKL) as a therapeutic target for suppression and prevention of OS. Combined conditional osteoblast-specific deletions of Rb, p53, and the protein kinase A (PKA) regulatory subunit Prkar1α genes in genetically engineered mouse models (GEMMs) generate aggressive osteosarcomas, characterized by PKA, RANKL, and osteoclast hyperactivity. Whole-body Rankl deletion completely abrogates tumorigenesis. Although osteoblastic Rank deletion has little effect, osteoclastic Rank deletion delays tumorigenesis and prolongs life span. The latter is associated with inactivation of osteoclastogenesis and up-regulation of the tumor suppressor phosphatase and tensin homolog (PTEN). Further, we use these GEMMs as preclinical platforms to show that RANKL blockade with RANK-Fc arrests tumor progression and improves survival and also inhibits lung metastasis. Moreover, preemptive administration of RANK-Fc completely prevents tumorigenesis in mice highly predisposed to this aggressive cancer. Denosumab, a fully human monoclonal antibody against RANKL, is currently used to treat patients with osteoporosis or bone metastases. Our studies provide a strong rationale to consider RANKL blockade for the treatment and prevention of aggressive RANKL-overexpressing OS in humans.
RESEARCH BACKGROUND: Currently, multiple myeloma is the second most common hematological malignancy in the U.S., constituting 1% of all cancers. With conventional treatment, the median survival time is typically 3-4 years, although it can be extended to 5-7 years or longer with advanced treatments. Recent research indicated that an increase in osteoclast (OC) activity is often associated withmultiple myeloma (MM) and that a decrease inosteoblast (OB) activity contributesto the osteolytic lesions in MM. Normally, the populations of OCs and OBs are inequilibrium, and an imbalance in this statecontributes to the development of lesions.
RESEARCH PROCEDURES: A multi-scale agent-based multiple myeloma model was developed to simulate the proliferation, migration and death of OBs and OCs. Subsequently, this model was employed to investigate the efficacy of thethree most commonly used drugs for MM treatment under the following two premises: the reduction in the progression of MM and the re-establishment of the equilibrium between OCs and OBs.
RESEARCH PURPOSES: The simulated results not only demonstrated the capacity of the model to choose optimal combinations of the drugs but also showed that the optimal use of the three drugs can restore the balance between OCs and OBs as well as kill MMs. Furthermore, the drug synergism analysis function of the model revealed that restoring the balance between OBs and OCs can significantly increase the efficacy of drugs against tumor cells.
Widschwendter M, Burnell M, Fraser L, et al.Osteoprotegerin (OPG), The Endogenous Inhibitor of Receptor Activator of NF-κB Ligand (RANKL), is Dysregulated in BRCA Mutation Carriers.
EBioMedicine. 2015; 2(10):1331-9 [PubMed
] Free Access to Full Article Related Publications
Breast cancer development in BRCA1/2 mutation carriers is a net consequence of cell-autonomous and cell nonautonomous factors which may serve as excellent targets for cancer prevention. In light of our previous data we sought to investigate the consequences of the BRCA-mutation carrier state on RANKL/osteoprotegerin (OPG) signalling. We analysed serum levels of RANKL, OPG, RANKL/OPG complex, oestradiol (E2), and progesterone (P) during menstrual cycle progression in 391 BRCA1/2-mutation carriers and 782 noncarriers. These studies were complemented by analyses of RANKL and OPG in the serum and mammary tissues of female cynomolgus macaques (n = 88) and serum RANKL and OPG in postmenopausal women (n = 150). BRCA-mutation carriers had lower mean values of free serum OPG in particular in BRCA1-mutation carriers (p = 0.018) compared with controls. Among BRCA1/2 mutation carriers, lower OPG levels were associated with germline mutation locations known to confer an increased breast cancer risk (p = 0.003). P is associated with low OPG levels in serum and tissue, particularly in BRCA-mutation carriers (rho = - 0.216; p = 0.002). Serum OPG levels were inversely correlated (rho = - 0.545, p < 0.001) with mammary epithelial proliferation measured by Ki67 expression and increased (p = 0.01) in postmenopause. The P-RANKL/OPG system is dysregulated in BRCA-mutation carriers. These and previously published data provide a strong rationale for further investigation of antiprogestogens or an anti-RANKL antibody such as denosumab for breast cancer prevention.
FYN is a SRC family kinase (SFK) that has been shown to be up-regulated in human prostate cancer (PCa) tissues and cell lines. In this study, we observed that FYN is strongly up-regulated in human neuroendocrine PCa (NEPC) tissues and xenografts, as well as cells derived from a NEPC transgenic mouse model. In silico analysis of FYN expression in prostate cancer cell line databases revealed an association with the expression of neuroendocrine (NE) markers such as CHGA, CD44, CD56, and SYP. The loss of FYN abrogated the invasion of PC3 and ARCaPM cells in response to MET receptor ligand HGF. FYN also contributed to the metastatic potential of NEPC cells in two mouse models of visceral metastasis with two different cell lines (PC3 and TRAMPC2-RANKL). The activation of MET appeared to regulate neuroendocrine (NE) features as evidenced by increased expression of NE markers in PC3 cells with HGF. Importantly, the overexpression of FYN protein in DU145 cells was directly correlated with the increase of CHGA. Thus, our data demonstrated that the neuroendocrine differentiation that occurs in PCa cells is, at least in part, regulated by FYN kinase. Understanding the role of FYN in the regulation of NE markers will provide further support for ongoing clinical trials of SFK and MET inhibitors in castration-resistant PCa patients.
BACKGROUND: Osteoprotegerin (OPG) is a glycoprotein that has multifaceted role and is associated with several cancer malignancies like that of bladder carcinoma, gastric carcinoma, prostate cancer, multiple myeloma and breast cancer. Also OPG has been associated with several organ pathologies. The widespread expression of OPG suggests that OPG may have multiple biological activities that are yet to be explored.
METHODS: The anchorage-independent sphere cultures of the adherent cells were instrumental in our study as it provided a deeper insight into the complexity of a 3D tumor. Cytokine profiling was performed for OPG's detection in the microenvironment. ELISA and western blotting were performed to quantify the OPG secretion and measure the protein levels respectively. OPG expression was detected in human breast cancer tissue samples by IHC. To decipher OPG's role in tumor aggressiveness both recombinant human OPG as well as OPG rich and depleted breast cancer cell conditioned media were tested. Western blotting and MTT assay were performed to detect changes in signaling pathways and proliferation that were induced in presence of OPG. Onset of aneuploidy, in presence of OPG, was measured by cell cycle analysis and western blotting. Finally, human Breast Cancer qBiomarker Copy Number PCR Array was used to detect how OPG remarkably induced gene copy numbers for oncogenic pathway regulators.
RESULTS: SUM149PT and SUM1315M02 cells secrete high levels of the cytokine OPG compared to primary human mammary epithelial cells (HMEC). High expression of OPG was also detected in human breast cancer tissue samples compared to the uninvolved tissue from the same patient. OPG induced proliferation of control HMEC spheres and triggered the onset of aneuploidy in HMEC sphere cultures. OPG induced the expression of aneuploidy related kinases Aurora-A Kinase (IAK-1), Bub1 and BubR1 probably through the receptor activator of nuclear factor kappa-B ligand (RANKL) and syndecan-1 receptors via the Erk, AKT and GSK3(3 signaling pathway. Gene copy numbers for oncogenic pathway regulators such AKT1, Aurora-A Kinase (AURKA or IAK-1), epidermal growth factor receptor (EGFR) and MYC with a reduction in the copy numbers of cyclin dependent kinase inhibitor 2A (CDKN2A), PTEN and DNA topoisomerase 2 alpha (TOP2A) were induced in presence of OPG.
CONCLUSIONS: These results highlight the role of OPG in reprogramming normal mammary epithelial cells to a tumorigenic state and suggest promising avenues for treating inflammatory breast cancer as well as highly invasive breast cancer with new therapeutic targets.
Zong JC, Wang X, Zhou X, et al.Gut-derived serotonin induced by depression promotes breast cancer bone metastasis through the RUNX2/PTHrP/RANKL pathway in mice.
Oncol Rep. 2016; 35(2):739-48 [PubMed
] Related Publications
Breast cancer metastasizes to the bone in a majority of patients with advanced disease resulting in bone destruction. The underlying mechanisms are complex, and both processes are controlled by an interaction between locally and systemically derived signals. Clinically, breast cancer patients with depression have a higher risk of bone metastasis, yet the etiology and mechanisms are yet to be elucidated. MDA‑MB‑231 breast cancer cells were used to establish a bone metastasis model by using intracardiac injection in nude mice. Chronic mild stress (CMS) was chosen as a model of depression in mice before and after inoculation of the cells. Knockdown of the RUNX‑2 gene was performed by transfection of the cells with shRNA silencing vectors against human RUNX‑2. A co‑culture system was used to test the effect of the MDA‑MB‑231 cells on osteoclasts and osteoblasts. RT‑PCR and western blotting were used to test gene and protein expression, respectively. We confirmed that depression induced bone metastasis by promoting osteoclast activity while inhibiting osteoblast differentiation. Free serotonin led to an increase in the expression of RUNX2 in breast cancer cells (MDA‑MB‑231), which directly inhibited osteoblast differentiation and stimulated osteoclast differentiation by the PTHrP/RANKL pathway, which caused bone destruction and formed osteolytic bone lesions. Additionally, the interaction between depression and breast cancer cells was interrupted by LP533401 or RUNX2 knockdown. In conclusion, depression promotes breast cancer bone metastasis partly through increasing levels of gut‑derived serotonin. Activation of RUNX2 in breast cancer cells by circulating serotonin appears to dissociate coupling between osteoblasts and osteoclasts, suggesting that the suppression of gut‑derived serotonin decreases the rate of breast cancer bone metastasis induced by depression.
Beuselinck B, Jean-Baptiste J, Couchy G, et al.RANK/OPG ratio of expression in primary clear-cell renal cell carcinoma is associated with bone metastasis and prognosis in patients treated with anti-VEGFR-TKIs.
Br J Cancer. 2015; 113(9):1313-22 [PubMed
] Free Access to Full Article Related Publications
BACKGROUND: Bone metastases (BMs) are associated with poor outcome in metastatic clear-cell renal carcinoma (m-ccRCC) treated with anti-vascular endothelial growth factor tyrosine kinase inhibitors (anti-VEGFR-TKIs). We aimed to investigate whether expression in the primary tumour of genes involved in the development of BM is associated with outcome in m-ccRCC patients treated with anti-VEGFR-TKIs.
METHODS: Metastatic clear-cell renal cell carcinoma patients with available fresh-frozen tumour and treated with anti-VEGFR-TKIs. Quantitative real-time PCR (qRT-PCR) for receptor activator of NF-kB (RANK), RANK-ligand (RANKL), osteoprotegerin (OPG), the proto-oncogene SRC and DKK1 (Dickkopf WNT signalling pathway inhibitor-1). Time-to-event analysis by Kaplan-Meier estimates and Cox regression.
RESULTS: We included 129 m-ccRCC patients treated between 2005 and 2013. An elevated RANK/OPG ratio was associated with shorter median time to metastasis (HR 0.50 (95% CI 0.29-0.87); P=0.014), shorter time to BM (HR 0.54 (95% CI 0.31-0.97); P=0.037), shorter median overall survival (mOS) since initial diagnosis (HR 2.27 (95% CI 1.44-3.60); P=0.0001), shorter median progression-free survival (HR 0.44 (95% CI 0.28-0.71); P=0.001) and mOS (HR 0.31 (95% CI 0.19-0.52); P<0.0001) on first-line anti-VEGFR-TKIs in the metastatic setting. Higher RANK expression was associated with shorter mOS on first-line anti-VEGFR-TKIs (HR 0.46 (95% CI 0.29-0.73); P=0.001).
CONCLUSIONS: RANK/OPG ratio of expression in primary ccRCC is associated with BM and prognosis in patients treated with anti-VEGFR-TKIs. Prospective validation is warranted.
Shang WQ, Li H, Liu LB, et al.RANKL/RANK interaction promotes the growth of cervical cancer cells by strengthening the dialogue between cervical cancer cells and regulation of IL-8 secretion.
Oncol Rep. 2015; 34(6):3007-16 [PubMed
] Related Publications
Receptor activator for nuclear factor κB ligand (RANKL) is a member of the tumor necrosis factor (TNF) family. The interaction between RANKL and its receptor RANK plays an important role in the development and function of diverse tissues. However, the expression and role of RANKL in cervical cancer are still unknown. In the present study, we found that RANKL and RANK were highly co-expressed in cervical cancer. HeLa and SiHa cells secreted soluble RANKL (sRANKL), expressed member RANKL (mRANKL) and RANK. Recombinant human RANKL protein had no effect on the viability of HeLa and SiHa cells. Yet, blocking RANKL with an anti-human RANKL neutralizing antibody (α-RANKL) or recombinant human osteoprotegrin (OPG) protein resulted in the downregulation of Ki-67 and B-cell lymphoma 2 (Bcl-2) expression and an increase in Fas and Fas ligand (FasL) expression, as well as a high level of viability and a low level of apoptosis in the HeLa and SiHa cells. In addition, α-RANKL led to a decrease in IL-8 secretion. Recombinant human IL-8 protein reversed the effect of α-RANKL on the expression of proliferation- and apoptosis‑related molecules, and proliferation and apoptosis in the HeLa and SiHa cells. The present study suggests that a high level of mRANKL/RANK expression in cervical cancer lesions plays an important role in the rapid growth of cervical cancer cells possibly through strengthening the dialogue between cervical cancer cells and regulation of IL-8 secretion, which may be a possible target for cervical cancer therapy.
Fluorosis is caused by excess of fluoride intake over a long period of time. Aberrant change in the Runt-related transcription factor 2 (RUNX2) mediated signaling cascade is one of the decisive steps during the pathogenesis of fluorosis. Up to date, role of fluoride on the epigenetic alterations is not studied. In the present study, global expression profiling of short noncoding RNAs, in particular miRNAs and snoRNAs, was carried out in sodium fluoride (NaF) treated human osteosarcoma (HOS) cells to understand their possible role in the development of fluorosis. qPCR and in silico hybridization revealed that miR-124 and miR-155 can be directly involved in the transcriptional regulation of Runt-related transcription factor 2 (RUNX2) and receptor activator of nuclear factor κ-B ligand (RANKL) genes. Compared to control, C/D box analysis revealed marked elevation in the number of UG dinucleotides and D-box sequences in NaF exposed HOS cells. Herein, we report miR-124 and miR-155 as the new possible players involved in the development of fluorosis. We show that the alterations in UG dinucleotides and D-box sequences of snoRNAs could be due to NaF exposure.
Non-small cell lung cancer (NSCLC) frequently metastasizes to bone, which is associated with significant morbidity and a dismal prognosis. RUNX3 functions as a tumour suppressor in lung cancer and loss of expression occurs more frequently in invasive lung adenocarcinoma than in pre-invasive lesions. Here, we show that RUNX3 and RUNX3-regulated chemokines are linked to NSCLC-mediated bone resorption. Notably, the receptor activator of nuclear factor-κB ligand (RANKL)/osteoprotegerin (OPG) ratio, an index of osteoclastogenic stimulation, was significantly increased in human osteoblastic cells treated with conditioned media derived from RUNX3-knockdown NSCLC cells. We aimed to identify RUNX3-regulated factors that modify the osteoblastic RANKL/OPG ratio and found that RUNX3 knockdown led to CCL5 up-regulation and down-regulation of CCL19 and CXCL11 in NSCLC cells. Tumour size was noticeably increased and more severe osteolytic lesions were induced in the calvaria and tibiae of mice that received RUNX3-knockdown cells. In response to RUNX3 knockdown, serum and tissue levels of CCL5 increased, whereas CCL19 and CXCL11 decreased. Furthermore, CCL5 increased the proliferation, migration, and invasion of lung cancer cells in a dose-dependent manner; however, CCL19 and CXCL11 did not show any significant effects. The RANKL/OPG ratio in osteoblastic cells was increased by CCL5 but reduced by CCL19 and CXCL11. CCL5 promoted osteoclast differentiation, but CCL19 and CXCL11 reduced osteoclastogenesis in RANKL-treated bone marrow macrophages. These findings suggest that RUNX3 and related chemokines are useful markers for the prediction and/or treatment of NSCLC-induced bone destruction.
Li R, Zhang K, Penedo TL, et al.The RANK Pathway in Advanced Breast Cancer: Does Src Play a Role?
Appl Immunohistochem Mol Morphol. 2016; 24(1):42-50 [PubMed
] Related Publications
Receptor activator of NF-κB (RANK) and its ligand, RANKL, are essential for osteoclastogenesis and modulate osteolytic bone metastasis. The RANKL/RANK system is also fundamental for mammary gland development and plays a potential role in breast carcinogenesis. c-Src, a nonreceptor tyrosine kinase downstream of RANK, is overexpressed in most breast cancers and plays a key role in several transduction pathways. The aim of the study was to examine the expression of these molecules in tissue microarrays constructed from 62 advanced breast cancers and 10 breast cancers controls (no metastasis after follow-up). Significantly higher levels of RANK and lower levels of RANKL were found in triple-negative (ER-/PR-/HER2-) tumors when compared with luminal subtypes, whereas their levels in the HER2 subtype were quantitatively in between. RANK expression was significantly associated with tumor grade/differentiation by multivariate analysis. Despite their high expression in bone, neither molecule in primary tumors seemed to be related to a bone-seeking phenotype. Rather, they were significantly correlated with a brain-metastatic phenotype. RANKL and RANK were significantly associated with survival outcomes. Further, Src expression showed a significantly positive linear relationship with RANK, suggesting a potential mechanism of the RANKL-RANK axis in regulating breast cancer cell differentiation and antiapoptosis. Thus, these molecules may be potential therapeutic targets, especially in triple-negative tumors, for which the only systemic treatment option is cytotoxic chemotherapy.
Brodowicz T, Hemetsberger M, Windhager RDenosumab for the treatment of giant cell tumor of the bone.
Future Oncol. 2015; 11(13):1881-94 [PubMed
] Related Publications
Giant cell tumor of bone is typically composed of neoplastic stromal cells and non-neoplastic osteoclastic giant cells. RANK-expressing osteoclastic giant cells are recruited by RANK ligand excreted by the stromal cells, and used by these neoplastic cells to create expansion space. Denosumab specifically binds to and inhibits RANK ligand, thereby eradicating osteoclastic giant cells from the tumor and thus reducing osteolytic activity. Clinical studies reported disease stabilization and clinical benefit in terms of reduced pain and analgesics use, avoided surgeries or surgeries with less morbid procedures. Adverse events observed in patients with giant cell tumor of bone were consistent with the known safety profile of denosumab with a very low incidence of hypocalcemia and osteonecrosis. Overall, denosumab was shown to suppress osteolytic activity and slow disease progression and is thus a treatment option for patients with giant cell tumor of bone.
Pitari MR, Rossi M, Amodio N, et al.Inhibition of miR-21 restores RANKL/OPG ratio in multiple myeloma-derived bone marrow stromal cells and impairs the resorbing activity of mature osteoclasts.
Oncotarget. 2015; 6(29):27343-58 [PubMed
] Free Access to Full Article Related Publications
miR-21 is an oncogenic microRNA (miRNA) with an emerging role as therapeutic target in human malignancies, including multiple myeloma (MM). Here we investigated whether miR-21 is involved in MM-related bone disease (BD). We found that miR-21 expression is dramatically enhanced, while osteoprotegerin (OPG) is strongly reduced, in bone marrow stromal cells (BMSCs) adherent to MM cells. On this basis, we validated the 3'UTR of OPG mRNA as miR-21 target. Constitutive miR-21 inhibition in lentiviral-transduced BMSCs adherent to MM cells restored OPG expression and secretion. Interestingly, miR-21 inhibition reduced RANKL production by BMSCs. Overexpression of protein inhibitor of activated STAT3 (PIAS3), which is a direct and validated target of miR-21, antagonized STAT3-mediated RANKL gene activation. Finally, we demonstrate that constitutive expression of miR-21 inhibitors in BMSCs restores RANKL/OPG balance and dramatically impairs the resorbing activity of mature osteoclasts. Taken together, our data provide proof-of-concept that miR-21 overexpression within MM-microenviroment plays a crucial role in bone resorption/apposition balance, supporting the design of innovative miR-21 inhibition-based strategies for MM-related BD.
Li Y, Li N, Yan Z, et al.Dysregulation of the NLRP3 inflammasome complex and related cytokines in patients with multiple myeloma.
Hematology. 2016; 21(3):144-51 [PubMed
] Related Publications
OBJECTIVE: The NLRP3 inflammasome complex, an important regulatory factor of inflammation and cell apoptosis, has attracted considerable attention in the development of tumor. Here, we analyzed the expression and clinical significance of NLRP3 inflammasome complex and related cytokines in patients with multiple myeloma (MM).
METHODS: Peripheral blood and bone marrow of 38 newly diagnosed myeloma patients and 25 age- and gender-matched healthy people were studied. NLRP3 and caspase-1 were analyzed using quantitative real-time polymerase chain reaction and Western blot and IL-1beta, IL-18, RANKL, and OPG were evaluated by enzyme-linked immunosorbent assay.
RESULTS: We showed that aberrant NLRP3 and caspase-1 expression were observed in MM and down-regulated compared with the healthy people. We further demonstrated that NLRP3 mRNA was negatively correlated with beta2-microglobulin and plasma cell percentage in MM. The downstream cytokines IL-18 and sRANKL/OPG in MM patients were higher than that in control group. Moreover, the lower mRNA levels of NLRP3 and caspase-1 were shown to be positively correlated with IL-1beta in newly diagnosed MM patients.
CONCLUSIONS: This study demonstrated that dysregulated expression of NLRP3-caspase-1-IL-1beta axis was observed in patients with MM, suggesting they might be involved in the pathogenesis of MM.
Autophagy is a cellular process to recycle nutrients and has been implicated in cancer treatment. Oral squamous cell carcinoma (OSCC) is the most common oral cancer which ranks 3% of cancers in men and 2% in women. In this study, immunohistochemical staining of OSCC tumor specimens from human subjects and an athymic mouse model demonstrated high levels of autophagy markers LC3-II and ATG5 expression. Further, we identified high levels LC3-II expression in OSCC tumor cell lines (SCC-1, SCC-12, and SCC-14a) compared to normal human epithelial (RWPE-1) cells. OSCC cells express high levels of RANK ligand (RANKL); however, a functional role in autophagy is unknown. Interestingly, RANKL stimulation significantly increased autophagosome-related gene expressions such as LC3, ATG5, BECN1, and PI3KC3 mRNA expression in OSCC cells. Further, Western blot analysis of total cell lysates demonstrated a dose-dependent increase in LC3-II and ATG5 expression in RANKL-stimulated cells. In addition, RANKL increased expression of LC3-I and LC3-II, essential for autophagosome formation. Confocal microscopy analysis of LC3-II and localization with lysosome further confirms autophagosome formation in response to RANKL treatment in OSCC cells. Collectively, our results indicate a novel function of RANKL to induce autophagosome formation, and could be a potential therapeutic target to control OSCC tumor progression.
Tsai YM, Chong IW, Hung JY, et al.Syringetin suppresses osteoclastogenesis mediated by osteoblasts in human lung adenocarcinoma.
Oncol Rep. 2015; 34(2):617-26 [PubMed
] Related Publications
Bone metastasis in lung cancer results in an unfavorable outcome for patients by not only impairing the quality of life, yet also increasing the cancer-related death rates. In the present study, we discuss a novel treatment strategy that may benefit these patients. Human CD14+ monocytes treated with macrophage-colony stimulating factor (M-CSF)/receptor activator of nuclear factor κB ligand (RANKL) differentiated into osteoclasts, whereas syringetin (SGN), a flavonoid derivative found in both grapes and wine, suppressed the osteoclastogenesis in vitro in a dose-dependent manner. In addition, SGN inhibited osteoclast formation induced by human lung adenocarcinoma A549 and CL1-5 cells. The associated signaling transduction pathway in osteoclastogenesis and SGN inhibition was found to be via the AKT/mammalian target of rapamycin (mTOR) signaling pathway. Blocking AKT and mTOR by respective inhibitors significantly decreased lung adenocarcinoma-mediated osteoclastogenesis. Moreover, SGN regulated the lung adenocarcinoma-mediated interaction between osteoblasts and osteoclasts by suppressing the stimulatory effect of lung adenocarcinoma on M-CSF and RANKL production in osteoblasts, and reversing the inhibitory effect of the lung adenocarcinoma on OPG production in osteoblasts. The present study has two novel findings. It is the first to illustrate lung adenocarcinoma-mediated interaction between osteoblasts and osteoclasts, leading to osteolytic bone metastasis. It also reveals that SGN, a flavonoid derivative, directly inhibits osteoclastogenesis and reverses lung adenocarcinoma-mediated osteoclastogenesis. In conclusion, the present study suggests that SGN, a natural compound, prevents and treats bone metastasis in patients with lung cancer.
BACKGROUND: DNA methylation regulates gene expression, through the inhibition/activation of gene transcription of methylated/unmethylated genes. Hence, DNA methylation profiling can capture pivotal features of gene expression in cancer tissues from patients at the time of diagnosis. In this work, we analyzed a breast cancer case series, to identify DNA methylation determinants of metastatic versus non-metastatic tumors.
METHODS: CpG-island methylation was evaluated on a 56-gene cancer-specific biomarker microarray in metastatic versus non-metastatic breast cancers in a multi-institutional case series of 123 breast cancer patients. Global statistical modeling and unsupervised hierarchical clustering were applied to identify a multi-gene binary classifier with high sensitivity and specificity. Network analysis was utilized to quantify the connectivity of the identified genes.
RESULTS: Seven genes (BRCA1, DAPK1, MSH2, CDKN2A, PGR, PRKCDBP, RANKL) were found informative for prognosis of metastatic diffusion and were used to calculate classifier accuracy versus the entire data-set. Individual-gene performances showed sensitivities of 63-79 %, 53-84 % specificities, positive predictive values of 59-83 % and negative predictive values of 63-80 %. When modelled together, these seven genes reached a sensitivity of 93 %, 100 % specificity, a positive predictive value of 100 % and a negative predictive value of 93 %, with high statistical power. Unsupervised hierarchical clustering independently confirmed these findings, in close agreement with the accuracy measurements. Network analyses indicated tight interrelationship between the identified genes, suggesting this to be a functionally-coordinated module, linked to breast cancer progression.
CONCLUSIONS: Our findings identify CpG-island methylation profiles with deep impact on clinical outcome, paving the way for use as novel prognostic assays in clinical settings.
Multiply myeloma (MM) grows in and destroys bone, where osteocytes secrete FGF23, a hormone which affects phosphate homeostasis and aging. We report that multiple myeloma (MM) cells express receptors for and respond to FGF23. FGF23 increased mRNA for EGR1 and its target heparanase, a pro-osteolytic factor in MM. FGF23 signals through a complex of klotho and a classical FGF receptor (FGFR); both were expressed by MM cell lines and patient samples. Bone marrow plasma cells from 42 MM patients stained positively for klotho, while plasma cells from 8 patients with monoclonal gammopathy of undetermined significance (MGUS) and 6 controls were negative. Intact, active FGF23 was increased 2.9X in sera of MM patients compared to controls. FGF23 was not expressed by human MM cells, but co-culture with mouse bone increased its mRNA. The FGFR inhibitor NVP-BGJ398 blocked the heparanase response to FGF23. NVP-BGJ398 did not inhibit 8226 growth in vitro but significantly suppressed growth in bone and induction of the osteoclast regulator RANK ligand, while decreasing heparanase mRNA. The bone microenvironment provides resistance to some anti-tumor drugs but increased the activity of NVP-BGJ398 against 8226 cells. The FGF23/klotho/heparanase signaling axis may offer targets for treatment of MM in bone.
Pigmented villonodular synovitis (PVNS) is a benign, translocation-derived neoplasm. Because of its high local recurrence rate after surgery and occurrence of osteochondral destruction, a novel therapeutic target is required. The present study aimed to evaluate the significance of protein expression possibly associated with the pathogenesis during the clinical course of PVNS. In 40 cases of PVNS, positivity of colony-stimulated factor 1 (CSF1), its receptor (CSF1R), and receptor activator of nuclear factor kappa-B ligand (RANKL) were immunohistochemically determined. The relationship between the positivity and clinical outcomes was investigated. High positivity of CSF1 staining intensity was associated with an increased incidence of osteochondral lesions (bone erosion and osteoarthritis) (p = 0.009), but not with the rate of local recurrence. Positivity of CSF1R and RANKL staining was not associated with any clinical variables. The number of giant cells was not correlated with positivity of any of the three proteins, or with the clinical outcome. Focusing on knee cases, CSF1 positivity was also associated with the incidence of osteochondal change (p = 0.02). CSF1R positivity was high in cases which had local recurrence, but not significantly so (p = 0.129). Determination of CSF1 and CSF1R expression may be useful as a prognosticator of the clinical course and/or outcomes of PVNS.