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1.
J Bone Oncol ; 17: 100244, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31236323

RESUMO

BACKGROUND: Bone metastasis is one of the most common complications of advanced breast cancer. During dissemination to bone, breast cancer cells locate in a putative 'metastatic niche', a microenvironment that regulates the colonisation, maintenance of tumour cell dormancy and subsequent tumour growth. The precise location and composition of the bone metastatic niche is not clearly defined. We have used in vivo models of early breast cancer dissemination to provide novel evidence that demonstrates overlap between endosteal, perivascular, HSC and the metastatic niche in bone. METHODS: Estrogen Receptor (ER) +ve and -ve breast cancer cells were labelled with membrane dyes Vybrant-DiD and Vybrant-CM-DiI and injected via different routes in BALBc/nude mice of different ages. Two-photon microscopy was used to detect and quantitate tumour cells and map their location within the bone microenvironment as well as their distance to the nearest bone surface compared to the nearest other tumour cell. To investigate whether the metastatic niche overlapped with the HSC niche, animals were pre-treated with the CXCR4 antagonist AMD3100 to mobilise hematopoietic (HSCs) prior to injection of breast cancer cells. RESULTS: Breast cancer cells displayed a characteristic pattern of homing in the long bones, with the majority of tumour cells seeded in the trabecular regions, regardless of the route of injection, cell-line characteristics (ER status) or animal age. Breast cancer cells located in close proximity to the nearest bone surface and the average distance between individual tumour cells was higher than their distance to bone. Mobilisation of HSCs from the niche to the circulation prior to injection of cell lines resulted in increased numbers of tumour cells disseminated in trabecular regions. CONCLUSION: Our data provide evidence that homing of breast cancer cells is independent of their ER status and that the breast cancer bone metastasis niche is located within the trabecular region of bone, an area rich in osteoblasts and microvessels. The increased number of breast cancer cells homing to bone after mobilisation of HSCs suggests that the HSC and the bone metastasis niche overlap.

2.
Drug Discov Today ; 24(3): 763-772, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30496853

RESUMO

The current main goal of diagnostic medicine is to detect crucial events in 'infinitely' small samples. The key question now is how to determine whether the rare cell events isolated and characterized from these samples reliably reflect the disease and heterogeneity of the tumor. In this review, we provide a short overview of the most recent methods developed for the isolation and characterization of rare cell events in clinical practice, with a specific focus on circulating tumor cells. We discuss the biological value to studying these cells at the single cell level and how these rare cell events can reflect tumor heterogeneity. The potential biomedical applications are also critically discussed in light of precision medicine.


Assuntos
Células Neoplásicas Circulantes , Animais , Humanos , Neoplasias/genética
3.
Int J Mol Sci ; 19(10)2018 Sep 26.
Artigo em Inglês | MEDLINE | ID: mdl-30261597

RESUMO

Breast cancer cells colonize the skeleton by homing to specific niches, but the involvement of osteoblasts in tumour cell seeding, colonization, and progression is unknown. We used an in vivo model to determine how increasing the number of cells of the osteoblast lineage with parathyroid hormone (PTH) modified subsequent skeletal colonization by breast cancer cells. BALB/c nude mice were injected for five consecutive days with PBS (control) or PTH and then injected with DiD-labelled breast cancer cells via the intra-cardiac route. Effects of PTH on the bone microenvironment and tumour cell colonization and growth was analyzed using bioluminescence imaging, two-photon microscopy, and histological analysis. PTH treatment caused a significant, transient increase in osteoblast numbers compared to control, whereas bone volume/structure in the tibia was unaffected. There were no differences in the number of tumour cells seeding to the tibias, or in the number of tumours in the hind legs, between the control and PTH group. However, animals pre-treated with PTH had a significantly higher number of tumour colonies distributed throughout skeletal sites outside the hind limbs. This is the first demonstration that PTH-induced stimulation of osteoblastic cells may result in alternative skeletal sites becoming available for breast cancer cell colonization.


Assuntos
Osso e Ossos/efeitos dos fármacos , Neoplasias da Mama/patologia , Osteoblastos/efeitos dos fármacos , Hormônio Paratireóideo/farmacologia , Animais , Apoptose/efeitos dos fármacos , Osso e Ossos/patologia , Linhagem Celular Tumoral , Feminino , Humanos , Camundongos Endogâmicos BALB C , Camundongos Nus , Microscopia de Fluorescência por Excitação Multifotônica , Tíbia/efeitos dos fármacos , Tíbia/patologia , Transplante Heterólogo
4.
J Bone Oncol ; 12: 83-90, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-30123735

RESUMO

Osteosarcoma is a rare primary bone tumor, which mainly affects children and adolescents and has a poor prognosis, especially for patients with metastatic disease. A poor therapeutic response to the conventional chemotherapy is observed with the development of lung metastases, highlighting the need for improving the current regimens and the identification of early markers of the recurrent and metastatic disease. Circulating Tumour Cells (CTCs) play a key role in the metastatic process and could be powerful biomarkers of the progressive disease. The present study aimed to isolate CTCs by using a pre-clinical model of human osteosarcoma and to monitor their kinetic of release and their modulation by ifosfamide. CTCs were detectable into the bloodstream before any palpable primary tumors. Ifosfamide increased CTCs count and in contrast decreased the number of lung tumor nodules. On established tumors, ifosfamide slowed down the tumour growth and did not modulate CTC count that could be explained by a release of cancer cells from the primary tumour with reduced properties for inducing lung metastases. This report highlights the biological interest of CTCs in osteosarcoma.

5.
Cancer Lett ; 430: 193-200, 2018 08 28.
Artigo em Inglês | MEDLINE | ID: mdl-29802930

RESUMO

Osteosarcoma (OS) is the most frequent paediatric bone cancer, responsible for 9% of all cancer-related deaths in children. In this paper, a new strategy based on delivering edelfosine (ET) in lipid nanoparticles (LN) was explored in order to target the primary tumour and eliminate metastases. The in vitro and in vivo efficacy of the free drug, drug loaded into lipid nanoparticles (ET-LN) and doxorubicin (DOX) against osteosarcoma (OS) cells was analysed. ET and ET-LN decreased the growth of OS cells in vitro in a time- and dose-dependent manner. Interestingly, the uptake of ET and ET-LN was lower when OS cells were pre-treated with DOX. In vivo studies revealed that ET and ET-LN slowed down the primary tumour growth in two OS models. However, the combination of both drugs showed no additional anti-tumour effect. Importantly, ET-LN successfully prevented the metastatic spread of OS cells from the primary tumour to the lungs. On the whole, ET-LN are a promising candidate for OS chemotherapy.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Neoplasias Ósseas/tratamento farmacológico , Neoplasias Pulmonares/prevenção & controle , Osteossarcoma/tratamento farmacológico , Éteres Fosfolipídicos/administração & dosagem , Administração Oral , Animais , Neoplasias Ósseas/patologia , Linhagem Celular Tumoral , Doxorrubicina/administração & dosagem , Portadores de Fármacos/química , Composição de Medicamentos , Feminino , Humanos , Lipídeos/química , Neoplasias Pulmonares/secundário , Camundongos , Camundongos Nus , Nanopartículas/química , Osteossarcoma/patologia , Resultado do Tratamento , Ensaios Antitumorais Modelo de Xenoenxerto
6.
Calcif Tissue Int ; 102(2): 174-195, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29238848

RESUMO

Bone sarcomas are tumours belonging to the family of mesenchymal tumours and constitute a highly heterogeneous tumour group. The three main bone sarcomas are osteosarcoma, Ewing sarcoma and chondrosarcoma each subdivided in diverse histological entities. They are clinically characterised by a relatively high morbidity and mortality, especially in children and adolescents. Although these tumours are histologically, molecularly and genetically heterogeneous, they share a common involvement of the local microenvironment in their pathogenesis. This review gives a brief overview of their specificities and summarises the main therapeutic advances in the field of bone sarcoma.


Assuntos
Neoplasias Ósseas/etiologia , Sarcoma/etiologia , Neoplasias Ósseas/diagnóstico , Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/terapia , Feminino , Tumor de Células Gigantes do Osso/patologia , Humanos , Masculino , Sarcoma/diagnóstico , Sarcoma/tratamento farmacológico , Sarcoma/terapia , Microambiente Tumoral
8.
Expert Opin Drug Discov ; 12(4): 379-389, 2017 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-28277839

RESUMO

INTRODUCTION: Patients with metastatic cancer suffer the highest rate of cancer-related death, but existing animal models of metastasis have disadvantages that limit our ability to understand this process. The zebrafish is increasingly used for cancer modelling, particularly xenografting of human cancer cell lines, and drug discovery, and may provide novel scientific and therapeutic insights. However, this model system remains underexploited. Areas covered: The authors discuss the advantages and disadvantages of the zebrafish xenograft model for the study of cancer, metastasis and drug discovery. They summarise previous work investigating the metastatic cascade, such as tumour-induced angiogenesis, intravasation, extravasation, dissemination and homing, invasion at secondary sites, assessing metastatic potential and evaluation of cancer stem cells in zebrafish. Expert opinion: The practical advantages of zebrafish for basic biological study and drug discovery are indisputable. However, their ability to sufficiently reproduce and predict the behaviour of human cancer and metastasis remains unproven. For this to be resolved, novel mechanisms must to be discovered in zebrafish that are subsequently validated in humans, and for therapeutic interventions that modulate cancer favourably in zebrafish to successfully translate to human clinical studies. In the meantime, more work is required to establish the most informative methods in zebrafish.


Assuntos
Antineoplásicos/farmacologia , Descoberta de Drogas/métodos , Neoplasias/tratamento farmacológico , Animais , Humanos , Metástase Neoplásica , Neoplasias/patologia , Células-Tronco Neoplásicas/metabolismo , Neovascularização Patológica/patologia , Especificidade da Espécie , Ensaios Antitumorais Modelo de Xenoenxerto/métodos , Peixe-Zebra
9.
Eur J Cell Biol ; 96(2): 110-118, 2017 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-28187910

RESUMO

Gap junctions are transmembrane structures that directly connect the cytoplasm of adjacent cells, making intercellular communications possible. It has been shown that the behaviour of several tumours - such as bone tumours - is related to gap junction intercellular communications (GJIC). Several methodologies are available for studying GJIC, based on measuring different parameters that are useful for multiple applications, such as the study of carcinogenesis for example. These methods nevertheless have several limitations. The present manuscript describes the setting up of a dielectrophoresis (DEP)-based lab-on-a-chip platform for the real-time study of Gap Junctional Intercellular Communication between osteosarcoma cells and the main cells accessible to their microenvironment. We conclude that using the DEParray technology for the GJIC assessment has several advantages comparing to current techniques. This methodology is less harmful for cells integrity; cells can be recovered after interaction to make further molecular analysis; it is possible to study GJIC in real time; we can promote cell interactions using up to five different populations. The setting up of this new methodology overcomes several difficulties to perform experiments for solving questions about GJIC process that we are not able to do with current technics.


Assuntos
Comunicação Celular/fisiologia , Eletroforese em Microchip/instrumentação , Eletroforese em Microchip/métodos , Junções Comunicantes/fisiologia , Neoplasias Ósseas/patologia , Diferenciação Celular/fisiologia , Linhagem Celular Tumoral , Humanos , Células-Tronco Mesenquimais/citologia , Osteoblastos/citologia , Osteossarcoma/patologia , Imagem com Lapso de Tempo/métodos
10.
Cancer Lett ; 386: 189-195, 2017 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-27894960

RESUMO

Osteosarcoma is the most common primary bone tumour in children and adolescents and advanced osteosarcoma patients with evidence of metastasis share a poor prognosis. Osteosarcoma frequently gains resistance to standard therapies highlighting the need for improved treatment regimens and identification of novel therapeutic targets. Cancer stem cells (CSC) represent a sub-type of tumour cells attributed to critical steps in cancer including tumour propagation, therapy resistance, recurrence and in some cases metastasis. Recent published work demonstrates evidence of cancer stem cell phenotypes in osteosarcoma with links to drug resistance and tumorigenesis. In this review we will discuss the commonly used isolation techniques for cancer stem cells in osteosarcoma as well as the identified biochemical and molecular markers.


Assuntos
Antineoplásicos/uso terapêutico , Neoplasias Ósseas/patologia , Células-Tronco Neoplásicas/patologia , Osteossarcoma/patologia , Animais , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/genética , Neoplasias Ósseas/metabolismo , Separação Celular/métodos , Resistencia a Medicamentos Antineoplásicos , Humanos , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/metabolismo , Osteossarcoma/tratamento farmacológico , Osteossarcoma/genética , Osteossarcoma/metabolismo , Fenótipo
11.
Front Oncol ; 6: 202, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-27656422

RESUMO

Sarcomas are a heterogeneous group of malignant neoplasms of mesenchymal origin, many of which have a propensity to develop distant metastases. Cancer cells that have escaped from the primary tumor are able to invade into surrounding tissues, to intravasate into the bloodstream to become circulating tumor cells (CTCs), and are responsible for the generation of distant metastases. Due to the rarity of these tumors and the absence of specific markers expressed by sarcoma tumor cells, the characterization of sarcoma CTCs has to date been relatively limited. Current techniques for isolating sarcoma CTCs are based on size criteria, the identification of circulating cells that express either common mesenchymal markers, sarcoma-specific markers, such as CD99, CD81, or PAX3, and chromosomal translocations found in certain sarcoma subtypes, such as EWS-FLI1 in Ewing's sarcoma, detection of osteoblast-related genes, or measurement of the activity of specific metabolic enzymes. Further studies are needed to improve the isolation and characterization of sarcoma CTCs, to demonstrate their clinical significance as predictive and/or prognostic biomarkers, and to utilize CTCs as a tool for investigating the metastatic process in sarcoma and to identify novel therapeutic targets. The present review provides a short overview of the most recent literature on CTCs in sarcoma.

12.
Expert Opin Investig Drugs ; 25(11): 1265-1280, 2016 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-27633385

RESUMO

INTRODUCTION: Osteosarcomas are the main malignant primary bone tumours found in children and young adults. Conventional treatment is based on diagnosis and resection surgery, combined with polychemotherapy. This is a protocol that was established in the 1970s. Unfortunately, this therapeutic approach has reached a plateau of efficacy and the patient survival rate has not improved in the last four decades. New therapeutic approaches are thus required to improve the prognosis for osteosarcoma patients. Areas covered: From the databases available and published scientific literature, the present review gives an overview of the drugs currently in early clinical development for the treatment of osteosarcoma. For each drug, a short description is given of the relevant scientific data supporting its development. Expert opinion: Multidrug targeted approaches are set to emerge, given the heterogeneity of osteosarcoma subtypes and the multitude of therapeutic responses. The key role played by the microenvironment in the disease increases the number of therapeutic targets (such as macrophages or osteoclasts), as well as the master proteins that control cell proliferation or cell death. Ongoing phase I/II trials are important steps, not only for identifying new therapies with greater safety and efficacy, but also for better defining the role played by the microenvironment in the pathogenesis of osteosarcoma.


Assuntos
Antineoplásicos/uso terapêutico , Neoplasias Ósseas/tratamento farmacológico , Osteossarcoma/tratamento farmacológico , Animais , Antineoplásicos/farmacologia , Neoplasias Ósseas/patologia , Criança , Desenho de Drogas , Humanos , Terapia de Alvo Molecular , Osteossarcoma/patologia , Prognóstico , Microambiente Tumoral , Adulto Jovem
13.
J Exp Clin Cancer Res ; 34: 124, 2015 Oct 20.
Artigo em Inglês | MEDLINE | ID: mdl-26480944

RESUMO

BACKGROUND: While both preclinical and clinical studies suggest that the frequency of growing skeletal metastases is elevated in individuals with higher bone turnover, it is unclear whether this is a result of increased numbers of tumour cells arriving in active sites or of higher numbers of tumour cells being induced to divide by the bone micro-environment. Here we have investigated how the differences in bone turnover affect seeding of tumour cells and/or development of overt osteolytic bone metastasis using in vivo models of hormone-independent breast and prostate cancer. METHODS: Cohorts of 6 (young) and 16 (mature)-week old BALB/c nude mice were culled 1, 7 and 21 days after received intracardiac injection of luciferase expressing human prostate (PC3) or breast cancer (MDA-MB-231) cell lines labelled with a fluorescent cell membrane dye (Vybrant DiD). The presence of growing bone metastases was determined by bioluminescence using an in vivo imaging system (IVIS) and followed by anatomical confirmation of tumour metastatic sites post mortem, while the presence of individual fluorescently labelled tumour cells was evaluated using two-photon microscopy ex vivo. The bone remodelling activities were compared between young and mature naïve mice (both male and female) using micro-CT analysis, ELISA and bone histomorphometry. RESULTS: Both prostate and breast cancer cells generated higher numbers of overt skeletal lesions in young mice (~80%) than in mature mice (~20%). Although mature mice presented with fewer overt bone metastases, the number of tumour cells arriving/colonizing in the tibias was comparable between young and mature animals. Young naïve mice had lower bone volume but higher bone formation and resorption activities compared to mature animals. CONCLUSIONS: Our studies suggest that higher frequencies of growing osteolytic skeletal metastases in these models are linked to increased bone turnover and not to the initial number of tumour cells entering the bone microenvironment.


Assuntos
Neoplasias Ósseas/patologia , Neoplasias da Mama/patologia , Células Neoplásicas Circulantes/patologia , Osteólise/patologia , Neoplasias da Próstata/patologia , Animais , Neoplasias Ósseas/secundário , Contagem de Células , Linhagem Celular Tumoral , Modelos Animais de Doenças , Feminino , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Transplante de Neoplasias , Transplante Heterólogo
14.
FASEB J ; 29(8): 3141-50, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-25888599

RESUMO

This study aimed to identify subpopulations of prostate cancer cells that are responsible for the initiation of bone metastases. Using rapidly dividing human prostate cancer cell lines, we identified mitotically quiescent subpopulations (<1%), which we compared with the rapidly dividing populations for patterns of gene expression and for their ability to migrate to the skeletons of athymic mice. The study used 2-photon microscopy to map the presence/distribution of fluorescently labeled, quiescent cells and luciferase expression to determine the presence of growing bone metastases. We showed that the mitotically quiescent cells were very significantly more tumorigenic in forming bone metastases than fast-growing cells (55 vs. 15%) and had a unique gene expression profile. The quiescent cells were not uniquely stem cell like, with no expression of CD133 but had the same level expression of other putative prostate stem cell markers (CD44 and integrins α2/ß1), when compared to the rapidly proliferating population. In addition, mitotic quiescence was associated with very high levels of C-X-C chemokine receptor type 4 (CXCR4) production. Inhibition of CXCR4 activity altered the homing of quiescent tumor cells to bone. Our studies suggest that mitotic dormancy is a unique phenotype that facilitates tumor cell colonization of the skeleton in prostate cancer.


Assuntos
Biomarcadores Tumorais/metabolismo , Neoplasias Ósseas/patologia , Mitose/fisiologia , Neoplasias da Próstata/patologia , Antígeno AC133 , Animais , Antígenos CD/metabolismo , Neoplasias Ósseas/metabolismo , Glicoproteínas/metabolismo , Humanos , Receptores de Hialuronatos/metabolismo , Cadeias alfa de Integrinas/metabolismo , Cadeias beta de Integrinas/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Peptídeos/metabolismo , Fenótipo , Neoplasias da Próstata/metabolismo , Receptores CXCR4/metabolismo , Células Tumorais Cultivadas
15.
Int J Cancer ; 137(4): 968-77, 2015 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-25603921

RESUMO

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.


Assuntos
Neoplasias Ósseas/tratamento farmacológico , Neoplasias da Mama/patologia , Osteoprotegerina/administração & dosagem , Ovariectomia/efeitos adversos , Animais , Células da Medula Óssea/efeitos dos fármacos , Células da Medula Óssea/patologia , Neoplasias Ósseas/patologia , Neoplasias Ósseas/secundário , Reabsorção Óssea/tratamento farmacológico , Reabsorção Óssea/patologia , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/cirurgia , Proliferação de Células/efeitos dos fármacos , Modelos Animais de Doenças , Feminino , Humanos , Fragmentos Fc das Imunoglobulinas/administração & dosagem , Fragmentos Fc das Imunoglobulinas/imunologia , Camundongos , Osteoclastos , Osteoprotegerina/imunologia , Ligante RANK/metabolismo
16.
J Bone Miner Res ; 29(12): 2688-96, 2014 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-24956445

RESUMO

It has been suggested that metastasis-initiating cells gain a foothold in bone by homing to a metastastatic microenvironment (or "niche"). Whereas the precise nature of this niche remains to be established, it is likely to contain bone cell populations including osteoblasts and osteoclasts. In the mouse tibia, the distribution of osteoblasts on endocortical bone surfaces is non-uniform, and we hypothesize that studying co-localization of individual tumor cells with resident cell populations will reveal the identity of critical cellular components of the niche. In this study, we have mapped the distribution of three human prostate cancer cell lines (PC3-NW1, LN-CaP, and C4 2B4) colonizing the tibiae of athymic mice following intracardiac injection and evaluated their interaction with potential metastatic niches. Prostate cancer cells labeled with the fluorescent cell membrane dye (Vybrant DiD) were found by two-photon microscopy to be engrafted in the tibiae in close proximity (∼40 µm) to bone surfaces and 70% more cancer cells were detected in the lateral compared to the medial endocortical bone regions. This was associated with a 5-fold higher number of osteoblasts and 7-fold higher bone formation rate on the lateral endocortical bone surface compared to the medial side. By disrupting cellular interactions mediated by the chemokine (C-X-C motif) receptor 4 (CXCR4)/chemokine ligand 12 (CXCL12) axis with the CXCR4 inhibitor AMD3100, the preferential homing pattern of prostate cancer cells to osteoblast-rich bone surfaces was disrupted. In this study, we map the location of prostate cancer cells that home to endocortical regions in bone and our data demonstrate that homing of prostate cancer cells is associated with the presence and activity of osteoblast lineage cells, and suggest that therapies targeting osteoblast niches should be considered to prevent development of incurable prostate cancer bone metastases.


Assuntos
Neoplasias Ósseas/metabolismo , Neoplasias Ósseas/secundário , Neoplasias Experimentais/metabolismo , Osteoblastos/metabolismo , Neoplasias da Próstata/metabolismo , Animais , Neoplasias Ósseas/patologia , Linhagem Celular Tumoral , Quimiocina CXCL12/metabolismo , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Metástase Neoplásica , Neoplasias Experimentais/patologia , Osteoblastos/patologia , Neoplasias da Próstata/patologia , Receptores CXCR4/metabolismo
17.
Bone ; 66: 240-50, 2014 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-24971713

RESUMO

INTRODUCTION: Bone metastasis is the most common complication of advanced breast cancer. The associated cancer-induced bone disease is treated with bone-sparing agents like zoledronic acid. Clinical trials have shown that zoledronic acid also reduces breast cancer recurrence in bone; potentially by modifying the bone microenvironment surrounding disseminated tumour cells. We have characterised the early effects of zoledronic acid on key cell types of the metastatic niche in vivo, and investigated how these modify the location of breast tumour cells homing to bone. METHODS: Female mice were treated with a single, clinically achievable dose of zoledronic acid (100µg/kg) or PBS. Bone integrity, osteoclast and osteoblast activity and number/mm trabecular bone on 1, 3, 5 and 10days after treatment were assessed using µCT, ELISA (TRAP, PINP) and bone histomorphometry, respectively. The effect of zoledronic acid on osteoblasts was validated in genetically engineered mice with GFP-positive osteoblastic cells. The effects on growth plate cartilage were visualised by toluidine blue staining. For tumour studies, mice were injected i.c. with DID-labelled MDA-MB-231-NW1-luc2 breast cancer cells 5days after zoledronic acid treatment, followed by assessment of tumour cell homing to bone and soft tissues by multiphoton microscopy, flow cytometry and ex vivo cultures. RESULTS: As early as 3days after treatment, animals receiving zoledronic acid had significantly increased trabecular bone volume vs. control. This rapid bone effect was reflected in a significant reduction in osteoclast and osteoblast number/mm trabecular bone and reduced bone marker serum levels (day 3-5). These results were confirmed in mice expressing GFP in osteoblastic linage cells. Pre-treatment with zoledronic acid caused accumulation of an extra-cellular matrix in the growth plate associated with a trend towards preferential [1] homing of tumour cells to osteoblast-rich areas of bone, but without affecting the total number of tumour cells. The number of circulating tumour cells was reduced in ZOL treated animals. CONCLUSION: A single dose of zoledronic acid caused significant changes in the bone area suggested to contain the metastatic niche. Tumour cells arriving in this modified bone microenvironment appeared to preferentially locate to osteoblast-rich areas, supporting that osteoblasts may be key components of the bone metastasis niche and therefore a potential therapeutic target in breast cancer.


Assuntos
Neoplasias Ósseas/secundário , Neoplasias da Mama/patologia , Difosfonatos/farmacologia , Imidazóis/farmacologia , Osteoblastos/patologia , Animais , Neoplasias Ósseas/tratamento farmacológico , Osso e Ossos/efeitos dos fármacos , Osso e Ossos/patologia , Linhagem Celular Tumoral , Difosfonatos/uso terapêutico , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Feminino , Proteínas de Fluorescência Verde/metabolismo , Humanos , Imidazóis/uso terapêutico , Hospedeiro Imunocomprometido , Camundongos Endogâmicos BALB C , Camundongos Nus , Tamanho do Órgão/efeitos dos fármacos , Osteoblastos/efeitos dos fármacos , Osteoclastos/efeitos dos fármacos , Osteoclastos/patologia , Reprodutibilidade dos Testes , Ácido Zoledrônico
18.
Clin Cancer Res ; 20(11): 2922-32, 2014 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-24687923

RESUMO

PURPOSE: Clinical trials in early breast cancer have suggested that benefits of adjuvant bone-targeted treatments are restricted to women with established menopause. We developed models that mimic pre- and postmenopausal status to investigate effects of altered bone turnover on growth of disseminated breast tumor cells. Here, we report a differential antitumor effect of zoledronic acid (ZOL) in these two settings. EXPERIMENTAL DESIGN: Twleve-week-old female Balb/c-nude mice with disseminated MDA-MB-231 breast tumor cells in bone underwent sham operation or ovariectomy (OVX), mimicking the pre- and postmenopausal bone microenvironment, respectively. To determine the effects of bone-targeted therapy, sham/OVX animals received saline or 100 µg/kg ZOL weekly. Tumor growth was assessed by in vivo imaging and effects on bone by real-time PCR, micro-CT, histomorphometry, and measurements of bone markers. Disseminated tumor cells were detected by two-photon microscopy. RESULTS: OVX increased bone resorption and induced growth of disseminated tumor cells in bone. Tumors were detected in 83% of animals following OVX (postmenopausal model) compared with 17% following sham operation (premenopausal model). OVX had no effect on tumors outside of bone. OVX-induced tumor growth was completely prevented by ZOL, despite the presence of disseminated tumor cells. ZOL did not affect tumor growth in bone in the sham-operated animals. ZOL increased bone volume in both groups. CONCLUSIONS: This is the first demonstration that tumor growth is driven by osteoclast-mediated mechanisms in models that mimic post- but not premenopausal bone, providing a biologic rationale for the differential antitumor effects of ZOL reported in these settings. Clin Cancer Res; 20(11); 2922-32. ©2014 AACR.


Assuntos
Conservadores da Densidade Óssea/farmacologia , Neoplasias Ósseas/secundário , Osso e Ossos/efeitos dos fármacos , Neoplasias da Mama/patologia , Microambiente Celular , Difosfonatos/farmacologia , Imidazóis/farmacologia , Animais , Linhagem Celular Tumoral , Modelos Animais de Doenças , Feminino , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Microscopia Confocal , Osteoclastos/efeitos dos fármacos , Osteoclastos/metabolismo , Ovariectomia , Pós-Menopausa/efeitos dos fármacos , Pós-Menopausa/metabolismo , Pré-Menopausa/efeitos dos fármacos , Pré-Menopausa/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Microtomografia por Raio-X , Ensaios Antitumorais Modelo de Xenoenxerto , Ácido Zoledrônico
19.
Endocr Relat Cancer ; 21(2): 327-41, 2014 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-24413608

RESUMO

Advanced breast cancer is associated with the development of incurable bone metastasis. The two key processes involved, tumour cell homing to and subsequent colonisation of bone, remain to be clearly defined. Genetic studies have indicated that different genes facilitate homing and colonisation of secondary sites. To identify specific changes in gene and protein expression associated with bone-homing or colonisation, we have developed a novel bone-seeking clone of MDA-MB-231 breast cancer cells that exclusively forms tumours in long bones following i.v. injection in nude mice. Bone-homing cells were indistinguishable from parental cells in terms of growth rate in vitro and when grown subcutaneously in vivo. Only bone-homing ability differed between the lines; once established in bone, tumours from both lines displayed similar rates of progression and caused the same extent of lytic bone disease. By comparing the molecular profile of a panel of metastasis-associated genes, we have identified differential expression profiles associated with bone-homing or colonisation. Bone-homing cells had decreased expression of the cell adhesion molecule fibronectin and the migration and calcium signal binding protein S100A4, in addition to increased expression of interleukin 1B. Bone colonisation was associated with increased fibronectin and upregulation of molecules influencing signal transduction pathways and breakdown of extracellular matrix, including hRAS and matrix metalloproteinase 9. Our data support the hypothesis that during early stages of breast cancer bone metastasis, a specific set of genes are altered to facilitate bone-homing, and that disruption of these may be required for effective therapeutic targeting of this process.


Assuntos
Neoplasias Ósseas/genética , Neoplasias Ósseas/secundário , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Animais , Neoplasias Ósseas/metabolismo , Neoplasias Ósseas/patologia , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Movimento Celular , Feminino , Perfilação da Expressão Gênica , Humanos , Interleucina-1beta/sangue , Interleucina-1beta/metabolismo , Camundongos , Camundongos Nus
20.
Breast Cancer Res ; 14(3): R86, 2012 May 25.
Artigo em Inglês | MEDLINE | ID: mdl-22632416

RESUMO

INTRODUCTION: The majority of deaths from breast cancer are a result of metastases; however, little is understood about the genetic alterations underlying their onset. Genetic profiling has identified the adhesion molecule plakoglobin as being three-fold reduced in expression in primary breast tumors that have metastasized compared with nonmetastatic tumors. In this study, we demonstrate a functional role for plakoglobin in the shedding of tumor cells from the primary site into the circulation. METHODS: We investigated the effects of plakoglobin knockdown on breast cancer cell proliferation, migration, adhesion, and invasion in vitro and on tumor growth and intravasation in vivo. MCF7 and T47D cells were stably transfected with miRNA sequences targeting the plakoglobin gene, or scramble vector. Gene and protein expression was monitored by quantitative polymerase chain reaction (qPCR) and Western blot. Cell proliferation, adhesion, migration, and invasion were measured by cell counting, flow cytometry, and scratch and Boyden Chamber assays. For in vivo experiments, plakoglobin knockdown and control cells were inoculated into mammary fat pads of mice, and tumor growth, shedding of tumor cells into the bloodstream, and evidence of metastatic bone lesions were monitored with caliper measurement, flow cytometry, and microcomputed tomography (µCT), respectively. RESULTS: Plakoglobin and γ-catenin expression were reduced by more than 80% in all knockdown cell lines used but were unaltered after transfection with the scrambled sequence. Reduced plakoglobin resulted in significantly increased in MCF7 and T47D cell proliferation in vitro and in vivo, compared with control, with significantly more tumor cells being shed into the bloodstream of mice bearing plakoglobin knockdown tumors. In addition, plakoglobin knockdown cells showed a >250% increase in invasion through basement membrane and exhibited reduced cell-to-cell adhesion compared with control cells. CONCLUSION: Decreased plakoglobin expression increases the invasive behavior of breast cancer cells. This is the first demonstration of a functional role for plakoglobin/γ-catenin in the metastatic process, indicating that this molecule may represent a target for antimetastatic therapies.


Assuntos
Neoplasias Mamárias Animais/metabolismo , Neoplasias Mamárias Animais/patologia , Células Neoplásicas Circulantes , gama Catenina/metabolismo , Animais , Adesão Celular , Comunicação Celular , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Células MCF-7 , Neoplasias Mamárias Animais/genética , Camundongos , Camundongos Endogâmicos BALB C , Invasividade Neoplásica , Interferência de RNA , RNA Interferente Pequeno , gama Catenina/genética
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