Functional Analysis of Matrix Metalloproteinases and Tissue Inhibitors of Metalloproteinases Differentially Expressed by Variants of Human HT-1080 Fibrosarcoma Exhibiting High and Low Levels of Intravasation and Metastasis (original) (raw)
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Cancer Research, 2005
The human tumor/chick embryo model involving grafting of human HT-1080 fibrosarcoma cells on the chorioallantoic membrane was used in conjunction with quantitative real-time Alu PCR to select in vivo a pair of isogenic cell lines (HT-hi/diss and HT-lo/diss), dramatically differing in their ability to disseminate from the primary tumor (i.e., intravasate into the chorioallantoic membrane vasculature and metastasize to the lungs). During an immunohistochemical time course study, HT-hi/diss cells were sequentially visualized having escaped from the primary tumors, engaged with the blood vessels, and eventually observed inside the chorioallantoic membrane capillaries, thus reflecting early intravasating events. In contrast, HT-lo/diss cells seemed restricted to their primary tumor. Importantly, after i.v. inoculation, both variants arrested, extravasated, and proliferated in host tissues with similar efficiencies, highlighting that the observed earlier events at the periphery of the primary tumor could account for their differential dissemination. In a mechanistic probing of these events, we determined that HT-hi/diss intravasation was sensitive to a broad-range matrix metalloproteinase (MMP) inhibitor. To analyze the possible role of individual MMPs, membrane-bound MMP-14 and secreted MMP-9 were individually down-regulated in HT-hi/diss cells with their corresponding small interfering RNAs. Despite efficient down-regulation of MMP-14, neither intravasation nor metastasis of HT-hi/diss cells was affected significantly. However, a substantial down-regulation of MMP-9 was accompanied by a surprising 3-fold increase in intravasation and metastasis. The results emphasize a rising awareness that targeting certain MMPs might result in an enhanced malignancy, exemplified herein at the intravasation level as this step of the metastatic cascade is dissected and quantified.
Journal of Cell Biology, 2004
A independently of plasminogen, the gelatinase A/TIMP-2 axis, gelatinase B, collagenase-3, collagenase-2, or stromelysin-1. In contrast, deleting or suppressing expression of the membrane-tethered MMP, MT1-MMP, in fibroblasts or tumor cells results in a loss of collagenolytic and invasive activity in vitro or in vivo. Thus, MT1-MMP serves as the major cell-associated proteinase necessary to confer normal or neoplastic cells with invasive activity.
Tumor cell traffic through the extracellular matrix is controlled by the membrane-anchored …
Journal of Cell Biology
A independently of plasminogen, the gelatinase A/TIMP-2 axis, gelatinase B, collagenase-3, collagenase-2, or stromelysin-1. In contrast, deleting or suppressing expression of the membrane-tethered MMP, MT1-MMP, in fibroblasts or tumor cells results in a loss of collagenolytic and invasive activity in vitro or in vivo. Thus, MT1-MMP serves as the major cell-associated proteinase necessary to confer normal or neoplastic cells with invasive activity.
Membrane type-1 matrix metalloproteinase and TIMP-2 in tumor angiogenesis
Matrix Biology, 2003
The matrix metalloproteinases (MMPs) constitute a multigene family of over 23 secreted and cell-surface associated enzymes that cleave or degrade various pericellular substrates. In addition to virtually all extracellular matrix (ECM) compounds, their targets include other proteinases, chemotactic molecules, latent growth factors, growth factor-binding proteins and cell surface molecules. The MMP activity is controlled by the physiological tissue inhibitors of MMPs (TIMPs). There is much evidence that MMPs and their inhibitors play a key role during extracellular remodeling in physiological situations and in cancer progression. They have other functions that promoting tumor invasion. Indeed, they regulate early stages of tumor progression such as tumor growth and angiogenesis. Membrane type MMPs (MT-MMPs) constitute a new subset of cell surface-associated MMPs. The present review will focus on MT1-MMP which plays a major role at least, in the ECM remodeling, directly by degrading several of its components, and indirectly by activating pro-MMP2. As our knowledge on the field of MT1-MMP biology has grown, the unforeseen complexities of this enzyme and its interaction with its inhibitor TIMP-2 have emerged, often revealing unexpected mechanisms of action.
Matrix Metalloproteinase 1: Role in Sarcoma Biology
PLOS One, 2010
In carcinomas stromal cells participate in cancer progression by producing proteases such as MMPs. The expression MMP1 is a prognostic factor in human chondrosarcoma, however the role in tumor progression is unknown. Laser capture microdissection and In Situ hybridization were used to determine cellular origin of MMP1 in human sarcomas. A xenogenic model of tumor progression was then used and mice were divided in two groups: each harboring either the control or a stably MMP1 silenced cell line. Animals were sacrificed; the neovascularization, primary tumor volumes, and metastatic burden were assessed. LCM and RNA-ISH analysis revealed MMP1 expression was predominantly localized to the tumor cells in all samples of sarcoma (p = 0.05). The percentage lung metastatic volume at 5 weeks (p = 0.08) and number of spontaneous deaths secondary to systemic tumor burden were lower in MMP1 silenced cell bearing mice. Interestingly, this group also demonstrated a larger primary tumor size (p,0.04) and increased angiogenesis (p,0.01). These findings were found to be consistent when experiment was repeated using a second independent MMP1 silencing sequence. Prior clinical trials employing MMP1 inhibitors failed because of a poor understanding of the role of MMPs in tumor progression. The current findings indicating tumor cell production of MMP1 by sarcoma cells is novel and highlights the fundamental differences in MMP biology between carcinomas and sarcomas. The results also emphasize the complex roles of MMP in tumor progression of sarcomas. Not only does metastasis seem to be affected by MMP1 silencing, but also local tumor growth and angiogenesis are affected inversely.
International Journal of Cancer, 2002
Membrane type 1 matrix metalloproteinase (MT1-MMP) is frequently expressed by cancer cells and is believed to play an important role in cancer cell invasion and metastasis. However, little is known about the role of MT1-MMP in mediating invasiveness of cervical cancer cells. In this study, we examined MT1-MMP expression in 58 primary human cervical tissue specimens, including normal cervix, low-grade squamous intraepithelial lesions (LSIL), high-grade SILs (HSIL), and invasive carcinomas. We also evaluated MT1-MMP, MMP-2, and tissue inhibitor of metalloproteinase-2 expression in several cervical cancer-derived cell lines, human papillomavirus (HPV)-immortalized keratinocytes, and keratinocytes derived from a LSIL. Using in situ hybridization techniques to study the cervical tissue specimens, we found that MT1-MMP expression increases with cervical tumor progression (Spearman correlation coefficient = 0.66; P < 0.0001, exact test). Specifically, MT1-MMP expression is very low or absent in normal cervix and LSILs, is readily detectable in HSILs, and is very strongly expressed in nearly all invasive carcinomas. Most but not all cervical cancerderived cell lines also expressed significant levels of MT1-MMP and MMP-2. Constitutive expression of exogenous MT1-MMP in cervical carcinoma-derived cells and HPVimmortalized keratinocytes with low endogenous levels of MT1-MMP induced invasiveness in collagen I, but this effect was not observed in LSIL-derived keratinocytes. Our results show that MT1-MMP is a key enzyme mediating cervical cancer progression. However, MT1-MMP alone is not always sufficient for inducing keratinocyte invasiveness at least in the collagen I invasion assay used in this study. Further studies of gene expression in preinvasive and invasive cervical cancers should assist with identification of additional critical factors mediating cervical cancer progression.
Cancer Research, 2004
(TIMPs) regulate the degradation of extracellular matrix components and play important roles in the progression of select neoplastic processes. The locally invasive soft tissue tumor, aggressive fibromatosis (also called desmoid tumor), is caused by mutations resulting in -catenin-mediated T-cell factor (tcf)-dependent transcriptional activity. Because -catenin can regulate MMP expression, we investigated the expression of several MMPs and TIMPs in aggressive fibromatosis tumors that develop in Apc؉/Apc1638N mice. Mmp-3 and Timp-1 were differentially regulated (5-fold and 0.5-fold, respectively) in tumors compared with normal fibrous tissue. Conditioned media from tumor cells showed an increased ability to degrade collagen, and inhibition of MMPs using GM6001 decreased the ability of the tumor cells to invade through Matrigel. Both the treatment of Apc/Apc1638N mice with GM6001 or crossing with a transgenic mouse that overexpresses Timp-1 resulted in a significant reduction in tumor volume. Surprisingly, overexpression of Timp-1 also resulted in a 50% increase in tumor number. Although TIMP-1 can induce growth stimulatory effects in some cell types, we found no difference in proliferation or apoptosis rate in cells from tumors that developed in the Timp-1-transgenic mice compared with mice that did not express the Timp-1 transgene, suggesting that TIMP-1 promotes aggressive fibromatosis tumor formation through an alternate mechanism. These data suggest that MMPs play a crucial role in regulating the invasiveness of mesenchymal cells and in modulating aggressive fibromatosis tumor progression. Because this is a locally invasive tumor, MMP inhibition could slow tumor growth and may prove to be an effective adjuvant therapy.
Cancer Research, 2005
Increased matrix metalloproteinase-1 (MMP-1) expression is associated with advanced stages of breast cancer and may be a predictive marker for the development of invasive disease. In this report, we used short hairpin RNA (shRNA) molecules to investigate whether MMP-1 production in MDA-231 breast cancer cells contributed to the degradation of a collagen matrix or tumor formation in nude mice. We created two groups of MDA-231 cell lines. MDA-231 cells containing a vector producing shRNA specific for MMP-1 had a >90% decrease in MMP-1 mRNA and protein compared with cells containing an empty vector, and blocking MMP-1 expression inhibited the in vitro collagenolytic activity of MDA-231 cells. When the cells were injected into the mammary fat pad, there was no difference in the frequency of tumor formation in mice. However, the average tumor size was larger in mice injected with cells containing the empty vector (1,216 ± 334 mm3) than in mice injected with cells expressing the MMP-1 ...
Molecular Cancer Therapeutics, 2005
Membrane type 1 matrix metalloproteinase (MT1-MMP) is a potent modulator of the pericellular environment and promotes tumor cell invasion and proliferation in many types of tumor. The activation of proMMP-2 and processing of collagen I by MT1-MMP have been thought to be important for its tumor-promoting function. These activities can be inhibited by mutant forms of MT1-MMP lacking the catalytic domain. However, the effect of such dominantnegative mutants has never been evaluated in vivo. Various mutants lacking the catalytic domain (dCAT) were prepared and confirmed to inhibit MT1-MMP activity in human fibrosarcoma HT1080 cells, and tumor cells expressing these mutants were implanted s.c. into nude mice to monitor tumor formation. Only the membraneanchored form of a dCAT construct through the transmembrane domain [dCAT(1)] showed potent antitumor activity not only in HT1080 cells but also in gastric carcinoma MKN28 and MKN45 cells expressing MT1-MMP. A soluble form of dCAT lacking the transmembrane domain did not show such activity. The expression of dCAT(1) in MKN28 or MKN45 further prevented the metastatic spread of tumor cells into the peritoneal cavity; however, dCAT(1) showed no effect against TMK-1, another gastric carcinoma cell line expressing no MT1-MMP. It is of note that the tumorigenicity of TMK-1 cells enhanced by MT1-MMP overexpression was, in turn, canceled by the additional expression of dCAT(1). Thus, MT1-MMP expressed in tumor cells seems to play a pivotal role in tumor growth in mice. The results also suggest new possibilities to abrogate the tumorpromoting function of MT1-MMP other than the conventional protease inhibitor -based approach. [Mol Cancer Ther