Bone Metastasis Formation Research Articles

RANKL: A promising circulating marker for bone metastasis response.

Bone metastases are a frequent event in patients with solid tumors. Although great advances have been made in the treatment of these patients, the identification of novel, accurate indicators of bone response would greatly facilitate the clinical management of the disease. The receptor activator of nuclear factor-κB (RANK)/RANK ligand (RANKL)/osteoprotegerin (OPG) signaling pathway is significantly involved in bone metastasis formation. The main aim of the present study was to evaluate the role of circulating RANK, RANKL and OPG levels in predicting bone response. Marker accuracy was also compared with that of the conventional tumor marker N-terminal telopeptide of type I collagen (NTX). A prospective study was performed on 49 patients with bone metastases from breast, lung and prostate cancer, who were undergoing treatment with zoledronic acid. Patients were monitored for 1 year with blood tests, clinical evaluation and instrumental exams according to the response evaluation criteria of the University of Texas M. D. Anderson Cancer Center (Houston, TX, USA) and the Positron Emission Tomography Response Criteria in Solid Tumors. Circulating RANK/RANKL/OPG transcripts and NTX levels were evaluated by reverse transcription-quantitative polymerase chain reaction and immune enzymatic assay, respectively. The baseline RANKL levels differed significantly between responders and non-responders, whereas no differences in NTX levels were observed between the two groups. Receiver operating characteristic curve evaluation for all markers revealed that RANKL was the most accurate marker, with an area under the curve of 0.74 (95% confidence interval, 0.54-0.93). In addition, RANKL, which is the target of the novel monoclonal antibody denosumab, was the most accurate predictor of bone response in the present series of patients with bone metastases. Thus, the use of RANKL as a marker could potentially improve clinical practice, as current bone response evaluation is still somewhat problematic.

Abstract 2733: Novel function of the RAI2 protein in genomic integrity of breast cancer cells

Abstract We have recently identified the RAI2 protein as putative metastasis-suppressor related to dedifferentiation, early occurring bone metastasis formation, and survival of hormone dependent breast carcinomas. Nevertheless, low RAI2 expression is also predictive for poor patient outcome in hormone independent tumors and is furthermore associated with mutation of p53 in primary breast tumors. Thus, besides sustaining luminal differentiation, RAI2 might possess a general function as tumor suppressor that is, as indicated by association with p53 status, possibly related to maintenance of genomic stability. In order to characterize its potential role in genomic integrity, we screened lysates from RAI2 depleted MCF-7 and KPL-1 breast cancer cell lines with phosphorylation-specific antibody arrays. Additionally, we analyzed changes in expression of cell cycle related genes and proteins by qPCR and Western Blot analysis. We also conducted cytogenetic assays to investigate whether RAI2 depletion affects chromosomal stability and mitotic progression. Finally, we evaluated a possible association of RAI2 expression with aneuploidy in published expression data sets from breast cancer patients. As revealed by antibody array analysis, RAI2 depletion causes activation of p53 protein in MCF-7 and KPL-1 breast cancer cells, which was accompanied by a significant increase of senescent cells in the corresponding cell populations. Furthermore RAI2 depletion causes downregulation of several key factors of G2/M transition, like Aurora kinases A and B as well as Cyclins A2, B1 and B2. Concomitantly, we found in RAI2-depleted cell cultures reduction of mitotic index, increase of unattached chromosomes in metaphase and lagging chromosomes and during anaphase. This finding was further approved by an increase of numerical aberrations in RAI2 knockdown cells. Finally, we could show an association between low RAI2 expression and higher level of chromosomal instability in primary tumors from breast cancer patients. In conclusion, we found that loss of RAI2 function is associated with decreased mitotic fidelity. Thus, besides sustaining differentiation in hormone dependent breast tumor, RAI2 also acts as general tumor suppressor that maintains genomic integrity. Reference: Werner S et al., Cancer Discovery. 2015 May;5(5):506-19 Citation Format: Stefan Werner, Kerstin Borgmann, Klaus Pantel, Harriet Wikman. Novel function of the RAI2 protein in genomic integrity of breast cancer cells. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2733.

The role of microRNAs in bone metastasis

The skeleton represents a common site of metastases for osteotropic cancers such as prostate and breast tumors and novel therapeutic targets and new markers for the monitoring of bone lesions are urgently needed. The formation of bone metastases is a complex process that starts at the level of the confined tumor and that is characterized by a dynamic crosstalk between the primary cancer and the future metastatic site, the bone. Factors released by the primary tumor contribute to prepare a fertile “soil”, where a “pre-metastatic niche” is established prior to future colonization by cancer cells. When the primary cancer progress from the confined disease to its invasive phase, tumor cells will acquire an invasive phenotype, enter into the circulation and colonize the previously prepared site where they will establish a “metastatic niche”. Among the variety of molecules that participate in the metastatic cascade, microRNAs are a class of small non-coding RNA that play an important role in the development of metastatic bone lesions. Many studies have addressed the role of small non-coding RNAs (miRs) in metastasis in osteotropic cancers and have highlighted the role of miRs as oncogenes (oncomiRs) or tumor suppressor miRs.In this review we present describe the role of miRs in the processing of the supportive bone microenvironment prior and after the bone colonization by cancer cells. Finally, future therapeutic strategies and perspectives are also discussed.

Lysyl oxidase promotes survival and outgrowth of colon cancer cells in the bone marrow, enabling bone metastasis formation

Lysyl oxidase promotes survival and outgrowth of colon cancer cells in the bone marrow, enabling bone metastasis formation

Can the microRNA expression profile help to identify novel targets for zoledronic acid in breast cancer?

Zoledronic acid (ZOL), belonging to third generation bisphosphonate family, is a potent inhibitor of osteoclast-mediated bone resorption, widely used to effectively prevent osteolysis in breast cancer patients who develop bone metastases. Low doses of ZOL have been shown to exhibit a direct anticancer role, by inhibiting cell adhesion, invasion, cytoskeleton remodelling and proliferation in MCF-7 breast cancer cells. In order to identify the molecular mechanisms and signaling pathways underlying the anticancer activity exerted by ZOL, we analyzed for the first time the microRNA expression profile in breast cancer cells. A large-scale microarray analysis of 377 miRNAs was performed on MCF7 cells treated with 10 μM ZOL for 24 h compared to untreated cells. Furthermore, the expression of specific ZOL-induced miRNAs was analyzed in MCF-7 and SkBr3 cells through Real-time PCR. Low-dose treatment with ZOL significantly altered expression of 54 miRNAs. Nine upregulated and twelve downregulated miRNAs have been identified after 24 h of treatment. Also, ZOL induced expression of 11 specific miRNAs and silenced expression of 22 miRNAs. MiRNA data analysis revealed the involvement of differentially expressed miRNAs in PI3K/Akt, MAPK, Wnt, TGF-β, Jak-STAT and mTOR signaling pathways, and regulation of actin cytoskeleton. Our results have been shown to be perfectly coherent with the recent findings reported in literature concerning changes in expression of some miRNAs involved in bone metastasis formation, progression, therapy resistance in breast cancer. In conclusion, this data supports the hypothesis that ZOL-induced modification of the miRNA expression profile contributes to the anticancer efficacy of this agent.

The role of osteoclasts in breast cancer bone metastasis

Breast cancer frequently metastasises to the skeleton, interfering with the normal bone remodelling process and inducing bone degradation. Bone degradation is caused by osteoclasts, the normal bone-resorbing cells. Osteoclast-mediated bone degradation subsequently leads to the release of bone-derived factors that promote skeletal tumour growth. Osteoclasts themselves stimulate tumour growth. This Review describes the molecular mechanisms through which osteoclasts and breast cancer cells collaborate with each other, triggering the formation of osteolytic bone metastasis.

Potential Involvement of Jagged1 in Metastatic Progression of Human Breast Carcinomas.

Jagged1, the ligand of Notch, has been shown to be involved in formation of bone metastases in an experimental study. Here, clinical relevance of Jagged1 expression in tumor progression was assessed in human breast carcinomas. Jagged1 expression was evaluated by immunohistochemistry in 228 tumor tissue samples and compared to clinicopathologic parameters and patients' outcomes. Furthermore, circulating tumor cells (CTCs) from peripheral blood of 100 unmatched metastatic cancer patients with progressive disease were enriched using Ficoll density gradient centrifugation and detected by pan-keratin/Jagged1/CD45 immunofluorescent staining. Jagged1 expression was detected in 50% of 228 tumors. Jagged1 expression was correlated with higher tumor grade (P = 0.047), vascular invasion (P = 0.026), luminal B subtype (P = 0.016), overexpression of Her-2 (P = 0.001), high Ki-67 expression (P = 0.035), and aldehyde dehydrogenase 1 (ALDH1) positivity (P = 0.013). Jagged 1 expression indicated shorter disease-free survival (DFS) (P = 0.040) and metastasis-free survival (P = 0.048) in lymph node-negative breast cancer for which it was the only independent predictor of DFS (multivariate analysis, P = 0.046). Tumors characterized by the strongest Jagged1 staining intensity (7.5% of cases) correlated with lymph node positivity (P = 0.037), metastatic relapse (P = 0.049), and higher number of disseminated tumor cells in bone marrow aspirates (P = 0.041). Twenty-one unmatched metastatic breast cancer patients with progressive disease were positive for CTCs, and 85.7% of the CTCs also expressed Jagged1. The presence of Jagged1(+) CTCs was significantly associated with shorter progression-free survival in patients treated with bisphosphonates (P = 0.013). Jagged1 expression characterizes more aggressive breast carcinoma and might be involved in tumor cell dissemination, metastatic progression, and resistance to bone-targeting therapy in breast cancer patients.

Dissecting Tumor-Stromal Interactions in Breast Cancer Bone Metastasis.

Bone metastasis is a frequent occurrence in breast cancer, affecting more than 70% of late stage cancer patients with severe complications such as fracture, bone pain, and hypercalcemia. The pathogenesis of osteolytic bone metastasis depends on cross-communications between tumor cells and various stromal cells residing in the bone microenvironment. Several growth factor signaling pathways, secreted micro RNAs (miRNAs) and exosomes are functional mediators of tumor-stromal interactions in bone metastasis. We developed a functional genomic approach to systemically identified molecular pathways utilized by breast cancer cells to engage the bone stroma in order to generate osteolytic bone metastasis. We showed that elevated expression of vascular cell adhesion molecule 1 (VCAM1) in disseminated breast tumor cells mediates the recruitment of pre-osteoclasts and promotes their differentiation to mature osteoclasts during the bone metastasis formation. Transforming growth factor β (TGF-β) is released from bone matrix upon bone destruction, and signals to breast cancer to further enhance their malignancy in developing bone metastasis. We furthered identified Jagged1 as a TGF-β target genes in tumor cells that engaged bone stromal cells through the activation of Notch signaling to provide a positive feedback to promote tumor growth and to activate osteoclast differentiation. Substantially change in miRNA expression was observed in osteoclasts during their differentiation and maturation, which can be exploited as circulating biomarkers of emerging bone metastasis and therapeutic targets for the treatment of bone metastasis. Further research in this direction may lead to improved diagnosis and treatment strategies for bone metastasis.

Proteomic Profiling of In-Vitro Bone-Conditioned Skbr3 Breast Cancer Cells

A major complication of advanced breast cancer is the frequent formation of bone metastases. To our knowledge, however, the information on the phenotypic properties of cancer cells able to survive and colonize the bone tissue is still fragmentary. We report here a proteomic study of breast cancer cells (SKBR3) collected after a colonization of bone fragments in a special model of co-culture, used to simulate the phenotype of breast cancer metastasizing cells. The scanning electron microscopy of the seeded cancer cells into the bone fragments, showed their peculiar traits of multilayered growth and spiny surface, typical of a very aggressive phenotype. The comparative proteomic profiling of the bone-conditioned cells (SKBR3-B1) versus the parental cell line (SKBR3-WT), performed by applying two-dimensional electrophoresis (2D-IPG) and mass spectrometry (Maldi / TOF) revealed the existence of an interesting differential proteomic profiling. Within this pattern, an increase of several members of the S100 protein family, not described before in relation to bone metastasis from breast cancer, was observed in the SKBR3-B1. Other up-regulated proteins belong to protein classes involved mainly in cytoskeleton dynamics, chaperone / folding, negative regulation of apoptosis and cell metabolism. These results suggest that, rather than individual proteins, a coordinated network of strategic proteins is able to determine or drive the invasive and metastatic phenotype of breast cancer cells, namely: proteins primarily strategic for the cell penetration and homing into the bone, and proteins coming into play to satisfy the modified metabolic needs of the cells. We believe that the present study represents an innovative approach of co-culture with fresh human bone fragments, and that it may contribute greatly to improve the knowledge of the molecular mechanisms underlying the propensity of breast cancer cells to colonize, survive and proliferate into the bone microenvironment.

Functional screen identifies kinases driving prostate cancer visceral and bone metastasis.

Mutationally activated kinases play an important role in the progression and metastasis of many cancers. Despite numerous oncogenic alterations implicated in metastatic prostate cancer, mutations of kinases are rare. Several lines of evidence suggest that nonmutated kinases and their pathways are involved in prostate cancer progression, but few kinases have been mechanistically linked to metastasis. Using a mass spectrometry-based phosphoproteomics dataset in concert with gene expression analysis, we selected over 100 kinases potentially implicated in human metastatic prostate cancer for functional evaluation. A primary in vivo screen based on overexpression of candidate kinases in murine prostate cells identified 20 wild-type kinases that promote metastasis. We queried these 20 kinases in a secondary in vivo screen using human prostate cells. Strikingly, all three RAF family members, MERTK, and NTRK2 drove the formation of bone and visceral metastasis confirmed by positron-emission tomography combined with computed tomography imaging and histology. Immunohistochemistry of tissue microarrays indicated that these kinases are highly expressed in human metastatic castration-resistant prostate cancer tissues. Our functional studies reveal the strong capability of select wild-type protein kinases to drive critical steps of the metastatic cascade, and implicate these kinases in possible therapeutic intervention.

EpCAM expression in breast cancer cells is associated with enhanced bone metastasis formation.

Epithelial cell adhesion molecule (EpCAM) has been implicated in multiple cellular functions including cell adhesion. EpCAM has also recently been identified as a marker for cancer stem cells (CSCs). Here, we examined the roles of EpCAM in the development of bone metastasis of breast cancer by using well-characterized animal models. Morphological and real-time reverse transcriptase-polymerase chain reaction data showed that the EpCAM-negative and -positive (EpCAM(neg) and EpCAM(pos) ) cell populations isolated from breast cancer cell lines exhibited mesenchymal and epithelial phenotypes, respectively. Flow cytometric analysis revealed that EpCAM(pos) , but not EpCAM(neg) , cells possessed self-renewal and differentiation potentials. Tumorsphere formation in suspension cultures and tumorigenicity in the orthotopic mammary fat pad of mice were significantly greater in EpCAM(pos) cells than in EpCAM(neg) cells. The development of bone metastases induced by an intracardiac injection was markedly increased in mice inoculated with EpCAM(pos) cells. Furthermore, intracardiac inoculations of parental cells demonstrated that the EpCAM(pos) population in cancer cells that colonized in bone was significantly higher than that in parental cells. However, stable transduction of EpCAM into EpCAM(neg) cells failed to reproduce the phenotypes of EpCAM(pos) cells. These results suggest that the expression of EpCAM in breast cancer cells is associated with CSC-like phenotypes, which contribute to the promotion of bone metastases by enhancing tumorigenicity. Our results also support the possibility that the epithelial phenotypes of EpCAM-expressing cells confer advantageous properties for the development of bone metastases, at least after entering the circulation, while EpCAM is likely not solely responsible for the phenotypes of EpCAM(pos) cells.

PKCε Is an Essential Mediator of Prostate Cancer Bone Metastasis.

The bone is a preferred site for metastatic homing of prostate cancer cells. Once prostate cancer patients develop skeletal metastases, they eventually succumb to the disease; therefore, it is imperative to identify key molecular drivers of this process. This study examines the involvement of protein kinase C epsilon (PKCε), an oncogenic protein that is abnormally overexpressed in human tumor specimens and cell lines, on prostate cancer cell bone metastasis. PC3-ML cells, a highly invasive prostate cancer PC3 derivative with bone metastatic colonization properties, failed to induce skeletal metastatic foci upon inoculation into nude mice when PKCε expression was silenced using shRNA. Interestingly, while PKCε depletion had only marginal effects on the proliferative, adhesive, and migratory capacities of PC3-ML cells in vitro or in the growth of xenografts upon s.c. inoculation, it caused a significant reduction in cell invasiveness. Notably, PKCε was required for transendothelial cell migration (TEM) as well as for the growth of PC3-ML cells in a bone biomimetic environment. At a mechanistic level, PKCε depletion abrogates the expression of IL1β, a cytokine implicated in skeletal metastasis. Taken together, PKCε is a key factor for driving the formation of bone metastasis by prostate cancer cells and is a potential therapeutic target for advanced stages of the disease. This study uncovers an important new function of PKCε in the dissemination of cancer cells to the bone; thus, highlighting the promising potential of this oncogenic kinase as a therapeutic target for skeletal metastasis.

Estrogen-deficient osteoporosis enhances the recruitment and activity of osteoclasts by breast cancer cells.

To reduce the burden of bone metastases, the pathophysiology of the metastatic niche should be elucidated and targeted. The aim of the present study was to assess the effect of tumor cells on osteoclast (OC) recruitment and activity in the presence of altered bone remodelling. Peripheral blood mononuclear cells (PBMC) were isolated from healthy and ovariectomized (OVX) rats and co-cultured with MRMT-1 rat breast carcinoma cells or with their conditioned medium for 1 and 2 weeks. Alamar Blue viability test, synthesis of cathepsin K, transforming growth factor-beta 1 (TGF-β1), tumor necrosis factor alpha (TNF-α), vascular endothelial growth factor (VEGF), metalloproteinase (MMP)-7, MMP-9, FITC-conjugate phalloidin staining and tartrate-resistant acid phosphatase (TRAP) staining were evaluated. The results indicate that breast carcinoma cells induced different responses in PBMC derived from rats affected by estrogen deficiency osteoporosis (OP) in comparison with healthy ones, with a significant increase in proliferation rate, OC differentiation, synthesis of TNF-α, MMP-7 and MMP-9. The data support the "proof of concept" that OP due to estrogen deficiency might offer a receptive site for cancer cells to form bone metastases.

Abstract 47: KRAS-mutation dependent effect of zoledronic acid in human NSCLC preclinical models

Abstract In non-small cell lung cancer (NSCLC) KRAS-mutant status is a negative prognostic and predictive factor of both classic and target based therapy. In human malignancies bisphosphonate drugs are applied to treat or prevent formation of bone metastases. Nitrogen-containing bisphosphonates inhibit the posttranslational modification of small G-proteins (e.g. Ras, Rac, Rho) and thus may affect proliferation and migration. In our preclinical work, we investigated the effect of an aminobisphosphonate compound (zoledronic acid) on mutant and wild type KRAS-expressing human NSCLC cell lines. In vitro cell viability was measured by MTT assay in H358, LCLC-103H, H1650 and H1975 cells. Apoptotic cell death was quantified by flow cytometric detection of sub-G1-fraction in propidium-iodide stained cultures. Furthermore, we investigated the in vivo effect of zoledronic acid in a SCID mouse subcutaneous xenograft model. The extracted tumors were examined by traditional histological methods and the proportion of proliferating tumor cells was determined by Ki67 labeling. KRAS-mutant H358 human NSCLC cell line showed relative resistance to zoledronic acid on proliferation, while human lung cancer cell lines carried wild type KRAS proved to be sensitive. Zoledronic acid improved the cytostatic effect of traditional anti-cancer agent cisplatin. Zoledronic acid did not induce significant apoptosis in NSCLC cell lines in vitro, rather the blockade of the cell cycle appeared. In in vivo subcutaneous animal model, zoledronic acid significantly reduced the weight of LCLC-103H xenograft tumor harboring wild type KRAS, furthermore, the treatment decreased the number of Ki67-labeled tumor. Zoledronic acid induced the expression of VEGF in LCLC-103H cells and stimulated in vivo angiogenesis. We concluded that in human NSCLC cells the antiproliferative effect of zoledronic acid dependended on KRAS-status (wild-type or mutant). The inhibitory effect of zoledronic manifested through the in vitro and in vivo blockade of cell cycle. The in vivo angiogenic effect of zoledronic acid supported cytostatic effect of cisplatin. Our preclinical investigation suggests that patients with wild type KRAS-expressing NSCLC could potentially benefit from aminobisphosphonate therapy. Citation Format: István Kenessey, Krisztina Kói, Mihály Cserepes, Judit Dobos, Balázs Hegedűs, József Tóvári, József Tímár. KRAS-mutation dependent effect of zoledronic acid in human NSCLC preclinical models. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 47. doi:10.1158/1538-7445.AM2015-47

The hypoxic cancer secretome induces pre-metastatic bone lesions through lysyl oxidase.

Tumour metastasis is a complex process involving reciprocal interplay between cancer cells and host stroma at both primary and secondary sites, and is strongly influenced by microenvironmental factors such as hypoxia. Tumour-secreted proteins play a crucial role in these interactions and present strategic therapeutic potential. Metastasis of breast cancer to the bone affects approximately 85% of patients with advanced disease and renders them largely untreatable. Specifically, osteolytic bone lesions, where bone is destroyed, lead to debilitating skeletal complications and increased patient morbidity and mortality. The molecular interactions governing the early events of osteolytic lesion formation are currently unclear. Here we show hypoxia to be specifically associated with bone relapse in patients with oestrogen-receptor negative breast cancer. Global quantitative analysis of the hypoxic secretome identified lysyl oxidase (LOX) as significantly associated with bone-tropism and relapse. High expression of LOX in primary breast tumours or systemic delivery of LOX leads to osteolytic lesion formation whereas silencing or inhibition of LOX activity abrogates tumour-driven osteolytic lesion formation. We identify LOX as a novel regulator of NFATc1-driven osteoclastogenesis, independent of RANK ligand, which disrupts normal bone homeostasis leading to the formation of focal pre-metastatic lesions. We show that these lesions subsequently provide a platform for circulating tumour cells to colonize and form bone metastases. Our study identifies a novel mechanism of regulation of bone homeostasis and metastasis, opening up opportunities for novel therapeutic intervention with important clinical implications.

Tumour-derived miRNAs and bone metastasis.

Skeletal metastases are complications of epithelial cancers, among which breast, prostate and lung carcinomas are the most osteotropic. In primary tumours, a subset of cancer cells undergoes epithelial-mesenchymal transition, acquires mobility to migrate into the surrounding stroma and seeds at distant sites to grow. The specific development of bone metastasis requires the recruitment of circulating tumour cells in the bone marrow, their adaptation to survive in the surrounding microenvironment where they alter the functions of osteoclasts and osteoblasts, and hijack signals coming from the bone matrix. Each of the molecular pathways underlining these steps is regulated by multiple factors, through the tight control of genes expressed by cancer cells interacting with cells from the bone microenvironment. In this context, miRNAs can act as master regulators of gene expression to control multiple aspects of bone metastasis formation, including cancer cell escape from the primary tumour site, cancer cell dissemination to bone and invasion of the bone marrow, as well as secondary outgrowth and tumour-stroma cell interactions. In the clinic, specific miRNA signatures have been identified in osteotropic cancer cells, raising the possibility that miRNAs could be used as biomarkers of bone metastasis. The regulatory activity of miRNAs in the bone microenvironment also suggests that miRNAs could be promising therapeutic targets.

Animal and cellular models of hepatocellular carcinoma bone metastasis: establishment and characterisation.

An increasingly high occurrence of bone metastases in hepatocellular carcinoma (HCC) patients highlights the importance of fundamental research on HCC bone metastasis, which has been limited in its success due to the lack of a model system. Establishment of animal and cellular models of HCC bone metastasis and discovery of HCC bone metastasis-related genes. Luciferase-transfected HCC cell lines HCCLM3, MHCC97H, and SMMC-7721 were used to inoculate nude mice intracardially. Formation of bone metastases was examined by bioluminescence imaging, SPECT, and pathology study. Metastatic cells in bone were isolated and subcultured. Differences between bone metastatic cells and their parental cells were studied by in vitro/in vivo assays. Mouse model of HCC bone metastasis was successfully established. Injected tumour cells formed metastases in the skull, the spine, the hind limbs, and the sternum, causing osteolytic lesions via act of MMP-1 and recruitment of osteoclasts. Four bone metastatic cell lines were extracted from HCCLM3-inoculated mice and were demonstrated to exhibit a much stronger ability to form bone metastases as well as other phenotypes, including enhanced in vitro migration/invasion and colony formation. Moreover, the expression of PTHrP, MMP-1, and CTGF was significantly elevated in bone metastatic cells compared to parental HCC cells. The nude mouse model and bone metastatic cell lines together provide an effective simulation of HCC bone metastasis. This model system will become powerful tool with which to explore the mechanisms and therapies of HCC bone metastasis. Additionally, PTHrP, MMP-1, and CTGF are candidate genes related to HCC bone metastasis.

OPG-Fc inhibits ovariectomy-induced growth of disseminated breast cancer cells in bone.

Dormant disseminated tumour cells can be detected in the bone marrow of breast cancer patients several years after resection of the primary tumour. The majority of these patients will remain asymptomatic, however, ∼ 15% will go on to develop overt bone metastases and this condition is currently incurable. The reason why these dormant cells are stimulated to proliferate and form bone tumours in some patients and not others remains to be elucidated. We have recently shown that in an in vivo model, increasing bone turnover by ovariectomy stimulated proliferation of disseminated tumour cells, resulting in formation of bone metastasis. We now show for the first time that osteoclast mediated mechanisms induce growth of tumours from dormant MDA-MB-231 cells disseminated in the bone. We also show that disruption of RANK-RANKL interactions following administration of OPG-Fc inhibits growth of these dormant tumour cells in vivo. Our data support early intervention with anti-resorptive therapy in a low-oestrogen environment to prevent development of bone metastases.

The Impact of Immune System in Regulating Bone Metastasis Formation by Osteotropic Tumors.

Bone metastases are frequent and debilitating consequence for many tumors, such as breast, lung, prostate, and kidney cancer. Many studies report the importance of the immune system in the pathogenesis of bone metastasis. Indeed, bone and immune system are strictly linked to each other because bone regulates the hematopoietic stem cells from which all cells of the immune system derive, and many immunoregulatory cytokines influence the fate of bone cells. Furthermore, both cytokines and factors produced by immune and bone cells promote the growth of tumor cells in bone, contributing to supporting the vicious cycle of bone metastasis. This review summarizes the current knowledge on the interactions among bone, immune, and tumor cells aiming to provide an overview of the osteoimmunology field in bone metastasis from solid tumors.

Knockdown of TRAIL-R2 in breast cancer cells impairs bone metastases formation in a preclinical mouse model

Knockdown of TRAIL-R2 in breast cancer cells impairs bone metastases formation in a preclinical mouse model