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Introduction: Bone metastases are the most frequent site of secondary localization of prostate cancer (PCa) and are present in about 90% of cases of advanced disease. Consequently, an adequate management of bone involvement is of pivotal importance in the therapeutic approach and skeletal-related events (SREs) need to be closely monitored and promptly assessed and treated. Bone targeting agents (BTAs), consisting in bisphosphonates and denosumab, are an essential part of the treatment of metastatic prostate cancer that accompanies systemic treatments throughout the most part of the history of the disease. Activity and safety of bone targeting agents: These treatments are correlated to better outcomes in terms of reduction of SREs and, in metastatic castration resistant setting, of increased overall survival (OS), but several important adverse events have to be managed and prevented. Of these, osteonecrosis of the jaw (ONJ) is extremely invalidating and should be managed with a special attention. Discussion: The role of BTAs in prostate cancer is pivotal throughout many stages of the disease, but several toxicities should be quickly recognized and treated. We aim at recollecting evidence on clinical benefit of BTAs, common and specific toxicities, and explore the pathophysiology and clinical aspects of osteonecrosis of the jaw. We present a review of the literature to report the role of the different types of bone targeting agents in the management of prostate cancer with bone metastases with a particular focus on common toxicities and ONJ to recollect current evidences on the activity of these compounds and the correct management of their adverse events.

The clinical treatment of bone metastasis from breast cancer is currently based on the systemic administration of antiresorptive agents, radiopharmaceuticals, and/or local treatments such as radiation therapy, radiofrequency ablation and surgery. However, these therapeutic options are merely palliative and do not show to have a significant positive impact on patients' survival. In addition, the systemic administration of antiresorptive drugs and/or antitumour agents and/or radiopharmaceuticals may negatively affect normal bone metabolism with detrimental consequences for cancer patients. Hence, the need to identify alternative therapeutic strategies that, based on the hallmarks of bone metastasis, can effectively thwart tumour cell growth while, at the same time, overcoming antitumour drug-induced bone loss and preserving bone health. To this aim, current studies are directed toward the development of new molecules with low toxicity and/or novel drug delivery systems, which may efficiently hinder the growth of metastatic cells, while other studies are focussed on the design of new clinical treatments that, by acting on early stages of breast cancer, may prevent thehoming and the subsequent growth of cancercells inthebone. These treatments might also target occult micro-metastases before their development into clinically evident bone lesions, ultimately hindering bone relapse and disease progression. However, this therapeutic approach implies the identification of early stage breast cancer patients being at high risk of developing bone metastases. The discovery of novel specific biomarkers as predictor of metastatic bone disease may help clinicians in selecting these patients.

Simple Summary Over the disease course of metastatic prostate cancer, approximately the 90% of patients develops bone metastases, with bone involvement frequently leading to various skeletal complications including pathological fractures, spinal cord compression, and pain. Notably enough, the peculiar inclination of prostate cancer cells to seed the bone is considered an important cause of morbidity for prostate cancer patients. Recent years have witnessed the advent of several novel treatments for prostate cancer, and therapeutic paradigms are rapidly shifting. In this review, we aim at giving an overview of current knowledge on the relationship between prostate cancer and bone, especially focusing on the use of bone-targeted agents in this setting. Abstract Bone health represents a major issue in castration-resistant prostate cancer (CRPC) patients with bone metastases; in fact, the frequently prolonged use of hormonal agents causes important modifications in physiological bone turnover and most of these men will develop skeletal-related events (SREs), including spinal cord compression, pathologic fractures and need for surgery or radiation to bone, which are estimated to occur in almost half of this patient population. In the last decade, several novel therapeutic options have entered into clinical practice of bone metastatic CRPC, with recent approval of enzalutamide and abiraterone acetate, cabazitaxel chemotherapy and radium-223, on the basis of survival benefit suggested by landmark Phase III trials assessing these agents in this setting. Conversely, although bone-targeted agents (BTAs)—such as the bisphosphonate zoledronic acid and the receptor activator of nuclear factor kappa-B (RANK) ligand inhibitor denosumab—are approved for the prevention of SREs, these compounds have not shown benefit in terms of overall survival. However, emerging evidence has suggested that the combination of BTAs and abiraterone acetate, enzalutamide and the radiopharmaceutical radium-223 could result in improved clinical outcomes and prolonged survival in bone metastatic CRPC. In this review, we will provide an overview on bone tropism of prostate cancer and on the role of BTAs in metastatic hormone-sensitive and castration-resistant prostate cancer.

Bone is one of the most preferred sites of metastatic spread from different cancer types, including breast cancer. However, different breast cancer subtypes exhibit distinct metastatic behavior in terms of kinetics and anatomic sites of relapse. Despite advances in the diagnosis, the identification of patients at high-risk of bone recurrence is still an unmet clinical need. We conducted a retrospective analysis, by gene expression and immunohistochemical assays, on 90 surgically resected breast cancer samples collected from patients who experienced no evidence of distant metastasis, bone or visceral metastasis in order to identify a primary tumor-derived marker of bone recurrence. We identified trefoil factor-1 (pS2 or TFF1) as strictly correlated to bone metastasis from ER+ breast cancer. In silico analysis was carried out to confirm this observation, linking gene expression data with clinical characteristics available from public clinical datasets. Then, we investigated TFF1 function in ER+ breast cancer tumorigenesis and bone metastasis through xenograft in vivo models of MCF 7 breast cancer with gain and loss of function of TFF1. As a response to microenvironmental features in primary tumors, TFF1 expression could modulate ER+ breast cancer growth, leading to a less proliferative phenotype. Our results showed it may not play a role in late stages of bone metastasis, however further studies are warranted to understand whether it could contribute in the early-stages of the metastatic cascade. In conclusion, TFF1 upregulation in primary ER+ breast cancer could be useful to identify patients at high-risk of bone metastasis. This could help clinicians in the identification of patients who likely can develop bone metastasis and who could benefit from personalized treatments and follow-up strategies to prevent metastatic disease.

Bone metastasis (BM) is a dismal complication of cancer that frequently occurs in patients with advanced carcinomas and that often manifests as an osteolytic lesion. In bone, tumor cells promote an imbalance in bone remodeling via the release of growth factors that, directly or indirectly, stimulate osteoclast resorption activity. However, carcinoma cells are also characterized by an altered metabolism responsible for a decrease of extracellular pH, which, in turn, directly intensifies osteoclast bone erosion. Here, we speculated that tumor-derived acidosis causes the osteoblast–osteoclast uncoupling in BM by modulating the pro-osteoclastogenic phenotype of osteoblasts. According to our results, a low pH recruits osteoclast precursors and promotes their differentiation through the secretome of acid-stressed osteoblasts that includes pro-osteoclastogenic factors and inflammatory mediators, such as RANKL, M-CSF, TNF, IL-6, and, above the others, IL-8. The treatment with the anti-IL-6R antibody tocilizumab or with an anti-IL-8 antibody reverted this effect. Finally, in a series of BM patients, circulating levels of the osteolytic marker TRACP5b significantly correlated with IL-8. Our findings brought out that tumor-derived acidosis promotes excessive osteolysis at least in part by inducing an inflammatory phenotype in osteoblasts, and these results strengthen the use of anti-IL-6 or anti-IL-8 strategies to treat osteolysis in BM.

Background: Renal cell carcinoma (RCC) constitutes 3% of all cancers, with a higher incidence in patients with age between 60 and 70 years. RCC frequently present as a metastatic tumor at diagnosis, and bones represent one of the most frequent sites. Many cases, mainly in young patients, includes the Xp11 translocation RCC. The cytological diagnosis of Xp11 translocation RCC in adult population it is rarely performed, likely for the morphological overlap with other adult renal cell carcinoma subtypes. Methods: We retrospectively analyze a series of 92 adult patients with metastatic bone tumors, diagnosed on fine-needle aspiration cytology (FNAC) samples, focusing mainly on the cytological, immunophenotypic and molecular features of Xp11 translocation RCC. Results: In our series 6 of 92 (6.5%) cases were metastatic RCC (mRCC), among them 2 cases were metastasis from Xp11translocation RCC. Those cases showed a bloody background, with several groups of atypical cells arranged in syncytial groups or in papillary groups composed by atypical cells with abundant cytoplasm, with scattered clear cells. TFE3 was positive on immunocytochemical analysis and specific translocation t(Xp11.23) was detected by FISH analysis. Conclusions: In adult patients with mRCC, it is necessary to consider also Xp11 translocation RCC among the diagnostic hypotheses. FNAC represents a valid tool to investigate bone lesions but cytological features of Xp11 translocation RCC are still poorly described and must necessarily be better defined.

Bone is one of the most common sites for cancer metastasis. Bone tissue is composed by different kinds of cells that coexist in a coordinated balance. Due to the complexity of bone, it is impossible to capture the intricate interactions between cells under either physiological or pathological conditions. Hence, a variety of in vivo and in vitro approaches have been developed. Various models of tumor–bone diseases are routinely used to provide valuable information on the relationship between metastatic cancer cells and the bone tissue. Ideally, when modeling the metastasis of human cancers to bone, models would replicate the intra-tumor heterogeneity, as well as the genetic and phenotypic changes that occur with human cancers; such models would be scalable and reproducible to allow high-throughput investigation. Despite the continuous progress, there is still a lack of solid, amenable, and affordable models that are able to fully recapitulate the biological processes happening in vivo, permitting a correct interpretation of results. In the last decades, researchers have demonstrated that three-dimensional (3D) methods could be an innovative approach that lies between bi-dimensional (2D) models and animal models. Scientific evidence supports that the tumor microenvironment can be better reproduced in a 3D system than a 2D cell culture, and the 3D systems can be scaled up for drug screening in the same way as the 2D systems thanks to the current technologies developed. However, 3D models cannot completely recapitulate the inter- and intra-tumor heterogeneity found in patients. In contrast, ex vivo cultures of fragments of bone preserve key cell–cell and cell–matrix interactions and allow the study of bone cells in their natural 3D environment. Moreover, ex vivo bone organ cultures could be a better model to resemble the human pathogenic metastasis condition and useful tools to predict in vivo response to therapies. The aim of our review is to provide an overview of the current trends in bone metastasis modeling. By showing the existing in vitro and ex vivo systems, we aspire to contribute to broaden the knowledge on bone metastasis models and make these tools more appealing for further translational studies.

Bone is the third most frequent site of metastasis, with a particular incidence in breast and prostate cancer patients. For example, almost 70% of breast cancer patients develop several bone metastases in the late stage of the disease. Bone metastases are a challenge for clinicians and a burden for patients because they frequently cause pain and can lead to fractures. Unfortunately, current therapeutic options are in most cases only palliative and, although not curative, surgery remains the gold standard for bone metastasis treatment. Surgical intervention mostly provides the replacement of the affected bone with a bioimplant, which can be made by materials of different origins and designed through several techniques that have evolved throughout the years simultaneously with clinical needs. Several scientists and clinicians have worked to develop biomaterials with potentially successful biological and mechanical features, however, only a few of them have actually reached the scope. In this review, we extensively analyze currently available biomaterials-based strategies focusing on the newest and most innovative ideas while aiming to highlight what should be considered both a reliable choice for orthopedic surgeons and a future definitive and curative option for bone metastasis and cancer patients.

Bone primary tumors, such as osteosarcoma, are highly aggressive pediatric tumors that in 30% of the cases develop lung metastasis and are characterized by poor prognosis. Bone is also the third most common metastatic site in patients with advanced cancer and once tumor cells become homed to the skeleton, the disease is usually considered incurable, and treatment is only palliative. Bone sarcoma and bone metastasis share the same tissue microenvironment and niches. 3D cultures represent a new promising approach for the study of interactions between tumor cells and other cellular or acellular components of the tumor microenvironment (i.e., fibroblasts, mesenchymal stem cells, bone ECM). Indeed, 3D models can mimic physiological interactions that are crucial to modulate response to soluble paracrine factors, tumor drug resistance and aggressiveness and, in all, these innovative models might be able of bypassing the use of animal-based preclinical cancer models. To date, both static and dynamic 3D cell culture models have been shown to be particularly suited for screening of anticancer agents and might provide accurate information, translating in vitro cell cultures into precision medicine. In this mini-review, we will summarize the current state-of-the-art in the field of bone tumors, both primary and metastatic, illustrating the different methods and techniques employed to realize 3D cell culture systems and new results achieved in a field that paves the way toward personalized medicine.

Oleuropein (Ole) is the main bioactive phenolic compound present in olive leaves, fruits and olive oil. This molecule has been shown to exert beneficial effects on several human pathological conditions. In particular, recent preclinical and observational studies have provided evidence that Ole exhibits chemo-preventive effects on different types of human tumors. Studies undertaken to elucidate the specific mechanisms underlying these effects have shown that this molecule may thwart several key steps of malignant progression, including tumor cell proliferation, survival, angiogenesis, invasion and metastasis, by modulating the expression and activity of several growth factors, cytokines, adhesion molecules and enzymes involved in these processes. Interestingly, experimental observations have highlighted the fact that most of these signalling molecules also appear to be actively involved in the homing and growth of disseminating cancer cells in bones and, ultimately, in the development of metastatic bone diseases. These findings, and the experimental and clinical data reporting the preventive activity of Ole on various pathological conditions associated with a bone loss, are indicative of a potential therapeutic role of this molecule in the prevention and treatment of cancer-related bone diseases. This paper provides a current overview regarding the molecular mechanisms and the experimental findings underpinning a possible clinical role of Ole in the prevention and development of cancer-related bone diseases.

The skeleton is one of the most common sites of metastatic spread from advanced clear-cell renal carcinoma (ccRCC). Most of the bone lesions observed in RCC patients are classified as osteolytic, causing severe pain and morbidity due to pathological bone destruction. Nowadays, it is well known that cancer induced bone loss in lytic metastasis is caused by the triggering of a vicious cycle between cancer and bone resident cells that leads to an imbalance between bone formation and degradation. Targeting the mammalian target of rapamycin (mTOR) is an efficient treatment option for metastatic renal carcinoma patients. Moreover, bone targeted therapy could benefit bone metastatic cancer patients caused by advanced RCC. However, more data is needed to support the hypothesis of the beneficial effect of a combined therapy. The aim of this work is to investigate the effect of targeting mTOR and the sequential combination with bone targeted therapy as a strategy to break the vicious cycle between ccRCC cells and osteoclasts. A previously optimized fully human co-culture model is used to mimic the crosstalk between Caki-2 cells (ccRCC) and osteoclasts. Cells are treated at fixed timing with everolimus, zoledronic acid and denosumab as single or sequential combined treatment. We show that Caki-2 cells can induce osteoclast cells differentiation from isolated human monocytes, as demonstrated by specific tartrate-resistant acid phosphatase (TRAP) staining and f-actin ring formation, in a statistically significant manner. Moreover, differentiated osteoclasts proved to be functionally active by pit formation assay. Caki-2 cells co-cultured with osteoclasts acquire a more aggressive phenotype based on gene expression analysis. Interestingly, the sequential combined treatment of everolimus and zoledronic acid is the most effective in the inhibition of both Caki-2 cells survival and osteoclastogenic potential, making it an effective strategy to inhibit the vicious cycle of bone metastasis. At preclinical level, this observation confirms the value of our co-culture model as a useful tool to mimic the bone microenvironment and to assess drug sensitivity in vitro. A better understanding of the molecular mechanisms involved in tumor-bone cells crosstalk will be investigated next. Keywords: Bone metastasis, Renal carcinoma, Osteoclasts, Vicious cycle, Co-culture, Targeted therapy

Interventional radiology provides local management of bone metastases (BM) with a palliative intent in most cases, or with a curative intent in selected patients. Its role has rapidly expanded in the last decade, offering new treatment solutions often in combination with surgery, radiation therapy and medical treatments. The aim of the present paper is to increase awareness, acceptance and adoption of interventional radiology procedures for the treatment of BM; and to present the joint position of the Italian College of Musculoskeletal Radiology and the Italian College of Interventional Radiology.

Pembrolizumab is an immune checkpoint inhibitor approved for the treatment of patients with unresectable or metastatic melanoma. Appearance of bone metastases, either osteolytic or osteoblastic, during treatment qualifies as disease progression. We report the case of a 64-year-old White woman with a metastatic melanoma undergoing second-line treatment with pembrolizumab. At first evaluation, after 3 months of therapy, computed tomography scans showed the onset of osteosclerotic lesions and a significant reduction in all the previously identified metastases; on the contrary, a fluorine-18-fluorodeoxyglucose PET showed the normalization of fluorine-18-fluorodeoxyglucose uptake in all the baseline lesions, including bone metastases. Osteoblastic response, consisting of occurrence of new osteoblastic lesions on computed tomography imaging, as a consequence of an osteoblastic reaction of previously undetectable bone metastases, has been reported in some cancers that receive treatments such as chemotherapy, hormonal or targeted therapy. However, it had never been reported in patients with melanoma treated with immunotherapy. An apparent worsening of bone imaging on standard computed tomography scan in patients under checkpoint inhibitor should not lead to modification of treatment strategy, because misinterpretation as disease progression may lead to the premature cessation of a beneficial treatment and finally have a negative effect on patients' clinical outcome.

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Giandomenico Roviello,1,2 Silvia Paola Corona,3 Alberto Bonetta,4 Maria Rosa Cappelletti,5 Daniele Generali2,5 1Medical Oncology Unit, Department of Oncology, San Donato Hospital, Arezzo, 2Department of Medical, Surgery and Health Sciences, University of Trieste, Trieste, Italy; 3Radiation Oncology Department, Peter MacCallum Cancer Centre, Moorabbin Campus, East Bentleigh, VIC, Australia; 4Radiotherapy Department, 5Breast Cancer Unit, Azienda Socio-Sanitaria Territoriale di Cremona, Cremona, Italy Abstract: The aim of this study was to evaluate the correlation between circulating tumor cells (CTCs) and patterns of recurrence in patients with hormone-sensitive prostate cancer. The study involved patients with histologically confirmed, advanced prostatic adenocarcinoma, who were tested for CTCs (Veridex®) when they developed recurrence after radical prostatectomy or external beam radiation between 2008 and 2014. Forty-two prostate cancer patients were evaluated. CTCs were detected in 14 out of 42 (33.3%) patients (Group A), while the remaining 28 (66.7%) showed undetectable levels of CTCs (Group B). The mean prostate-specific antigen value was higher in Group A in comparison to Group B (6.2 vs 3.3 ng/dL) (P=0.48). Presence of bone metastases alone or along with nodal metastases was more common in GroupA (57.1%) in comparison to Group B (25%) (P=0.04). In a univariate analysis, the presence of CTCs at diagnosis correlated with the development of bone recurrence (OR: 4; 95% CI: 1.0–15.9; P=0.05). Even if the study enrolled only a small number of patients, the detection of CTCs in the blood appears to correlate with the pattern of progression in patients with hormone-sensitive prostate cancer, suggesting a possible role in anticipating recurrence at the bone in men with higher tumor load. Further prospective studies are warranted in this setting. Keywords: prostate cancer, CTCs, bone metastasis, biochemical failure

The purpose of this article is to outline the current approach to patients affected by metastasis to the long bones and to present a clinical and surgical algorithm available for clinicians and for future research. A modern approach to patients affected by long bone metastasis in fact requires a multidisciplinary contest where oncologists, radiotherapists, surgeons and physical therapists cooperate with a shared vision, in order to provide the best possible integrated treatments available. The authors of this article constitute the Bone Metastasis Study Group of the Italian Orthopaedic Society (SIOT): a national group of orthopedic tumor surgeons who are dedicated to studying the approach, techniques and outcomes of surgery for metastatic tumours of the musculoskeletal system.

Introduction. The problem of the diagnosis and treatment of the patients with cancer, in particular, with localization of it in the stomach, is one of the most pressing problems of today, both in Ukraine and throughout the world. The incidence of cancer is traditionally high in many countries, with males being twice as high as women. Annually in the world about 750 thousand new cases of gastric cancer are registered. Ukraine ranks eighth among the 48 countries in the world, which account the oncological diseases, by the indices of cancer. In the structure of mortality from malignant tumors, stomach cancer takes second place as in men (after malignant neoplasms of the lungs) and women (after breast cancer). In 2006, the incidence of stomach cancer in Ukraine was 27.4 per 100 thousand people, which significantly exceeds the world population – 15.6 per 100 thousand population. Aim. To make an overview of modern literature on the complications of gastric cancer in the practice of a doctor-rheumatologist and describe a clinical case. Materials and methods. The content analysis, the method of system and comparative analysis, the bibliosemantic method of study of the actual scientific researches concerning complications of gastric cancer in the practice of the doctor-rheumatologist, and the clinical case have been described. Results. It is presented the clinical case of arthralgic syndrome caused by the remote bone stomach cancer metastases in the practice of the doctor-rheumatologist. Diagnostic algorithm for the patient’s examination, directed by a family doctor with a suspicion of reactive arthritis, in the conditions of a rheumatologic hospital presupposes the use of a number of laboratory tests, most of which are performed in private laboratories. These tests were performed, signs of rheumatologic disease were not detected, but an increase of alkaline phosphatase level in serum by almost 12 times became the reason for reversing the search for the cause of bone tissue damage. X-ray examination of bone: focal changes has not been detected, chest x-ray has suspected metastasis in the lungs. Scintigraphic studies have revealed multiple metastatic bone defects. The ways of optimizing of the diagnosis search in patients with bone pain, elevated levels of alkaline phosphatase and burdened oncological history are outlined. It is noted that the optimal and most rational method of diarrhea in these situations is to study the level of alkaline phosphatase in the serum of the patient and make the scintigraphy with 99mTc-MDP. Conclusions. Taking into account the practical rheumatology, the article focuses on the optimal diagnosis of the patients experiencing pain in the bones, having the elevated levels of alkaline phosphatase, and the patients burdened with a cancer anamnesis. The studies on the problem show that the most effective and rational diagnostic method of gastric cancer is 99mTc-MDP scintigraphy.

Introduction Metastases to the proximal femur are usually managed surgically by tumor resection and reconstruction with an endoprosthesis, or by fixation with osteosynthesis. Still controversy remains regarding the most appropriate surgical treatment. We posed the following questions: (1) Is the frequency of surgical revision greater in patients treated with internal fixation than endoprosthetic reconstruction, and (2) Do complications that do not require surgery occur more frequently in patients treated with internal fixation rather than in those with endoprosthetic reconstruction? Materials and Methods A systematic review was performed of those studies reporting on surgical revision and complication rates comparing the two surgical methods. Ten studies including 1107 patients met the inclusion criteria, three with high methodological quality, three intermediate, and four with lowquality, according to the STROBE guidelines. Results At present, prosthetic dislocation is the most common complication observed in patients managed by prosthesis replacement of the proximal femur, while loosening was the main cause of reoperation in the fixation group. Time to reintervention ranged from 3 to 11.6 months for the prosthetic replacement and from 7.8 to 22.3 months for the fixation group. Non surgical complications, (mainly dislocations and infections) were more commonly observed in patients operated on by prosthetic replacement. Conclusions Implant related complications and surgery-related morbidity should be taken into account in the decision-making process for the surgical management of these patients. These data can improve the surgeon-patient communication and guide further studies on patients’ survival and complications with respect to surgery.

This study was directed to assess the clinical impact of the circulating cathepsin L, cystatin C, activin A, and follistatin in breast cancer patients. The serum concentrations of these molecules were determined by immunoenzymatic assays, and their association with some clinico-pathological parameters of breast cancer progression was evaluated. Our results identified cystatin C and activin A as predictive markers for the presence of breast cancer and bone metastasis, respectively. Therefore, these proteins may have a clinical role as circulating biomarkers in the diagnosis and therapeutic monitoring of breast cancer patients.

Advanced breast cancer frequently metastasizes to bone through a multistep process involving the detachment of cells from the primary tumor, their intravasation into the bloodstream, adhesion to the endothelium and extravasation into the bone, culminating with the establishment of a vicious cycle causing extensive bone lysis. In recent years, the crosstalk between tumor cells and secondary organs microenvironment is gaining much attention, being indicated as a crucial aspect in all metastatic steps. To investigate the complex interrelation between the tumor and the microenvironment, both in vitro and in vivo models have been exploited. In vitro models have some advantages over in vivo, mainly the possibility to thoroughly dissect in controlled conditions and with only human cells the cellular and molecular mechanisms underlying the metastatic progression. In this article we will review the main results deriving from in vitro co-culture models, describing mechanisms activated in the crosstalk between breast cancer and bone cells which drive the different metastatic steps.

[No abstract available]

Persistent circadian rhythm of bone turnover in bone metastatic breast cancer suggests greater skeletal retention of bisphosphonates if administered in the night. We assessed differential effects of night vs morning administration of zoledronic acid (ZA) on bone turnover. Forty-four breast cancer patients with bone metastases were randomised to receive intravenous ZA (4 mg) at 1100 or 2300 hours every 28 days for four times. Urinary concentration N-telopeptide of type-I collagen (NTX) and deoxypyridinolines, and serum C-telopeptide of type-I collagen (CTX), bone alkaline phosphatase (ALP), osteocalcin and Parathyroid hormone (PTH) was measured in the morning at baseline and after 4, 7, 14, 28, 56 and 84 days. Urinary ZA concentration was also measured. Zoledronic acid caused significant decreases of NTX and CTX (P

// Daniele Fanale 1, * , Valeria Amodeo 1, * , Viviana Bazan 1, * , Lavinia Insalaco 1 , Lorena Incorvaia 1 , Nadia Barraco 1 , Marta Castiglia 1 , Sergio Rizzo 1 , Daniele Santini 2 , Antonio Giordano 3 , Sergio Castorina 4, 5, # , Antonio Russo 1, # 1 Department of Surgical, Oncological and Oral Sciences, Section of Medical Oncology, University of Palermo, Palermo, Italy 2 University Campus Bio-Medico, Department of Medical Oncology, Rome, Italy 3 Sbarro Institute for Cancer Research and Molecular Medicine, Center for Biotechnology, College of Science and Technology, Temple University, Philadelphia, PA, USA 4 Fondazione Mediterranea “G.B. Morgagni”, Catania, Italy 5 Department of Biomedical and Biotechnological Sciences, University of Catania, Catania, Italy # Co-last authors, these authors contributed equally to this work * These authors contributed equally to this work Correspondence to: Antonio Russo, e-mail: antonio.russo@usa.net Keywords: bone metastasis, breast cancer, microarray analysis, miRNA expression profile, zoledronic acid Received: March 08, 2016 Accepted: March 31, 2016 Published: April 13, 2016 ABSTRACT 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 ability to identify prostate cancer patients at 'high risk' for bone metastasis development could allow early selection of those most likely to benefit from interventions to prevent or delay bone metastasis. This review is aimed to identify potential predictors of risk for bone metastasis in newly diagnosed patients and in those who have already received treatment. At diagnosis, established predictors of prostate cancer aggressiveness (e.g. PSA level, clinical stage, Gleason score) can identify patients at risk for bone metastasis. Following treatment of the disease, increasing evidence suggests that absolute PSA levels and other measures of PSA kinetics are useful to aid prediction of bone metastasis risk in patients both with and without a history of ADT. However, which PSA parameter most accurately predicts risk and the cut-off values that should be employed are unclear. Inclusion of PSA parameters to identify a high risk population may be beneficial in whom bone-modifying treatments are being considered. Other novel (but unvalidated) biomarkers that potentially predict the development of bone metastases have been identified, although it is unclear whether they will have value as independent markers or when combined with other parameters (e.g. measures of PSA kinetics). Further prospective studies of PSA kinetics and other predictive markers are, therefore, required to define the optimal criteria for identifying patients at high risk of bone metastases and those who are most likely to benefit from intensive monitoring and therapeutic intervention. (C) 2013 Elsevier Ltd. All rights reserved.

Introduction The aim of our study was to compare the detection rate of anti-3-18F-FACBC PET/CT in comparison with 11C-choline PET/CT in the evaluation of disease recurrence of PCa after radical prostatectomy. Patients and Methods Twenty-eight consecutive patients with biochemical relapse after radical prostatectomy were submitted to anti-3-18F-FACBC PET/CT and 11C-choline PET/CT to evaluate the site of disease recurrence. Androgen deprivation therapy was avoided in all cases. The primary end point was the overall detection rate of the 2 radiotracers. A patient-based analysis and a lesion-based analysis was performed. The target to background ratio (TBR) of each lesion was reported. Results At the time of PET scan, mean age was 67 years and mean prostate specific antigen (PSA) relapse was 2.9 ng/mL (range: 0.2-14.6). In patient-based analyses, 11C-choline PET/CT was positive in 5 patients and negative in 23 (detection rate = 17.8%) and anti-3-18F-FACBC PET/CT was positive in 10 patients and negative in 18 (detection rate = 35.7%). All lesions that were positive using 11C-choline were positive using anti-3-18F-FACBC PET/CT but with the latter radiotracer, 11 (61.1%) additional tumors were identified including 5 (17.8%) additional patients. The TBR of anti-3-18F-FACBC was greater than 11C-choline in 15 of 18 lesions, confirming a better image quality and contrast. Conclusion This preliminary study demonstrated that the detection rate of anti-3-18F-FACBC PET/CT is greater in comparison with 11C-choline, with approximately 20% of additional patients and approximately 60% additional lesions detected. Further studies, however, are required to assess the exact added value of this new tracer.

Receptor activator of NFκB ligand (RANKL), RANK, and osteoprotegerin (OPG) represent the key regulators of bone metabolism both in normal and pathological conditions, including bone metastases. To our knowledge, no previous studies investigated and compared RANK expression in primary tumors and in bone metastases from the same patient. We retrospectively examined RANK expression by immunohistochemistry in 74 bone metastases tissues from solid tumors, mostly breast, colorectal, renal, lung, and prostate cancer. For 40 cases, tissue from the corresponding primary tumor was also analyzed. Sixty-six (89%) of the 74 bone metastases were RANK-positive and, among these, 40 (59.5%) showed more than 50% of positive tumor cells. The median percentage of RANK-positive cells was 60% in primary tumors and metastases, without any statistically significant difference between the two groups (P = 0.194). The same percentage was obtained by considering only cases with availability of samples both from primary and metastasis. Our study shows that RANK is expressed by solid tumors, with high concordance between bone metastasis and corresponding primary tumor. These data highlight the central role of RANK/RANKL/OPG pathway as potential therapeutic target not only in bone metastasis management, but also in the adjuvant setting.

Cathepsin L is a lysosomal cysteine proteinase primarily devoted to the metabolic turnover of intracellular proteins. However, accumulating evidence suggests that this endopeptidase might also be implicated in the regulation of other important biological functions, including bone resorption in normal and pathological conditions. These findings support the concept that cathepsin L, in concert with other proteolytic enzymes involved in bone remodeling processes, could contribute to facilitate bone metastasis formation. In support of this hypothesis, recent studies indicate that cathepsin L can foster this process by triggering multiple mechanisms which, in part, differ from those of the major cysteine proteinase of osteoclasts, namely cathepsin K. Therefore, cathepsin L can be regarded as an additional target in the treatment of patients with metastatic bone disease. This review discusses the clinical and therapeutic implications related to these findings.

The clinical significance of serum cathepsin K and cystatin C was assessed in patients with breast cancer (BCa) or prostate cancer (PCa) with confined disease (M0) or bone metastasis (BM). Cathepsin K and cystatin C circulating levels were determined by ELISAs in 63 cancer patients, in 35 patients with nonmalignant diseases and in 42 healthy blood donors (control group). In BCa patients, cathepsin K serum levels were significantly lower than in sex matched control group (HS; p = 0.0008) or in patients with primary osteoporosis (OP; p = 0.0009). On the contrary, cystatin C levels were significantly higher in BCa patients than in HS ( p = 0.0001) or OP ( p = 0.017). In PCa patients, cathepsin K concentrations did not significantly differ from those measured in sex matched HS or in patients with benign prostatic hyperplasia (BPH). Conversely, cystatin C was more elevated in cancer patients than in controls ( p = 0.0001) or BPH patients ( p = 0.0078). Furthermore, in PCa patients, a positive correlation was observed between cystatin C and cathepsin K ( r S = 0.34; p = 0.047). No further relationship was highlighted between these molecules and the clinicobiological parameters of BCa or PCa progression including the number of bone lesions. Moreover, ROC curve analysis showed a poor diagnostic performance of cathepsin K and cystatin C in the detection of BM patients. Interestingly, the administration of zoledronic acid (ZA), a bisphosphonate derivative endowed with a potent antiosteoclastic activity, induced in BM patients a marked increase of cathepsin K and cystatin C serum levels compared to baseline values. However, this phenomenon was statistically significant only in the PCa group. In conclusion Cystatin C and cathepsin K may be regarded as possible markers to monitor the therapeutic response to bisphosphonate treatments. Nevertheless, their clinical value as specific gauges of skeletal metastasis remains questionable.

BACKGROUND: Bone metastases are devastating events resulting in disruption of local bone remodeling processes. Physiological bone turnover has a circadian rhythm. No data are available on the circadian pattern of bone turnover markers in patients with bone metastases. METHODS: Twenty post-menopausal women with breast cancer (BC) at first disease relapse and at least one bone metastasis were consecutively recruited. Twenty healthy women served as controls. Patients were free from concomitant chemotherapy/endocrine therapy. Throughout a 24-h period, urine samples were collected at 4-h intervals, and blood samples were collected at 4-h intervals between 08:00 and 24:00, and at 2-h intervals between 24:00 and 08:00. Serum osteocalcin (OC), total and bone-alkaline phosphatase (tALP and bALP, respectively) and C-terminal telopeptide of type I collagen (CTX), and urinary NTX and free deoxypyridinoline (fDPD) were measured together with serum parathyroid hormone (PTH) and serum and urinary calcium and phosphorus. Temporal variations of measured analytes were assessed by ANOVA and the COSINOR model. RESULTS: At 08:00, patients had higher levels of bone resorption indices (NTX, CTX and fDPD) than controls (p

Recent studies have highlighted that Activin A, a member of the transforming growth factor-beta (TGF-beta) superfamily, may be involved in the regulation of osteoblastic activity and in osteoclast differentiation. Therefore, we have investigated the clinical significance of its circulating levels in patients with bone metastasis. Activin A serum concentrations were determined, by a commercially available enzyme-linked immunosorbent assay kit, in 72 patients with breast cancer (BC) or prostatic cancer (PC) with (BM+) or without (BM-) bone metastases, in 15 female patients with age-related osteoporosis (OP), in 20 patients with benign prostatic hypertrophy (BPH) and in 48 registered healthy blood donors (HS) of both sex (25 female and 23 male). Activin A serum concentrations were significantly increased in BC or PC patients as compared to OP (P < 0.0001) or BPH (P = 0.045), respectively, or to sex matched HS (P < 0.0001). Additionally, these levels resulted more elevated in PC patients as compared to BC patients (P = 0.032). Interestingly, Activin A was significantly higher in BM+ patients than in BM- patients (BC, P = 0.047; PC, P = 0.016). In BC patients, a significant correlation was observed only between Activin A and number of bone metastases (P = 0.0065) while, in PC patients, Activin A levels were strongly correlated with the Gleason score (P = 0.011) or PSA levels (P = 0.0001) and, to a lessen extent, with the number of bone metastases (P = 0.056). Receiver operating characteristic curve (ROC) analysis showed a fair diagnostic accuracy of Activin A to discriminate between BM+ and BM- patients (BC: AUC = 0.71 +/- 0.09, P = 0.03; PC: AUC = 0.73 +/- 0.081, P = 0.005). These findings indicate that Activin A may be implicated in the pathogenesis of bone metastasis. Therefore, this cytokine may be considered a novel potential target for a more selective therapeutic approach in the treatment of skeletal metastasis and may be also useful as additional biochemical marker of metastatic bone disease.

Background: The effects of the bisphosphonate derivative zoledronic acid (ZA) on the > circulating levels of matrix metalloproteinase-2 (MMP-2), matrix metallo-proteinases-9 > (MMP-9), cathepsin B (Cath B) and urokinase-type plasminogen activator (uPA) in > patients with bone metastasis (BMTS) and the possible correlation with the symptomatic > response induced by this drug in these patients were evaluated. Patients and Methods: > Proteinase levels were determined by enzyme-linked immunosorbent assay (ELISA) in the > plasma of 30 patients with painful bone metastases from breast or prostate cancer > undergoing multiple treatment with ZA (4 mg i.v., every 4 weeks). Healthy subjects > (HS) of both genders (12 female and 30 male) served as the control group. The > symptomatic response to ZA was assessed by the visual analog scale score (VAS). > Results: The median MMP-2 and MMP-9 pretreatment levels were more elevated in BMTS as > compared to HS (p¡Ü0.0001). Conversely, uPA levels were lower in BMTS p=0.0033; no > significant difference was observed for Cath B. ZA administration was associated with > a symptomatic response (VAS score¡Ü4) in 25/30 patients (83.3%) (p phenomenon paralleled a decrease of Cath B and MMP-2 plasma concentrations from > baseline values on week 12 (p=0.05). A similar trend, although not statistically > significant, was also noted for MMP-9 and uPA. However, no direct relationship was > observed between the analgesic effect induced by ZA and changes in the circulating > levels of these enzymes. Conclusion: These data show that ZA administration may > provide relief from bone pain in patients with diffuse skeletal metastases and confirm > a possible implication of cysteine proteinases and matrix metalloproteinases in bone > metastasis formation, but not in the pathogenesis of metastatic bone pain.

Over the years, an increasing proportion of metastatic prostate cancer patients has been found to experience an initial bone flare phenomenon under both standard therapies (androgen deprivation therapy, chemotherapy, radiotherapy, abiraterone, enzalutamide) and novel agents (immunotherapy, bone-targeting radioisotopes). The underlying biological mechanisms of the flare phenomenon are still elusive and need further clarification, particularly in relation to different types of treatment and their treatment response assessment. Flare phenomenon is often underestimated and, in some cases, can negatively affect clinical outcome. In cases with suspected bone flare, the treatment should be continued for a minimum of 12 more weeks before further decisions about efficacy can be taken. Physicians and patients should be aware of this effect to avoid unwarranted anxiety and inadequate early discontinuation of treatment. This review aims at highlighting new evidence on flare phenomenon arising after the introduction of new drugs extending across the biochemical, radiographic and clinical spectrum of the disease.

Purpose: Aim of this study was to investigate for the presence of existing prognostic factors in patients with bone metastases (BMs) from RCC since bone represents an unfavorable site of metastasis for renal cell carcinoma (mRCC). Materials and methods: Data of patients with BMs from RCC were retrospectively collected. Age, sex, ECOG-Performance Status (PS), MSKCC group, tumor histology, presence of concomitant metastases to other sites, time from nephrectomy to bone metastases (TTBM, classified into three groups: 5 years) and time from BMs to skeletal-related event (SRE) were included in the Cox analysis to investigate their prognostic relevance. Results: 470 patients were enrolled in this analysis. In 19 patients (4%),bone was the only metastatic site; 277 patients had concomitant metastases in other sites. Median time to BMs was 16 months (range 0 - 44y) with Median OS of 17 months. Number of metastatic sites (including bone, p = 0.01), concomitant metastases, high Fuhrman grade (p < 0.001) and non-clear cell histology (p = 0.013) were significantly associated with poor prognosis. Patients with TTBM >5 years had longer OS (22 months) compared to patients with TTBM