NM23 and metastasis suppressor genes: update

Metastatic dissemination represents a leading cause of death in cancer patients. Elucidating the mechanisms of the metastatic process is therefore essential to control it. Since 1988, when the NME (NM23) gene was discovered, several genes specifically suppressing the metastatic potential of tumor cells, have been identified. These metastasis suppressor genes, which exhibit a reduced expression in metastatic tumor cells, are defined by their capacity to suppress metastatic dissemination in vivo without inhibiting primary tumor growth when transfected into metastatic cell lines and injected into experimental animals. Their decreased expression in a subset of human tumor cohorts is associated with a high metastatic potential, thus confirming the data obtained in experimental models. Most of these genes affect key signal transduction pathways, including mitogen-activated protein kinases, Rho-GTPases and G-protein-coupled receptors. These signaling categories control cell-cell and cell-matrix interactions, which are important in monitoring adhesion, invasion and migration properties of metastatic tumor cells. Reduced expression of metastasis suppressor genes is most often due to epigenetic mechanisms, suggesting that their re-expression could constitute a new anti-metastatic therapy. In this paper, we review the literature on metastasis suppressor genes, with a particular focus on NM23.     

Pathological angiogenesis facilitates tumor cell dissemination and metastasis

Clinically detectable metastases represent an ultimate consequence of the metastatic cascade that consists of distinct processes including tumor cell invasion, dissemination, metastatic niche formation, and re-growth into a detectable metastatic mass. Although angiogenesis is known to promote tumor growth, its role in facilitating early events of the metastatic cascade remain poorly understood. We have recently developed a zebrafish tumor model that enables us to study involvement of pathological angiogenesis in tumor invasion, dissemination and metastasis. This non-invasive in vivo model allows detection of single malignant cell dissemination under both normoxia and hypoxia. Further, hypoxia-induced VEGF significantly facilitates tumor cell invasion and dissemination. These findings demonstrate that VEGF-induced pathological angiogenesis is essential for tumor dissemination and further corroborates potentially beneficial effects of clinically ongoing anti-VEGF drugs for the treatment of various malignancies.     

Metastasis prevention: How to catch metastatic seeds

Despite considerable advances in the evolution of anticancer therapies, metastasis still remains the main cause of cancer mortality. Therefore, current strategies for cancer cure should be redirected towards prevention of metastasis. Targeting metastatic pathways represents a promising therapeutic opportunity aimed at obstructing tumor cell dissemination and metastatic colonization. In this review, we focus on preclinical studies and clinical trials over the last five years that showed high efficacy in suppressing metastasis through targeting lymph node dissemination, tumor cell extravasation, reactive oxygen species, pre-metastatic niche, exosome machinery, and dormancy.     

The role of the tumor microenvironment in tumor cell intravasation and dissemination

Metastasis, a process that requires tumor cell dissemination followed by tumor growth, is the primary cause of death in cancer patients. An essential step of tumor cell dissemination is intravasation, a process by which tumor cells cross the blood vessel endothelium and disseminate to distant sites. Studying this process is of utmost importance given that intravasation in the primary tumor, as well as the secondary and tertiary metastases, is the key step in the systemic spread of tumor cells, and that this process continues even after removal of the primary tumor. High-resolution intravital imaging of the tumor microenvironment of breast carcinoma has revealed that tumor cell intravasation exclusively occurs at doorways, termed “Tumor MicroEnvironment of Metastasis” (TMEM), composed of three different cell types: a Tie2^high/VEGF^high perivascular macrophage, a Mena overexpressing tumor cell, and an endothelial cell, all in direct contact. In this review article, we discuss the interactions between these cell types, the subsequent signaling events which lead to tumor cell intravasation, and the role of invadopodia in supporting tumor cell invasion and dissemination. We end our review by discussing how the knowledge acquired from the use of intravital imaging is now leading to new clinical trials targeting tumor cell dissemination and preventing metastatic progression.     

Gene expression sigantures, cancer cell evolution and metastatic progression

Global gene expression profiling and molecular cytogenetics for single cells provide new opportunities to resolve important questions, such as the mechanisms of tumor cell selection during cancer progression. While gene expression analyses show that metastatic progression correlates with the deregulation of certain gene sets in the primary tumor, the direct analysis of disseminated cancer cells, the putative precursors of metachronous metastases, suggests that dissemination is a very early event in the genetic development of human cancers.     

The Biochemistry of Cancer Dissemination

Abstract

The progression of a tumor cell from one of benign delimited proliferation to invasive and metastatic growth is the major cause of poor clinical outcome of cancer patients. Recent research has revealed that this complex process requires many components for successful dissemination and growth of the tumor cell at secondary sites. These include angiogenesis, enhanced extracellular matrix degradation via tumor and host-secreted proteases, tumor cell migration, and modulation of tumor cell adhesion. Each individual component is multifaceted and is discussed within this review with respect to historical and recent findings. The identification of components and their interrelationship have yielded new therapeutic targets leading to the development of agents that may prove effective in the treatment of cancer and its metastatic progression.     

The Calpain/Calpastatin System Has Opposing Roles in Growth and Metastatic Dissemination of Melanoma

Conventional calpains are ubiquitous cysteine proteases whose activity is promoted by calcium signaling and specifically limited by calpastatin. Calpain expression has been shown to be increased in human malignant cells, but the contribution of the calpain/calpastatin system in tumorigenesis remains unclear. It may play an important role in tumor cells themselves (cell growth, migration, and a contrario cell death) and/or in tumor niche (tissue infiltration by immune cells, neo-angiogenesis). In this study, we have used a mouse model of melanoma as a tool to gain further understanding of the role of calpains in tumor progression. To determine the respective importance of each target, we overexpressed calpastatin in tumor and/or host in isolation. Our data demonstrate that calpain inhibition in both tumor and host blunts tumor growth, while paradoxically increasing metastatic dissemination to regional lymph nodes. Specifically, calpain inhibition in melanoma cells limits tumor growth in vitro and in vivo but increases dissemination by amplifying cell resistance to apoptosis and accelerating migration process. Meanwhile, calpain inhibition restricted to host cells blunts tumor infiltration by immune cells and angiogenesis required for antitumor immunity, allowing tumor cells to escape tumor niche and disseminate. The development of highly specific calpain inhibitors with potential medical applications in cancer should take into account the opposing roles of the calpain/calpastatin system in initial tumor growth and subsequent metastatic dissemination.     

Cell adhesion in tumor invasion and metastasis: loss of the glue is not enough.

Tumor cells often show a decrease in cell-cell and/or cell-matrix adhesion. An increasing body of evidence indicates that this reduction in cell adhesion correlates with tumor invasion and metastasis. Two main groups of adhesion molecules, cadherins and CAMs, have been implicated in tumor malignancy. However, the specific role that these proteins play in the context of tumor progression remains to be elucidated. In this review, we discuss recent data pointing to a causal relationship between the loss of cell adhesion molecules and tumor progression. In addition, the direct involvement of these molecules in specific signal transduction pathways will be considered, with particular emphasis on the alterations of such pathways in transformed cells. Finally, we review recent observations on the molecular mechanisms underlying metastatic dissemination. In many cases, spreading of tumor cells from the primary site to distant organs has been characterized as an active process involving the loss of cell-cell adhesion and gain of invasive properties. On the other hand, various examples of metastases exhibiting a relatively benign (i.e. not invasive) phenotype have been reported. Together with our recent results on a mouse tumor model, these findings indicate that 'passive' metastatic dissemination can occur, in particular as a consequence of impaired cell-matrix adhesion and of tumor tissue disaggregation.     

Cell adhesion in tumor invasion and metastasis: loss of the glue is not enough

Tumor cells often show a decrease in cell–cell and/or cell–matrix adhesion. An increasing body of evidence indicates that this reduction in cell adhesion correlates with tumor invasion and metastasis. Two main groups of adhesion molecules, cadherins and CAMs, have been implicated in tumor malignancy. However, the specific role that these proteins play in the context of tumor progression remains to be elucidated. In this review, we discuss recent data pointing to a causal relationship between the loss of cell adhesion molecules and tumor progression. In addition, the direct involvement of these molecules in specific signal transduction pathways will be considered, with particular emphasis on the alterations of such pathways in transformed cells. Finally, we review recent observations on the molecular mechanisms underlying metastatic dissemination. In many cases, spreading of tumor cells from the primary site to distant organs has been characterized as an active process involving the loss of cell–cell adhesion and gain of invasive properties. On the other hand, various examples of metastases exhibiting a relatively benign (i.e. not invasive) phenotype have been reported. Together with our recent results on a mouse tumor model, these findings indicate that ‘passive’ metastatic dissemination can occur, in particular as a consequence of impaired cell–matrix adhesion and of tumor tissue disaggregation.     

Nm23/NDP kinases in hepatocellular carcinoma

One of the most aggressive cancers is hepatocellular carcinoma, which is associated with a very poor patient outcome due to a high recurrence rate and metastatic spread. NM23, the first metastasis suppressor gene to be identified, has been widely studied in human cancers. However, conflicting results have been obtained depending on the tumor type and the evaluation protocol. The current knowledge of NM23 as a diagnostic and/or prognostic marker in hepatocellular carcinoma is reviewed herein. Most studies demonstrate an inverse association between the expression of NM23-H1 and the metastatic potential, which is not observed with the closely related NM23-H2 isoform. Transfection of metastatic hepatoma cells with NM23 reduced their metastatic potential, as for other tumor cell lines. The demonstration of a causative role of NM23 in metastatic dissemination in a mouse model of hepatocarcinoma suggests that hepatocarcinoma-derived cells could be good models for the analysis of the molecular mechanisms involved in NM23 action.     

Epithelial–mesenchymal transition markers expressed in circulating tumor cells in hepatocellular carcinoma patients with different stages of disease

The presence of circulating tumor cells (CTCs) in peripheral blood is associated with metastasis and prognosis in hepatocellular carcinoma (HCC) patients. The epithelial–mesenchymal transition (EMT) has a pivotal role in tumor invasion and dissemination. To identify more sensitive biomarkers for evaluating metastasis and prognosis, we investigated the expression of EMT markers, including vimentin, twist, ZEB1, ZEB2, snail, slug and E-cadherin in CTCs, primary HCC tumors and adjacent non-tumoral liver tissues. After isolating viable CTCs from the peripheral blood of HCC patients using asialoglycoprotein receptors (ASGPRs), the CTCs were identified with immunofluorescence staining. CTCs were detected in the peripheral blood obtained from 46 of 60 (76.7%) HCC patients. Triple-immunofluorescence staining showed that twist and vimentin expression could be detected in CTCs obtained from 39 (84.8%) and 37 (80.4%) of the 46 patients, respectively. The expression of both twist and vimentin in CTCs was significantly correlated with portal vein tumor thrombus. Coexpression of twist and vimentin in CTCs could be detected in 32 (69.6%) of the 46 patients and was highly correlated with portal vein tumor thrombus, TNM classification and tumor size. Quantitative fluorescence western blot analysis revealed that the expression levels of E-cadherin, vimentin and twist in HCC tumors were significantly associated with the positivity of isolated CTCs (P=0.013, P=0.012, P=0.009, respectively). However, there was no significant difference in ZEB1, ZEB2, snail and slug expression levels in CTCs, primary HCC tumors and adjacent non-tumoral liver tissues across samples with regard to the clinicopathological parameters. Our results demonstrate that the EMT has a role in promoting the blood-borne dissemination of primary HCC cells, and the twist and vimentin expression levels in CTCs could serve as promising biomarkers for evaluating metastasis and prognosis in HCC patients.     

Age and metastasis – How age influences metastatic spread in cancer. Colorectal cancer as a model

Metastasis is the major cause of death in cancer patients. Whereas colorectal cancer (CRC) incidence increases with age, metastatic spread seems to decline. Furthermore, the epidemiology of CRC is changing. There is an increase in CRC incidence in the young, presenting at an advanced stage with higher likelihood of synchronous or metachronous metastases, and a decline in CRC incidence and metastatic spread in the oldest-old. Emerging data suggest that age-related changes with regard to tumor biology (e.g. genomic instability), the tumor microenvironment (e.g. inflammaging) and the immune system (e.g. immunosenescence), complemented by interaction between the genome and exposome might contribute to the observed metastatic patterns. As aging is a key prognostic factor, this highlights the need for further studies investigating age-related patterns and underlying mechanisms of tumor growth and dissemination. Eventually, this might allow for better risk stratification, refinement of screening strategies and follow-up care as well as therapies tailored to reflect patient age and that might possibly target responsible biomarkers in a precision medicine approach. This review aims to discuss the influence of aging on metastatic spread in colorectal cancer and elucidate underlying mechanisms responsible for the observed metastatic patterns.     

间质-上皮转化在肿瘤转移中的作用

肿瘤转移是一个多步骤、多因素参与的复杂过程,是目前临床上绝大多数肿瘤患者的致死因素.上皮-间质转化(epithelial-mesenchymal transition, EMT)过程已被证实可促使肿瘤细胞发生转移.近年来许多研究表明,间质-上皮转化(mesenchymal-epithelial transition, MET)即EMT的逆过程,与肿瘤也密切相关,特别是肿瘤转移即形成继发性的肿瘤转移灶.深入研究肿瘤MET有望为肿瘤转移的预防和诊治提供新思路.     

Involvement of CXCR4 Chemokine Receptor in Metastastic HER2-Positive Esophageal Cancer

A functional linkage of the structurally unrelated receptors HER2 and CXCR4 has been suggested for breast cancer but has not been evaluated for esophageal carcinoma. The inhibition of HER2 leads to a reduction of primary tumor growth and metastases in an orthotopic model of esophageal carcinoma. The chemokine receptor CXCR4 has been implicated in metastatic dissemination of various tumors and correlates with poor survival in esophageal carcinoma. The aim of this study was to investigate a correlation between the expression levels of HER2 and CXCR4 and to evaluate the involvemnent of CXCR4-expression in HER2-positive esophageal carcinoma. The effects of HER2-inhibition with trastuzumab and of CXCR4-inhibition with AMD3100 on primary tumor growth, metastatic homing, and receptor expression were evaluated in vitro and in an orthotopic model of metastatic esophageal carcinoma using MRI for imaging. The clinical relevance of HER2- and CXCR4-expression was examined in esophageal carcinoma patients. A significant correlation of HER2- and CXCR4-expression in primary tumor and metastases exists in the orthotopic model. Trastuzumab and AMD3100 treatment led to a significant reduction of primary tumor growth, metastases and micrometastases. HER2-expression was significantly elevated under AMD3100 treatment in the primary tumor and particularly in the metastases. The positive correlation between HER2- and CXCR4-expression was validated in esophageal cancer patients. The correlation of CXCR4- and HER2-expression and the elevation of HER2-expression and reduction of metastases through CXCR4-inhibition suggest a possible functional linkage and a role in tumor dissemination in HER2-positive esophageal carcinoma.     

The immunobiology of metastatic processes: analysis of NK sensitivity and the metastatic potential of H-2 gene transfected fibrosarcoma cells

The transformation of a potentially neoplastic cell into an autonomous highly malignant and metastatic tumor cell involves a multifactorial cascade of events. This will eventually lead not only to the emergence of a tumor cell with an unlimited potential of replication, but more important will contribute to its ability to ignore and evade homeostatic immune and nonimmune regulatory mechanisms. Specifically, those mechanisms which may restrict and direct its growth, dissemination, patterns of differentiation and interaction with the cellular and humoral factors comprising its environment. In the present studies we have investigated the contribution of three major factors which may be the cause or result of alterations at the level of the cell membrane: MHC encoded antigen expression, susceptibility to the cytolytic activity of NK cells and enhanced expression of the c-K-ras proto-oncogene, as to their development of the metastatic capacity of a malignant cell. To address these questions we used metastatic (IE7) and nonmetastatic (IC9) variants of the murine 3-methylcholanthrene-induced T-10 fibrosarcoma. Using this system, the following major conceptually important observations were made: (A) The restoration by transfection of the expression of membrane associated H-2K encoded glycoproteins abrogates the metastatic capacity of the highly metastatic tumor cell clone, IE7, irrespective of the degree of susceptibility to NK or c-K-ras oncogene expression.(ABSTRACT TRUNCATED AT 250 WORDS)