Metastasis is the primary cause of cancer morbidity and mortality. The process involves a complex interplay between intrinsic tumor cell properties as well as interactions between cancer cells and multiple microenvironments. The outcome is the development of a nearby or distant discontiguous secondary mass. To successfully disseminate, metastatic cells acquire properties in addition to those necessary to become neoplastic. Heterogeneity in mechanisms involved, routes of dissemination, redundancy of molecular pathways that can be utilized, and the ability to piggyback on the actions of surrounding stromal cells makes defining the hallmarks of metastasis extraordinarily challenging. Nonetheless, this review identifies four distinguishing features that are required: motility and invasion, ability to modulate the secondary site or local microenvironments, plasticity, and ability to colonize secondary tissues. By defining these first principles of metastasis, we provide the means for focusing efforts on the aspects of metastasis that will improve patient outcomes.

Breast cancer metastasis suppressor 1 (BRMS1) is a predominantly nuclear protein that differentially regulates expression of multiple genes, leading to suppression of metastasis without blocking orthotopic tumor growth in multiple human and murine cancer cells of diverse origins. We hypothesized that miR-146 may be involved in the ability of BRMS1 to supress metastasis because miR-146 expression is altered by BRMS1 and because BRMS1 and miR-146 are both associated with decreased signaling through the nuclear factor-KB pathway. BRMS1 significantly up-regulates miR-146a by 6-to 60-fold in metastatic MDA-MB-231 and MDA-MB-435 cells, respectively, and miR-146b by 40-fold in MDA-MB-435 as measured by real-time quantitative reverse transcription-PCR. Transduction of miR-146a or miR-146b into MDA-MB-231 down-regulated expression of epidermal growth factor receptor, inhibited invasion and migration in vitro, and suppressed experimental lung metastasis by 69% and 84%, respectively (mean F SE: empty vector = 39 F 6, miR-146a = 12 F 1, miR-146b = 6 F 1). These results further support the recent notion that modulating the levels of miR-146a or miR-146b could have a therapeutic potential to suppress breast cancer metastasis.

Despite advancements in knowledge from more than a century of metastasis research, the genetic programs and molecular mechanisms required for cancer metastasis are still incompletely understood. Genes that specifically regulate the process of metastasis are useful tools to elucidate molecular mechanisms and may become markers and/or targets for antimetastatic therapy. Recently, several noncoding regulatory RNA genes, microRNA (miRNA), were identified, which play roles in various steps of metastasis, some without obvious roles in tumorigenesis. Understanding how these metastasis-associated miRNA, which we term metastamir, are involved in metastasis will help identify possible biomarkers or targets for the most lethal attribute of cancer: metastasis.

KISS1 secretion was required for multiple organ metastasis suppression and for maintenance of disseminated cells in a dormant state. The absence of GPR54 expression in C8161.9 cells (whose metastatic spread was suppressed by KFM) suggests that metastasis suppression is not mediated through this receptor. The results imply the existence of another KISS1 receptor and/or paracrine signaling. The findings raise the possibility that soluble KISS1, kisspeptins, or mimetics could be used to maintain tumor dormancy, rendering treatment of already disseminated tumor cells (i.e., micrometastases) a legitimate target.

Heparanase acts as a master regulator of the aggressive tumor phenotype in part by enhancing expression of proteins known to drive tumor progression (e.g. VEGF, MMP-9, hepatocyte growth factor (HGF), and RANKL). However, the mechanism whereby this enzyme regulates gene expression remains unknown. We previously reported that elevation of heparanase levels in myeloma cells causes a dramatic reduction in the amount of syndecan-1 in the nucleus. Because syndecan-1 has heparan sulfate chains and because exogenous heparan sulfate has been shown to inhibit the activity of histone acetyltransferase (HAT) enzymes in vitro, we hypothesized that the reduction in nuclear syndecan-1 in cells expressing high levels of heparanase would result in increased HAT activity leading to stimulation of protein transcription. We found that myeloma cells or tumors expressing high levels of heparanase and low levels of nuclear syndecan-1 had significantly higher levels of HAT activity when compared with cells or tumors expressing low levels of heparanase. High levels of HAT activity in heparanase-high cells were blocked by SST0001, an inhibitor of heparanase. Restoration of high syndecan-1 levels in heparanase-high cells diminished nuclear HAT activity, establishing syndecan-1 as a potent inhibitor of HAT. Exposure of heparanase-high cells to anacardic acid, an inhibitor of HAT activity, significantly suppressed their expression of VEGF and MMP-9, two genes known to be up-regulated following elevation of heparanase. These results reveal a novel mechanistic pathway driven by heparanase expression, which leads to decreased nuclear syndecan-1, increased HAT activity, and up-regulation of transcription of multiple genes that drive an aggressive tumor phenotype.Heparanase, an endoglycosidase that cleaves heparan sulfate, is up-regulated in many cancers where it promotes tumor growth, angiogenesis, and metastasis (1, 2). High levels of heparanase in cancer patients are associated with shorter postoperative survival time compared with patients with low levels of heparanase (1). Although some of the tumor promoting effects of heparanase can be attributed to its ability to remodel the extracellular matrix barrier by cleaving heparan sulfate, heparanase is also known to regulate cell signaling and gene transcription (1, 3-5). Elevation of heparanase levels in myeloma cells, either by transfection of cells or by addition of recombinant active heparanase enzyme to cells, up-regulates expression of MMP-9, VEGF, HGF, 2 and RANKL, which together drive an aggressive tumor phenotype (6 -9). Although the mechanism whereby heparanase drives gene expression remains unknown, the enzyme is present and active in the nucleus where it could act locally to regulate gene expression (10).Acetylation of the N-terminal tails of histones by histone acetyltransferase enzymes has been known for many years to be a process correlating with transcriptional activation (11)(12)(13)(14). This process is balanced by the activity of histone deacetylases (HDACs), which selectivel...

Breast cancer metastasis suppressor 1 (BRMS1) inhibits formation of macroscopic lung metastases in breast, ovary, and melanoma xenograft models. Because it is unclear which step(s) of the metastatic cascade are affected by BRMS1, the major aim of this study was to determine when and how BRMS1 acts to suppress metastasis. We also examined whether BRMS1 expression globally blocks metastasis or selectively inhibits metastatic outgrowths in specific tissues. The overwhelming majority of morbidity and mortality for patients with cancer is associated with metastatic disease. In breast cancer, metastases are relatively widely distributed, with the most common sites being bone, regional lymph nodes, lung, liver, and brain.1 Significant improvements in survival and quality of life have been realized over several decades due to earlier detection and more effective treatment of metastases. However, there is still much room for improvement.A relatively new class of molecules, metastasis suppressors, hold promise for providing new avenues for therapeutic intervention. Metastasis suppressors are defined by the ability to prevent metastasis without blocking orthotopic tumor growth.2-5 Most have been discovered in the past decade, but their mechanisms of action remain largely unexplained.Breast cancer metastasis suppressor 1 (BRMS1) was functionally defined by its ability to block lung and regional lymph node metastases in experimental breast, melanoma, and ovarian models.6 -10 Decreased BRMS1 protein expression in human breast carcinomas has been correlated with reduced disease-free survival when stratified by loss of estrogen or progesterone receptor or

The BRMS1 metastasis suppressor interacts with the protein AT-rich interactive domain 4A (ARID4A, RBBP1) as part of SIN3⅐histone deacetylase chromatin remodeling complexes. These transcriptional co-repressors regulate diverse cell phenotypes depending upon complex composition. To define BRMS1 complexes and their roles in metastasis suppression, we generated BRMS1 mutants (BRMS1 mut ) and mapped ARID4A interactions. BRMS1L174D disrupted direct interaction with ARID4A in yeast two-hybrid genetic screens but retained an indirect association with ARID4A in MDA-MB-231 and -435 human breast cancer cell lines by co-immunoprecipitation. Deletion of the first coiled-coil domain (BRMS1 ⌬CC1 ) did not disrupt direct interaction in yeast two-hybrid screens but did prevent association by co-immunoprecipitation. These results suggest altered complex composition with BRMS1 mut . Although basal transcription repression was impaired and the pro-metastatic protein osteopontin was differentially down-regulated by BRMS1 L174D and BRMS1 ⌬CC1 , both down-regulated the epidermal growth factor receptor and suppressed metastasis in MDA-MB-231 and -435 breast cancer xenograft models. We conclude that BRMS1 mut , which modifies the composition of a SIN3⅐histone deacetylase chromatin remodeling complex, leads to altered gene expression profiles. Because metastasis requires the coordinate expression of multiple genes, down-regulation of at least one important gene, such as the epidermal growth factor receptor, had the ability to suppress metastasis. Understanding which interactions are necessary for particular biochemical/ cellular functions may prove important for future strategies targeting metastasis.The ability of a cancer cell to complete all steps of the metastatic cascade requires diverse tumor-host interactions that are dependent on the coordinate expression of specific genes both intrinsically and extrinsically (1-3). The metastasis suppressor BRMS1 3 has been shown to regulate the expression of multiple genes leading to the suppression of metastasis in multiple model systems, including human breast carcinoma (4, 5), melanoma (6), and ovarian carcinoma (7), without preventing orthotopic tumor growth. Specifically, down-regulation of the pro-metastatic genes osteopontin (OPN) and urokinase-type plasminogen activator has been linked to BRMS1 expression (8, 9). Gap junctional intercellular communication is restored by BRMS1 through a change in connexin expression (10). Microarray and proteomic analyses have also been performed showing multiple changes in gene and protein expression when BRMS1 was introduced (11-13). Clinically, loss of BRMS1 protein has been correlated with progesterone receptor expression and inversely correlated with HER2 expression in breast cancer patients (14).BRMS1 has been proposed to regulate transcription of genes by interaction with a large SIN3⅐HDAC chromatin remodeling complex through interaction with the protein AT-rich interactive domain 4A (ARID4A) that suppresses basal transcription in vivo using a Gal...

Matrix metalloproteinases (MMPs, 1 matrixins) are believed to participate in angiogenesis, embryonic development, morphogenesis, reproduction, tissue resorption and remodeling, and tumor growth, progression, invasion, and metastasis through breakdown of the extracellular matrix, cell surface proteins, and processing growth factors, cytokines, and chemokines (1-3). Recently, human MMP-26 (endometase/matrilysin 2) was identified and its mRNA expression was detected in normal tissues of the human uterus and placenta, and in many types of malignant tumors (4 -7). Characterization of the MMP-26 promoter suggests that this proteinase may be expressed in cancer cells of epithelial origin (8). MMP-26 may play an important role in human prostate and breast cancer invasion (9 -10).MMP-26 cleaves type I gelatin, ␣ 1 -proteinase inhibitor, fibrinogen, fibronectin, vitronectin, type IV collagen, and insulin-like growth factor binding protein-1 (4, 7, 11). Studies of MMP-26 indicate that it has substrate specificity similar to other MMPs, with the exception of a preference for Ile at the P 2 and P 2 Ј positions, for small residues at the P 3 Ј and P 4 Ј positions, and Lys at the P 4 position (11). MMP-26 also hydrolyzes several synthetic fluorogenic peptide substrates designed for stromelysin-1, gelatinases, collagenases, and tumor necrosis factor-␣ converting enzyme (4, 11). According to these peptide substrate studies, MMP-26 may be capable of cleaving a broad range of substrates, although it has less catalytic efficiency than other MMPs.X-ray crystal structures of MMPs illustrate that overall topology and secondary structures are conserved (12-18). The S 1 Ј pocket, a hydrophobic pocket of variable depth, is a well defined substrate P 1 Ј-binding site in MMPs. Three types of S 1 Ј pockets can be distinguished from the available structures of . One type is a shallow pocket, as found in MMP-1 (human fibroblast collagenase; 13) and 16), where the pockets are limited by the side chains of Arg and Tyr, respectively, crossing the pockets. Many of the structurally known MMPs possess Leu at the corresponding site, and its side chain forms the top of the pocket rather than crossing the pocket. These Leu-containing MMPs may be further classified as deep and intermediate S 1 Ј pocket MMPs. A deep, tunnel-like pocket is found in MMP-3 (stromelysin-1; 12), MMP-12 (metalloelastase; 17), and MMP-14 (MT1-MMP; 21), whereas MMP-2 (gelatinase A; 22), MMP-8 (human neutrophil collagenase; 15), and MMP-9 (gelatinase B; 23) possess an intermedi-