Bone morphogenetic protein 7 (BMP7) counteracts the physiological epithelial-to-mesenchymal transition (EMT), a process that is indicative of epithelial plasticity. Because EMT is involved in cancer, we investigated whether BMP7 plays a role in breast cancer growth and metastasis. In this study, we show that decreased BMP7 expression in primary breast cancer is significantly associated with the formation of clinically overt bone metastases in patients with z10 years of follow-up. In line with these clinical observations, BMP7 expression is inversely related to tumorigenicity and invasive behavior of human breast cancer cell lines. Moreover, BMP7 decreased the expression of vimentin, a mesenchymal marker associated with invasiveness and poor prognosis, in human MDA-MB-231 (MDA-231)-B/Luc + breast cancer cells under basal and transforming growth factor-B (TGF-B)-stimulated conditions. In addition, exogenous addition of BMP7 to TGF-B-stimulated MDA-231 cells inhibited Smad-mediated TGF-B signaling. Furthermore, in a well-established bone metastasis model using whole-body bioluminescent reporter imaging, stable overexpression of BMP7 in MDA-231 cells inhibited de novo formation and progression of osteolytic bone metastases and, hence, their metastatic capability. In line with these observations, daily i.v. administration of BMP7 (100 Mg/kg/d) significantly inhibited orthotopic and intrabone growth of MDA-231-B/Luc + cells in nude mice. Our data suggest that decreased BMP7 expression during carcinogenesis in the human breast contributes to the acquisition of a bone metastatic phenotype. Because exogenous BMP7 can still counteract the breast cancer growth at the primary site and in bone, BMP7 may represent a novel therapeutic molecule for repression of local and bone metastatic growth of breast cancer. [Cancer Res 2007;67(18):8742-51]

In breast cancer bone metastasis, tumor cells stimulate osteoclast-mediated bone resorption, and bone-derived growth factors released from resorbed bone stimulate tumor growth. The A v B 3 integrin is an adhesion receptor expressed by breast cancer cells and osteoclasts. It is implicated in tumor cell invasion and osteoclast-mediated bone resorption. Here, we hypothesized that the therapeutic targeting of tumor A v B 3 integrin would prevent bone metastasis formation. We first showed that, compared with mock-transfected cells, the i.v. inoculation of A v B 3 -overexpressing MDA-MB-231 breast cancer cells in animals increased bone metastasis incidence and promoted both skeletal tumor burden and bone destruction. The direct inoculation of A v B 3 -overexpressing transfectants into the tibial bone marrow cavity did not however enhance skeletal tumor burden and bone destruction, suggesting that A v B 3 controls earlier events during bone metastasis formation. We next examined whether a nonpeptide antagonist of A v B 3 (PSK1404) exhibits meaningful antitumor effects in experimental breast and ovarian cancer bone metastasis. A continuous PSK1404 treatment, which inhibited osteoclast-mediated bone resorption in an animal model of bone loss, substantially reduced bone destruction and decreased skeletal tumor burden. Importantly, a short-term PSK1404 treatment that did not inhibit osteoclast activity also decreased skeletal tumor burden and bone destruction. This dosing regimen caused a profound and specific inhibition of bone marrow colonization by green fluorescent protein, A v B 3 -expressing tumor cells in vivo and blocked tumor cell invasion in vitro. Overall, our data show that tumor A v B 3 integrin stands as a therapeutic target for the prevention of skeletal metastases. [Cancer Res 2007;67(12):5821-30]

miRNAs are master regulators of gene expression that play key roles in cancer metastasis. During bone metastasis, metastatic tumor cells must rewire their biology and express genes that are normally expressed by bone cells (a process called osteomimicry), which endow tumor cells with full competence for outgrowth in the bone marrow. Here, we establish miR-30 family members miR-30a, miR-30b, miR-30c, miR-30d, and miR-30e as suppressors of breast cancer bone metastasis that regulate multiple pathways, including osteomimicry. Low expression of miR-30 in primary tumors from patients with breast cancer were associated with poor relapse-free survival. In addition, estrogen receptor (ER)-negative/progesterone receptor (PR)-negative breast cancer cells expressed lower miR-30 levels than their ER/PR-positive counterparts. Overexpression of miR-30 in ER/PR-negative breast cancer cells resulted in the reduction of bone metastasis burden , miR-30 did not affect tumor cell proliferation, but did inhibit tumor cell invasion. Furthermore, overexpression of miR-30 restored bone homeostasis by reversing the effects of tumor cell-conditioned medium on osteoclastogenesis and osteoblastogenesis. A number of genes associated with osteoclastogenesis stimulation (), osteoblastogenesis inhibition (), tumor cell osteomimicry (), and invasiveness () were identified as targets for repression by miR-30. Among these genes, silencing or in ER-/PR-negative breast cancer cells recapitulated inhibitory effects of miR-30 on skeletal tumor burden Overall, our findings provide evidence that miR-30 family members employ multiple mechanisms to impede breast cancer bone metastasis and may represent attractive targets for therapeutic intervention. These findings suggest miR-30 family members may serve as an effective means to therapeutically attenuate metastasis in triple-negative breast cancer. .

Metastatic breast cancer cells exhibit the selective ability to seed and grow in the skeleton. We and others have previously reported that human breast tumors which metastasize to the skeleton overexpress bone matrix extracellular proteins. In an attempt to reveal the osteoblast-like phenotype of osteotropic breast cancer cells, we performed a microarray study on a model of breast cancer bone metastasis consisting of the MDA-MB-231 human cell line and its variant B02 selected for its high capacity to form bone metastases in vivo. Analysis of B02 cells transcriptional profile revealed that 11 and 9 out of the 50 most up- and down-regulated mRNAs, respectively, corresponded to genes which expression has been previously associated with osteoblastic differentiation process. Thus, osteoblast specific cadherin 11 which mediates the differentiation of mesenchymal cells into osteoblastic cells is up-regulated in B02. While S100A4, recently described as a key negative regulator of osteoblast differentiation, is the most down-regulated gene in B02 cells. RT-PCR and western blotting experiments allowed the validation of the modulation of several genes of interest. Using immunohistochemistry, performed on human breast primary tumors and their matched liver and bone metastases, we were able to confirm that the osteoblast-like pattern of gene expression observed in our model holds true in vivo. This is the first report demonstrating a gene-expression pattern corresponding to the acquisition of an osteomimetic phenotype by bone metastatic breast cancer cells.

Key Points• We demonstrate that PT promotes ECFCs dysfunction by inducing stress-induced premature senescence.• Our data reveal that SIRT1 deficiency drives PT-ECFC senescence, and acts as a critical determinant of the PT-ECFC angiogenic defect.Epidemiological and experimental studies indicate that early vascular dysfunction occurs in low-birth-weight subjects, especially preterm (PT) infants. We recently reported impaired angiogenic activity of endothelial colony-forming cells (ECFCs) in this condition. We hypothesized that ECFC dysfunction in PT might result from premature senescence and investigated the underlying mechanisms. Compared with ECFCs from term neonates (n 5 18), ECFCs isolated from PT (n 5 29) display an accelerated senescence sustained by growth arrest and increased senescence-associated b-galactosidase activity. Increased p16INK4a expression, in the absence of telomere shortening, indicates that premature PT-ECFC aging results from stress-induced senescence. SIRT1 level, a nicotinamide adenine dinucleotidedependent deacetylase with anti-aging activities, is dramatically decreased in PT-ECFCs and correlated with gestational age. SIRT1 deficiency is subsequent to epigenetic silencing of its promoter. Transient SIRT1 overexpression or chemical induction by resveratrol treatment reverses senescence phenotype, and rescues in vitro PT-ECFC angiogenic defect in a SIRT1-dependent manner. SIRT1 overexpression also restores PT-ECFC capacity for neovessel formation in vivo. We thus demonstrate that decreased expression of SIRT1 drives accelerated senescence of PT-ECFCs, and acts as a critical determinant of the PT-ECFC angiogenic defect. These findings lay new grounds for understanding the increased cardiovascular risk in individuals born prematurely and open perspectives for therapeutic strategy. (Blood. 2014;123(13):2116-2126

Germ-line alterations in BRCA1 are associated with an increased susceptibility to breast and ovarian cancer. BRCA1 is a 220-kDa protein that contains a tandem of two BRCA1 C-Terminal (BRCT) domains. Among missense and nonsense BRCA1 mutations responsible for family breast cancer, some are located into the BRCT tandem of BRCA1 coding sequence. In an attempt to understand how BRCT is critical for BRCA1 function, we search for partners of this BRCT tandem of BRCA1. Using a glutathione-S-transferase (GST) pull-down assay with murine cells, we isolated only one major BRCA1-interacting protein, further identified as Acetyl Coenzyme A (CoA) Carboxylase a (ACCA). We showed that this interaction is conserved through murine and human species. We also delineated the minimum interacting region as being the whole tandem of BRCT domains. We demonstrated that BRCA1 interacts in vitro and in vivo with ACCA. This interaction is completely abolished by five distinct germline BRCA1 deleterious mutations affecting the BRCT tandem of BRCA1. Interestingly, ACCA originally known as a rate-limiting enzyme for fatty acids biosynthesis, has been recently shown to be overexpressed in breast cancers and considered as a potential target for anti-neoplastic therapy. Furthermore, our observation is making a bridge between the genetic factors involved in susceptibility to breast and ovarian cancers, and environmental factors such as nutrition considered as key elements in the etiology of those cancers.

CD146 (MUC-18, MCAM) expression on cancer cells correlates with cancer progression and a bad prognosis in several tumors, including melanoma and pancreatic tumors. Deciphering the mechanism mediating the CD146 role in cancer is essential for generating new therapeutic strategies. We found that CD146 expression in cancer cells is associated with a secretion of soluble CD146 (sCD146) that constitutes an active player in tumor development. Indeed, sCD146 induces the overexpression of its binding protein, angiomotin, on both endothelial and cancer cells and promotes both paracrine effects on angiogenesis and autocrine effects on cancer cells proliferation and survival. These last effects are mediated in part through the induction and phosphorylation of c-myc in cancer cells. In mice models xenografted with human CD146-positive melanoma or pancreatic cancer cells, administration of a novel monoclonal antibody specifically targeting sCD146, but not its membrane form, successfully suppresses tumor vascularization and growth. Our findings demonstrate that sCD146 secreted by CD146-positive tumors mediates important pro-angiogenic and pro-tumoral effects. Targeting sCD146 with a novel neutralizing antibody could thus constitute an innovative therapeutic strategy for the treatment of CD146-positive tumors.

CD146 (cluster of differentiation 146) is an adhesion molecule that is expressed by different cells constituting vessels, particularly endothelial cells. The last 30 years of research in this field have shown that CD146 plays a key role in the control of several vessel functions. Three forms of CD146 have been described, including 2 transmembrane isoforms and a soluble protein that is detectable in the plasma. These CD146 forms mediate pleiotropic functions through homophilic and heterophilic interactions with proteins present on surrounding partners. Several studies used neutralizing antibodies, siRNA, or genetically modified mice to demonstrate the involvement of CD146 in the regulation of angiogenesis, vascular permeability, and leukocyte transmigration. In this review, we will focus on the current knowledge of the roles of CD146 in vascular homeostasis and diseases associated with endothelial dysfunction.