ABCC5 supports osteoclast formation and promotes breast cancer metastasis to bone

INTRODUCTION: Bone is the most common site of breast cancer metastasis, and complications associated with bone metastases can lead to a significantly decreased patient quality of life. Thus, it is essential to gain a better understanding of the molecular mechanisms that underlie the emergence and growth of breast cancer skeletal metastases. METHODS: To search for novel molecular mediators that influence breast cancer bone metastasis, we generated gene-expression profiles from laser-capture microdissected trephine biopsies of both breast cancer bone metastases and independent primary breast tumors that metastasized to bone. Bioinformatics analysis identified genes that are differentially expressed in breast cancer bone metastases compared with primary, bone-metastatic breast tumors. RESULTS: ABCC5, an ATP-dependent transporter, was found to be overexpressed in breast cancer osseous metastases relative to primary breast tumors. In addition, ABCC5 was significantly upregulated in human and mouse breast cancer cell lines with high bone-metastatic potential. Stable knockdown of ABCC5 substantially reduced bone metastatic burden and osteolytic bone destruction in mice. The decrease in osteolysis was further associated with diminished osteoclast numbers in vivo. Finally, conditioned media from breast cancer cells with reduced ABCC5 expression failed to induce in vitro osteoclastogenesis to the same extent as conditioned media from breast cancer cells expressing ABCC5. CONCLUSIONS: Our data suggest that ABCC5 functions as a mediator of breast cancer skeletal metastasis. ABCC5 expression in breast cancer cells is important for efficient osteoclast-mediated bone resorption. Hence, ABCC5 may be a potential therapeutic target for breast cancer bone metastasis

5′-Inositol phosphatase SHIP2 recruits Mena to stabilize invadopodia for cancer cell invasion

Invadopodia are specialized membrane protrusions that support degradation of extracellular matrix (ECM) by cancer cells, allowing invasion and metastatic spread. Although early stages of invadopodia assembly have been elucidated, little is known about maturation of invadopodia into structures competent for ECM proteolysis. The localized conversion of phosphatidylinositol(3,4,5)-triphosphate and accumulation of phosphatidylinositol(3,4)-bisphosphate at invadopodia is a key determinant for invadopodia maturation. Here we investigate the role of the 5′-inositol phosphatase, SHIP2, and reveal an unexpected scaffold function of SHIP2 as a prerequisite for invadopodia-mediated ECM degradation. Through biochemical and structure-function analyses, we identify specific interactions between SHIP2 and Mena, an Ena/VASP-family actin regulatory protein. We demonstrate that SHIP2 recruits Mena, but not VASP, to invadopodia and that disruption of SHIP2–Mena interaction in cancer cells leads to attenuated capacity for ECM degradation and invasion in vitro, as well as reduced metastasis in vivo. Together, these findings identify SHIP2 as a key modulator of carcinoma invasiveness and a target for metastatic disease

Molecular mediators of breast cancer bone metastasis

Breast cancer is the most frequently diagnosed and the second leading cause of cancer deaths in Canadian women. The most devastating and deadly feature of the disease is the emergence of metastases. Breast cancer most commonly metastasizes to bone, often leading to a significantly decreased quality of life in affected patients. Despite progress in understanding the underlying molecular biology of breast tumors that relapse to bone, to date there are no therapies capable of curing the disease. Hence, it is essential to gain a more in-depth knowledge of the molecular mechanisms that underlie the emergence and growth of breast cancer skeletal metastases. Consequently, it was attempted to: 1) examine the efficacy of targeting a known pathway important for breast cancer metastasis to bone, 2) identify novel mediators of this process and 3) develop a stratification tool capable of identifying patients with breast cancer that possesses a high likelihood of spreading to bone. Transforming growth factor-beta (TGF-β) signaling is a potent modulator of the invasive and metastatic behavior of breast cancer cells. The work in this thesis demonstrates that expression of a TGF-β ligand trap, which neutralizes TGF-β1 and TGF-β3 in breast cancer cells, diminished their outgrowth in bone and reduced the severity of osteolytic lesion formation. It is further shown that a reduction or loss of host-derived TGF-β1 reduced the incidence of breast tumor outgrowth in the skeleton. Moreover, tumor cells capable of growing within the bone of a TGF-β1 deficient host up-regulated expression of all three TGF-β isoforms within the tumor cells themselves, effectively bypassing the host-deficiency. Next, a gene discovery approach was undertaken to identify novel candidate mediators of breast cancer skeletal metastasis. Invasive breast epithelium was selectively isolated by laser capture microdissection (LCM) performed on bone metastases and primary tumors from patients displaying breast cancer with subsequent recurrence to the skeleton. In this search, ABCC5 was found to be overexpressed in osseous metastases compared to primary mammary tumors metastatic to bone. Furthermore, this protein was detected at substantially higher levels in human and mouse breast cancer cells, which metastasize to bone in animal models. Importantly, removal of this protein from these cells resulted in their decreased ability to induce osteolytic bone lesions, which was correlated with a decreased recruitment of osteoclasts, cells responsible for the bone resorption process. Finally, the molecular changes occurring within the primary breast tumor were investigated in an attempt to identify a prognostic bone metastatic signature. Gene expression profiling was performed on estrogen receptor (ER)-positive primary breast tumors metastastatic to bone and breast cancers, which spread to soft tissue. A 25-gene signature was derived from the top 100 differentially expressed probes and was found to be capable of discriminating breast tumors metastatic to bone from cancers recurring to visceral sites in an independent gene expression dataset.Le cancer du sein est le cancer le plus fréquemment diagnostiqué et la deuxième cause de décès par cancer chez les femmes canadiennes. La caractéristique la plus dramatique et la plus mortelle de cette maladie est l'apparition de métastases. Le cancer du sein métastase le plus souvent au niveau des os, conduisant fréquemment à une qualité de la vie sensiblement diminuée chez les patients atteints. Malgré les progrès réalisés dans la compréhension de la biologie moléculaire sous-jacente des tumeurs du sein qui colonisent l'os, il n'existe, à ce jour, aucun traitement capable de guérir cette affection. Il est ainsi primordial d'acquérir une connaissance plus approfondie des mécanismes moléculaires qui sont à l'origine de l'émergence et de la croissance des métastases du cancer du sein au niveau du squelette. Par conséquent, il fut envisagé: d'examiner l'efficacité du ciblage d'une voie connue, importante pour que le cancer du sein puisse métastaser au niveau de l'os; d'identifier les nouveaux médiateurs de ce processus et de développer un outil de discrimination capable d'identifier les patients atteints d'un cancer du sein qui possède une forte probabilité de dissémination à l'os. La voie de signalisation du facteur de croissance transformant bêta (TGF-β) est un puissant modulateur du comportement invasif et métastatique des cellules cancéreuses du sein. Le travail présenté dans cette thèse démontre que l'expression d'une molécule leurre du ligand du TGF-β, qui neutralise le TGF-β1 et le TGF-β3 dans les cellules cancéreuses du sein, réduit leur développement dans les os et ainsi que la gravité de la formation des lésions ostéolytiques. Il est également démontré qu'une réduction, ou une perte, de l'hôte dérivé du TGF-β1 réduit la fréquence d'apparition d'une excroissance de la tumeur mammaire au niveau du squelette. Par ailleurs, les cellules tumorales capables de croître au sein d'un tissu osseux hôte déficient en TGF-β1 ont montré une augmentation de l'expression des trois isoformes du TGF-β dans les cellules tumorales elles-mêmes, court-circuitant ainsi efficacement cette carence de l'hôte. Une approche de découverte de gènes a été ensuite entreprise pour identifier des nouveaux médiateurs candidats des métastases squelettiques des cancers du sein. L'épithélium invasif du sein a été sélectivement isolé par microdissection à capture laser (LCM), et ceci a été réalisé sur les métastases osseuses et des tumeurs primaires de patientes présentant un cancer du sein avec récidive ultérieure au niveau du squelette. Dans cette recherche, ABCC5 fut montré comme étant surexprimé dans les métastases osseuses, par rapport à des tumeurs primaires mammaires métastasant au niveau l'os. De plus, cette protéine a été détectée à des niveaux nettement plus élevés dans des cellules cancéreuses mammaires d'origine humaine et murine qui métastasent dans l'os dans des modèles animaux. Une autre donnée importante fut que la suppression de cette protéine dans les cellules concernées conduisit à une réduction de leur capacité à induire des lésions osseuses ostéolytiques, ce qui fut corrélée avec une diminution du recrutement d'ostéoclastes, les cellules responsables du processus de résorption osseuse. Pour terminer, les changements moléculaires qui se produisent au sein de la tumeur primaire du sein ont été étudiées dans le but d'identifier une signature pronostiquant des métastases osseuses. Un profilage de l'expression génique a été réalisé sur les tumeurs mammaires primaires positives pour le récepteur aux œstrogènes (ER) et métastasant à l'os mais aussi sur les cancers mammaires, qui se propagent aux tissus mous. Une signature de 25 gènes a été sélectionnée à partir des 100 meilleures sondes exprimées différentiellement et a montré sa capacité à discriminer d'un coté les tumeurs du sein métastasant à l'os et de l'autre les cancers récurrents sur des sites viscéraux et ceci à partir d'un ensemble de données sur l'expression de gènes indépendants

Transforming growth factor-\u3b21 is the predominant isoform required for breast cancer cell outgrowth in bone

Transforming growth factor (TGF)-\u3b2 signaling is a potent modulator of the invasive and metastatic behavior of breast cancer cells. Indeed, breast tumor responsiveness to TGF-\u3b2 is important for the development of osteolytic bone metastases. However, the specific TGF-\u3b2 isoforms that promote breast cancer outgrowth in bone is unknown. We demonstrate that expression of a TGF-\u3b2 ligand trap, which neutralizes TGF-\u3b21 and TGF-\u3b23, in MDA-MB-231 breast cancer cells diminished their outgrowth in bone and reduced the severity of osteolytic lesion formation when compared with controls. We further show that a reduction or loss of TGF-\u3b21 expression within the bone microenvironment of TGF-\u3b21+/- and TGF-\u3b21-/- mice significantly reduced the incidence of breast tumor outgrowth compared with wild-type animals. Interestingly, those tumors capable of growing within the tibiae of TGF-\u3b21- deficient mice had upregulated expression of all three TGF-\u3b2 isoforms. Finally, breast cancer cells expressing the TGF-\u3b2 ligand trap showed a pronounced reduction in their ability to form osteolytic lesions when injected into the tibiae of TGF-b1\ufe +/- mice. Thus, our studies show that both host- and tumor-derived TGF-\u3b2 expression plays a critical role during the establishment and outgrowth of breast cancer cells in bone.NRC publication: Ye

ABCC5 supports osteoclast formation and promotes breast cancer metastasis to bone

Abstract Introduction Bone is the most common site of breast cancer metastasis, and complications associated with bone metastases can lead to a significantly decreased patient quality of life. Thus, it is essential to gain a better understanding of the molecular mechanisms that underlie the emergence and growth of breast cancer skeletal metastases. Methods To search for novel molecular mediators that influence breast cancer bone metastasis, we generated gene-expression profiles from laser-capture microdissected trephine biopsies of both breast cancer bone metastases and independent primary breast tumors that metastasized to bone. Bioinformatics analysis identified genes that are differentially expressed in breast cancer bone metastases compared with primary, bone-metastatic breast tumors. Results ABCC5, an ATP-dependent transporter, was found to be overexpressed in breast cancer osseous metastases relative to primary breast tumors. In addition, ABCC5 was significantly upregulated in human and mouse breast cancer cell lines with high bone-metastatic potential. Stable knockdown of ABCC5 substantially reduced bone metastatic burden and osteolytic bone destruction in mice. The decrease in osteolysis was further associated with diminished osteoclast numbers in vivo. Finally, conditioned media from breast cancer cells with reduced ABCC5 expression failed to induce in vitro osteoclastogenesis to the same extent as conditioned media from breast cancer cells expressing ABCC5. Conclusions Our data suggest that ABCC5 functions as a mediator of breast cancer skeletal metastasis. ABCC5 expression in breast cancer cells is important for efficient osteoclast-mediated bone resorption. Hence, ABCC5 may be a potential therapeutic target for breast cancer bone metastasis