Inhibition of Angiogenesis In Vivo byets-1 Antisense Oligonucleotides—Inhibition of Ets-1 Transcription Factor Expression by the Antibiotic Fumagillin

Wernert, Nicolas; Stanjek, A.; Kiriakidis, Serafim; Hügel, A; Jha, Hem Chandra; Mazitschek, Ralph; Giannis, Athanassios

doi:10.1002/(sici)1521-3773(19991102)38:21<3228::aid-anie3228>3.0.co;2-8

Cited by 69 publications

(45 citation statements)

References 19 publications

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“…Tumour cell proliferation alone is not sufficient for continuous growth of metastases which also requires tumour vascularization (30,56,(67)(68)(69)(70). VEGF is considered the most important factor for tumour vascularization (71).…”

Section: Discussionmentioning

confidence: 99%

Tumour-stroma interactions between metastatic prostate cancer cells and fibroblasts

Kaminski

Hahne²,

Haddouti³

et al. 2006

Int J Mol Med

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Abstract.Previous work has shown the importance of tumourstroma interactions for prostate cancer development at the primary site. The aim of the present study was to find out whether evidence can be found for a tumour-stroma crosstalk also between metastatic prostate cancer cell lines and non-prostatic stromal fibroblasts which are encountered by metastatic cells at most sites. We addressed this issue in cell culture systems using 3 metastatic human prostate cancer cell lines (LnCaP, PC-3 and DU-145) on the one hand, and a human fibroblast line (HFF, human foreskin fibroblasts) on the other. We incubated fibroblasts with tumour cell-and tumour cells with fibroblast-conditioned media and evaluated several parameters important for the establishment of metastases such as cell proliferation, migration and expression of matrix degrading proteases. We also determined in the conditioned media the concentrations of several growth factors and cytokines which might be responsible for the observed effects. We found that media conditioned by all 3 metastatic prostate cancer cell lines stimulated fibroblast proliferation which corresponds to fibrous stroma induction in vivo. DU-145 cell conditioned media induced in fibroblasts expression of mmp-1 mRNA known to be important for tumour invasion. ELISA assays revealed that tumour cells secrete bFGF, PDGF and TNF· known to stimulate fibroblast proliferation and/or MMP-1 expression. Cultivation of DU-145 carcinoma cells in fibroblast conditioned medium resulted in an enhanced proliferation and anchorage-independent growth of this cell line in soft agar. Fibroblast conditioned medium also increased migration of PC-3 cells in the wound assay and slightly augmented mmp-1 expression. KGF (able to stimulate proliferation of normal and neoplastic prostate epithelial cells) was secreted by fibroblasts at higher concentrations than by all 3 tumour cell lines. In addition, fibroblasts secreted TNF·, bFGF, PDGF, HGF and also VEGF, the most important factor for tumour vascularization. Our results provide evidence that tumour-stroma interactions do not only exist at the primary site but also between metastatic prostate cancer cell lines and their fibroblastic microenvironment. These interactions, which are mediated through secreted factors, affect several steps of the metastatic cascade including proliferation, anchorage-independent growth, migration and the secretion of matrix-degrading proteases.

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Section: Discussionmentioning

confidence: 99%

Tumour-stroma interactions between metastatic prostate cancer cells and fibroblasts

Kaminski

Hahne²,

Haddouti³

et al. 2006

Int J Mol Med

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“…However, their action is not restricted to endothelial cells (Rodriguez-Nieto et al, 2001) and some effects have been shown on fibroblasts (Kria et al, 1998). Interestingly, fumagillin was also reported to inhibit Ets-1 expression induced by vascular endothelial growth factor (VEGF) (Wernert et al, 1999). In this context, we investigated the effect of fumagillin on Stromelysin-1 and Ets-1 expression in response to IL-1b and PMA in the HIG-82 cell line.…”

Section: Resultsmentioning

confidence: 94%

“…In the course of this study, we used the fungal antiangiogenic compound fumagillin because of its previously reported ability to downregulate the Ets-1 expression (Wernert et al, 1999). The precise mechanism of action of this drug and its derivatives remains unclear.…”

Section: Stromelysin-1 Promoter Regulation By Endogenous Ets-1 D Bailmentioning

confidence: 99%

Stromelysin-1 expression is activated in vivo by Ets-1 through palindromic head-to-head Ets binding sites present in the promoter

et al. 2006

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Regulation of the gene expression of Stromelysin-1 (matrix metalloproteinase-3), a member of the matrix metalloproteinase family, is critical for tissue homeostasis. The Stromelysin-1 promoter is known to be transactivated by Ets proteins through palindromic headto-head Ets binding sites (EBS), an unusual configuration among metalloproteinase promoters. Patterns of increased co-expression of Stromelysin-1 and Ets-1 genes have been observed in pathological processes such as rheumatoid arthritis, glomerulonephritis and tumor invasion. In this context, we show in a synovial fibroblastic model cell line (HIG-82), which is able to co-express Stromelysin-1 and Ets-1, that the EBS palindrome is essential for the expression of Stromelysin-1. More precisely, using electrophoretic mobility shift assays, DNA affinity purification and chromatin immunoprecipitation, we demonstrate that endogenous Ets-1, but not Ets-2, is present on this palindrome. The use of a dominant-negative form of Ets-1 and the decrease of Ets-1 amount either by fumagillin, an antiangiogenic compound, or by short interfering RNA show that the activation rate of the promoter and the expression of Stromelysin-1 correlate with the level of endogenous Ets-1. Thus, it is the first demonstration, using this cellular model, that endogenously expressed Ets-1 is actually a main activator of the Stromelysin-1 promoter through its effective binding to the EBS palindrome.

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“…Fumagillin inhibits the growth of vessels in tumors, and a derivative of the compound has been evaluated in clinical trials as an anticancer drug. The antiangiogenic effect of fumagillin and other inhibitors of MetAP-II have been attributed to the inhibition of the Ets-1 transcription factor expression and the activation of the p53 pathway (5,6). Besides from being anticancer drug targets, MetAPs have the potential to become the target proteins of antibacterial substances because the MetAP functionality is essential for cell growth and bacteria possess only one of the two known MetAP subtypes (7).…”

Section: Methionine Aminopeptidases (Metaps)mentioning

confidence: 96%

Protonation States of Methionine Aminopeptidase and Their Relevance for Inhibitor Binding and Catalytic Activity

Klein

Schiffmann

Folkers³

et al. 2003

Journal of Biological Chemistry

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We have performed a computational study of different protomeric states of the methionine aminopeptidase active site using a combined quantum-mechanical/molecular mechanical simulation approach. The aim of this study was to clarify the native protonation state of the enzyme, which is needed for the development of novel irreversible inhibitors that can possibly be used as antiangiogenic and antibiotic drugs by virtual screening and other drug design methods. The results of the simulations indicated that two protonation states are possible without disturbing the overall geometry of the active site. We then verified experimentally the presence of the two protonation states by studying the substrate hydrolysis and inhibitor binding reactions at different pH values and come to the conclusion that one of the protomeric states is relevant for inhibitor binding, whereas the other is relevant for substrate hydrolysis. This result has implications for the development of other inhibitors of this class of enzymes and adds a new perspective to the pharmacological properties of the antiangiogenic drug fumagillin, which is an irreversible inhibitor of the human methionine aminopeptidase type II.Methionine aminopeptidases (MetAPs) 1 play a central role for in vivo protein synthesis as they remove the starter methionine from newly synthetized proteins. The natural product fumagillin, an epoxide, covalently modifies one of the activesite histidines in the eukaryotic methionine aminopeptidase II (MetAP-II) (1-4) and other MetAPs (cf. Fig. 1). Fumagillin inhibits the growth of vessels in tumors, and a derivative of the compound has been evaluated in clinical trials as an anticancer drug. The antiangiogenic effect of fumagillin and other inhibitors of MetAP-II have been attributed to the inhibition of the Ets-1 transcription factor expression and the activation of the p53 pathway (5, 6). Besides from being anticancer drug targets, MetAPs have the potential to become the target proteins of antibacterial substances because the MetAP functionality is essential for cell growth and bacteria possess only one of the two known MetAP subtypes (7).MetAPs are metal-dependent enzymes. It is not clear which metal activates the MetAPs in vivo. For in vitro experiments, cobalt is commonly used because it activates all known MetAPs and the cobalt-substituted enzymes are usually the most active (8). Zinc and iron(II) have also been shown to activate some MetAPs (9, 10). The metal-chelating residues in all known MetAPs are two glutamates, two aspartates, and one histidine. The geometric arrangement of these residues is practically identical in all of the MetAP x-ray structures (11,12). Several three-dimensional structures of MetAPs have been determined by x-ray diffraction methods including the structure of human MetAP-II with a covalently bound fumagillin molecule (13-15).Density functional theory (16, 17) has long been used to study the reactivity of organic molecules and recently began to find its way into biochemistry (18). We present here an ex...

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Inhibition of Angiogenesis In Vivo byets-1 Antisense Oligonucleotides—Inhibition of Ets-1 Transcription Factor Expression by the Antibiotic Fumagillin

Cited by 69 publications

References 19 publications

Tumour-stroma interactions between metastatic prostate cancer cells and fibroblasts

Tumour-stroma interactions between metastatic prostate cancer cells and fibroblasts

Stromelysin-1 expression is activated in vivo by Ets-1 through palindromic head-to-head Ets binding sites present in the promoter

Protonation States of Methionine Aminopeptidase and Their Relevance for Inhibitor Binding and Catalytic Activity

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