G F Vande Woude

Publications (135)1331.44 Total impact

• Article: RhoA and RhoC are both required for the ROCK II-dependent promotion of centrosome duplication.

  M Kanai, M S Crowe, Y Zheng, G F Vande Woude, K Fukasawa
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  ABSTRACT: CDK2-cyclin E triggers centrosome duplication, and nucleophosmin (NPM/B23) is found to be one of its targets. NPM/B23 phosphorylated by CDK2-cyclin E acquires a high binding affinity to Rho-associated kinase (ROCK II), and physically associates with ROCK II. The NPM/B23-binding results in superactivation of ROCK II, which is a critical event for initiation of centrosome duplication. The activation of ROCK II also requires the binding of Rho small GTPase to the auto-inhibitory region; hence the availability of the active Rho protein is an important aspect of the centrosomally localized ROCK II to properly initiate centrosome duplication. There are three isoforms of Rho (RhoA, B and C), all of which are capable of binding to and priming the activation of ROCK II. Here, we investigated which Rho isoform(s) are involved in the activation of ROCK II in respect to the initiation of centrosome duplication. We found that both RhoA and RhoC, but not RhoB, were required for initiation of centrosome duplication, and overactivation of RhoA, as well as RhoC, but not RhoB, promoted centrosome duplication and centrosome amplification.
  Oncogene 11/2010; 29(45):6040-50. · 6.37 Impact Factor
• Article: Differential expression of c-Met, its ligand HGF/SF and HER2/neu in DCIS and adjacent normal breast tissue.

  K Lindemann, J Resau, J Nährig, E Kort, B Leeser, K Annecke, A Welk, J Schäfer, G F Vande Woude, E Lengyel, N Harbeck
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  ABSTRACT: Tyrosine kinase receptors Her2/neu and c-Met play an important role in breast cancer development and progression. Our aim was to determine the expression of c-Met, its ligand hepatocyte growth factor/scatter factor (HGF/SF) and Her2/neu in ductal carcinoma in situ (DCIS) lesions of the breast (n = 39) by two different immunocytochemical techniques, classical immunohistochemistry and immunofluorescence, and to correlate their expression levels with histopathological and clinical characteristics. Both methods revealed similar c-Met staining patterns in both the in situ component and the adjacent normal tissue (P < 0.001). However, an imbalance in c-Met expression between tumour and surrounding normal tissue was correlated with high-grade DCIS (Van Nuys Grade 3). No correlation existed between Her2/neu and c-Met expression. High HGF/SF immunoreactivity was observed in 43.6% of the cases, yet the adjacent cellular stroma revealed only low levels of HGF/SF. No correlation existed between c-Met, Her2/neu or HGF/SF expression and clinicopathological factors. An imbalance in c-Met expression between tumour and surrounding normal tissue is associated with an aggressive DCIS phenotype. Moreover, c-Met and HGF/SF may contribute to tumour development by different means than those controlled by Her2/neu.
  Histopathology 07/2007; 51(1):54-62. · 3.08 Impact Factor
• Article: Evidence that MIG-6 is a tumor-suppressor gene.

  Y-W Zhang, B Staal, Y Su, P Swiatek, P Zhao, B Cao, J Resau, R Sigler, R Bronson, G F Vande Woude
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  ABSTRACT: Mitogen-inducible gene 6 (MIG-6) is located in human chromosome 1p36, a locus frequently associated with human lung cancer. MIG-6 is a negative regulator of epidermal growth factor (EGF) signaling, and we show that Mig-6 - like EGF - is induced by hepatocyte growth factor/scatter factor (HGF/SF) in human lung cancer cell lines. Frequently, the receptors for both factors, EGFR and Met, are expressed in same lung cancer cell line, and MIG-6 is induced by both factors in a mitogen-activated protein kinase-dependent fashion. However, not all tumor lines express MIG-6 in response to either EGF or HGF/SF. In these cases, we find missense and nonsense mutations in the MIG-6 coding region, as well as evidence for MIG-6 transcriptional silencing. Moreover, germline disruption of Mig-6 in mice leads to the development of animals with epithelial hyperplasia, adenoma, and adenocarcinoma in organs like the lung, gallbladder, and bile duct. These data suggests that MIG-6 is a tumor-suppressor gene and is therefore a candidate gene for the frequent 1p36 genetic alterations found in lung cancer.
  Oncogene 02/2007; 26(2):269-76. · 6.37 Impact Factor
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  Available from: pnas.org

  Article: Neutralizing monoclonal antibodies to hepatocyte growth factor/scatter factor (HGF/SF) display antitumor activity in animal models.

  B Cao, Y Su, M Oskarsson, P Zhao, E J Kort, R J Fisher, L M Wang, G F Vande Woude
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  ABSTRACT: The hepatocyte growth factor (HGF/SF) receptor, Met, regulates mitogenesis, motility, and morphogenesis in a cell type-dependent fashion. Activation of Met via autocrine, paracrine, or mutational mechanisms can lead to tumorigenesis and metastasis and numerous studies have linked inappropriate expression of this ligand-receptor pair to most types of human solid tumors. To prepare mAbs to human HGF/SF, mice were immunized with native and denatured preparations of the ligand. Recloned mAbs were tested in vitro for blocking activity against scattering and branching morphogenesis. Our results show that no single mAb was capable of neutralizing the in vitro activity of HGF/SF, and that the ligand possesses a minimum of three epitopes that must be blocked to prevent Met tyrosine kinase activation. In vivo, the neutralizing mAb combination inhibited s.c. growth in athymic nu/nu mice of tumors dependent on an autocrine Met-HGF/SF loop. Importantly, growth of human glioblastoma multiforme xenografts expressing Met and HGF/SF were markedly reduced in the presence of HGF/SF-neutralizing mAbs. These results suggest interrupting autocrine and/or paracrine Met-HGF/SF signaling in tumors dependent on this pathway is a possible intervention strategy.
  Proceedings of the National Academy of Sciences 07/2001; 98(13):7443-8. · 9.68 Impact Factor
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  Available from: pnas.org

  Article: Suppression of ras-mediated transformation and inhibition of tumor growth and angiogenesis by anthrax lethal factor, a proteolytic inhibitor of multiple MEK pathways.

  N S Duesbery, J Resau, C P Webb, S Koochekpour, H M Koo, S H Leppla, G F Vande Woude
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  ABSTRACT: Lethal factor is a protease, one component of Bacillus anthracis exotoxin, which cleaves many of the mitogen-activated protein kinase kinases (MEKs). Given the importance of MEK signaling in tumorigenesis, we assessed the effects of anthrax lethal toxin (LeTx) on tumor cells. LeTx was very effective in inhibiting mitogen-activated protein kinase activation in V12 H-ras-transformed NIH 3T3 cells. In vitro, treatment of transformed cells with LeTx caused them to revert to a nontransformed morphology, and inhibited their abilities to form colonies in soft agar and to invade Matrigel without markedly affecting cell proliferation. In vivo, LeTx inhibited growth of ras-transformed cells implanted in athymic nude mice (in some cases causing tumor regression) at concentrations that caused no apparent animal toxicity. Unexpectedly, LeTx also greatly decreased tumor neovascularization. These results demonstrate that LeTx potently inhibits ras-mediated tumor growth and is a potential antitumor therapeutic.
  Proceedings of the National Academy of Sciences 04/2001; 98(7):4089-94. · 9.68 Impact Factor
• Article: Animal models for Ras-induced metastasis.

  C P Webb, G F Vande Woude
  Methods in Enzymology 02/2001; 333:318-29. · 2.04 Impact Factor
• Source

  Available from: Joseph Testa

  Article: Anti-apoptotic signaling by hepatocyte growth factor/Met via the phosphatidylinositol 3-kinase/Akt and mitogen-activated protein kinase pathways.

  G H Xiao, M Jeffers, A Bellacosa, Y Mitsuuchi, G F Vande Woude, J R Testa
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  ABSTRACT: Hepatocyte growth factor (HGF) is a ligand of the receptor tyrosine kinase encoded by the c-Met protooncogene. HGF/Met signaling has multifunctional effects on various cell types. We sought to determine the role of HGF/Met in apoptosis and identify signal transducers involved in this process. In experiments with human SK-LMS-1 leiomyosarcoma cells, we show that the Akt kinase is activated by HGF in a time- and dose-dependent manner by phosphatidylinositol 3-kinase (PI3-kinase). Akt is also activated by active tumorigenic forms of Met, i.e., ligand-independent Tpr-Met, a truncated and constitutively dimerized form of Met, and a mutationally activated version of Met corresponding to that found in human hereditary papillary renal carcinoma. In NIH 3T3 cells transfected with wild-type Met, HGF inhibits apoptosis induced by serum starvation and UV irradiation. HGF-induced survival correlates with Akt activity and is inhibited by the specific PI3-kinase inhibitor LY294002, indicating that HGF inhibits cell death through the PI3-kinase/Akt signal transduction pathway. Furthermore, transiently transfected Tpr-Met activates Akt (both Akt1 and Akt2) and protects cells from apoptosis. Mitogen-activated protein kinase (MAPK) also is activated by HGF and rescues cells from apoptosis, although the cytoprotective effect is less marked than for PI3-kinase/Akt. Blocking MAPK with the specific MAPK kinase inhibitor PD098059 impairs the ability of HGF to promote cell survival. Similar results were obtained with NIH 3T3 cells expressing the fusion protein Trk-Met and stimulated with nerve growth factor, the Trk ligand. These results demonstrate that HGF/Met is capable of protecting cells from apoptosis by using both PI3-kinase/Akt and, to a lesser extent, MAPK pathways.
  Proceedings of the National Academy of Sciences 02/2001; 98(1):247-52. · 9.68 Impact Factor
• Article: A novel germ line juxtamembrane Met mutation in human gastric cancer.

  J H Lee, S U Han, H Cho, B Jennings, B Gerrard, M Dean, L Schmidt, B Zbar, G F Vande Woude
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  ABSTRACT: Activating mutations in the Met receptor tyrosine kinase, both germline and somatic, have been identified in human papillary renal cancer. Here we report a novel germline missense Met mutation, P1009S, in a patient with primary gastric cancer. The dosage of the mutant Met DNA was elevated in the tumor when compared to its matched normal DNA. Therefore, as with hereditary renal papillary cancer, the mutant Met allele may also be selectively duplicated in the tumor. Different from previously reported Met mutations, which occur in the tyrosine kinase domain, this missense mutation is located at the juxtamembrane domain, and is not constitutively activated. However, following treatment with HGF/SF, the P1009S mutant Met protein, expressed in NIH3T3 cells, displays increased and persistent tyrosine phosphorylation compared to the wild-type Met. Importantly, these cells also form colonies in soft agar, and are highly tumorigenic in athymic nude mice. A second nucleotide change in this region of Met, T1010I, was found in a breast cancer biopsy and a large cell lung cancer cell line. Although this previously reported 'polymorphism' did not stimulate NIH3T3 cell growth in soft agar, it was more active than the wild-type Met in the athymic nude mice tumorigenesis assay, suggesting that it may have effects on tumorigenesis. Met has been shown to be highly expressed in human gastric carcinoma cell lines, and our results raise the possibility that activating missense Met mutations could contribute to tumorigenesis of gastric cancer.
  Oncogene 11/2000; 19(43):4947-53. · 6.37 Impact Factor
• Article: Genes that regulate metastasis and angiogenesis.

  C P Webb, G F Vande Woude
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  ABSTRACT: Genetic instability and an accumulation of genetic and epigenetic changes during tumor progression lead to an increasingly aggressive and treatment-resistant phenotype, and ultimately metastasis. In recent years it has become well established that angiogenesis, the process by which new vasculature is formed from pre-existing vessels, is an essential component to primary tumor growth and distant metastasis. A greater understanding of the complex multitude of factors involved in tumor angiogenesis and metastasis is fundamental to the development of potential therapeutics to treat malignant disease. As highlighted throughout this review, angiogenesis and metastasis share many common cellular and molecular features. We will briefly discuss the pertinent genes involved in the regulation of angiogenesis and metastasis.
  Journal of Neuro-Oncology 09/2000; 50(1-2):71-87. · 3.21 Impact Factor
• Article: Dominant negative Met reduces tumorigenicity-metastasis and increases tubule formation in mammary cells.

  M Firon, M Shaharabany, R T Altstock, J Horev, A Abramovici, J H Resau, G F Vande Woude, I Tsarfaty
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  ABSTRACT: Activation of the Met tyrosine kinase growth factor receptor by its ligand HGF/SF has been shown to increase in vitro invasiveness in epithelial cell lines. To study the effect of Met-HGF/SF signaling in breast cancer cells, we transfected met, hgf/sf and dominant negative (DN) forms of met into the poorly differentiated metastatic murine mammary adenocarcinoma cell line DA3. These cells express moderate levels of endogenous Met, which is rapidly phosphorylated in response to HGF/SF treatment. Met+hgf/sf transfection results in significantly increased tumorigenic and metastatic activity in vivo accompanied by reduced tubule formation. DA3 cells transfected with DN forms of Met (DN-DA3) exhibit reduced Met phosphorylation following exposure to HGF/SF. Furthermore, as compared to the parental cells, the DN-DA3 cells exhibit diminished in vitro scattering and invasiveness, while in vivo they display greatly reduced tumorigenicity and spontaneous metastasis. Tumors emanating from DN-DA3 cells injected to BALB/C mice are highly differentiated and display extensive tubule formation. These results suggest that Met-HGF/SF signaling is a determining factor in the delicate balance between differentiation/tubule formation and tumorigenicity-metastasis. Oncogene (2000) 19, 2386 - 2397
  Oncogene 06/2000; 19(20):2386-97. · 6.37 Impact Factor
• Article: Regulation of P311 expression by Met-hepatocyte growth factor/scatter factor and the ubiquitin/proteasome system.

  G A Taylor, E Hudson, J H Resau, G F Vande Woude
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  ABSTRACT: P311 is a mouse cDNA originally identified for its high expression in late-stage embryonic brain and adult cerebellum, hippocampus, and olfactory bulb. The protein product of P311, however, had not been identified previously, and its function remains unknown. We report here that P311 expression is regulated at multiple levels by pathways that control cellular transformation. P311 mRNA expression was decreased sharply in both neural and smooth muscle cells when the cells were transformed by coexpression of the oncogenic tyrosine kinase receptor Met and its ligand hepatocyte growth factor/scatter factor. The P311 mRNA was found to encode an 8-kDa polypeptide that was subject to rapid degradation by the lactacystin-sensitive ubiquitin/proteasome system and an unidentified metalloprotease, resulting in a protein half-life of about 5 min. These data suggest that P311 expression is dramatically decreased by several pathways that regulate cellular growth.
  Journal of Biological Chemistry 03/2000; 275(6):4215-9. · 4.77 Impact Factor
• Article: The geldanamycins are potent inhibitors of the hepatocyte growth factor/scatter factor-met-urokinase plasminogen activator-plasmin proteolytic network.

  C P Webb, C D Hose, S Koochekpour, M Jeffers, M Oskarsson, E Sausville, A Monks, G F Vande Woude
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  ABSTRACT: The Met receptor tyrosine kinase and its ligand, hepatocyte growth factor/scatter factor (HGF/SF), have been implicated in human tumor development and metastasis. HGF/SF induces the expression of urokinase plasminogen activator (uPA) and the uPA receptor (uPAR), important mediators of cell invasion and metastasis. We have developed a cell-based assay to screen for inhibitors of this signaling system using the induction of endogenous uPA and uPAR and the subsequent conversion of plasminogen to plasmin as the biological end point. Assay validation was established using a neutralizing antiserum to HGF/SF and a uPA inhibitor (B428), as well as inhibitors of the MKK-MAPK1/2 pathway, shown previously to be important in the induction of uPA and uPAR. Using this assay, we found several classes of molecules that exhibited inhibition of HGF/SF-dependent plasmin activation. However, we discovered that certain members of the geldanamycin family of anisamycin antibiotics are potent inhibitors of HGF/SF-mediated plasmin activation, displaying inhibitory properties at femtomolar concentrations and nine orders of magnitude below their growth inhibitory concentrations. At nanomolar concentrations, the geldanamycins down-regulate Met protein expression, inhibit HGF/SF-mediated cell motility and invasion, and also revert the phenotype of both autocrine HGF/SF-Met transformed cells as well as those transformed by Met proteins with activating mutations. Thus, the geldanamycins may have important therapeutic potential for the treatment of cancers in which Met activity contributes to the invasive/metastatic phenotype.
  Cancer Research 02/2000; 60(2):342-9. · 7.86 Impact Factor
• Article: Pathogen-specific loss of host resistance in mice lacking the IFN-gamma-inducible gene IGTP.

  G A Taylor, C M Collazo, G S Yap, K Nguyen, T A Gregorio, L S Taylor, B Eagleson, L Secrest, E A Southon, S W Reid, L Tessarollo, M Bray, D W McVicar, K L Komschlies, H A Young, C A Biron, A Sher, G F Vande Woude
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  ABSTRACT: Interferon-gamma (IFN-gamma) is critical for defense against pathogens, but the molecules that mediate its antimicrobial responses are largely unknown. IGTP is the prototype for a family of IFN-gamma-regulated genes that encode 48-kDa GTP-binding proteins that localize to the endoplasmic reticulum. We have generated IGTP-deficient mice and found that, despite normal immune cell development and normal clearance of Listeria monocytogenes and cytomegalovirus infections, the mice displayed a profound loss of host resistance to acute infections of the protozoan parasite Toxoplasma gondii. By contrast, IFN-gamma receptor-deficient mice have increased susceptibility to all three pathogens. Thus, IGTP defines an IFN-gamma-regulated pathway with a specialized role in antimicrobial resistance.
  Proceedings of the National Academy of Sciences 02/2000; 97(2):751-5. · 9.68 Impact Factor
• Article: Ras oncogene-induced sensitization to 1-beta-D-arabinofuranosylcytosine.

  H M Koo, M J McWilliams, W G Alvord, G F Vande Woude
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  ABSTRACT: Human tumor cells containing ras oncogenes display enhanced sensitivity to 1-beta-D-arabinofuranosylcytosine (Ara-C) and other deoxycytidine analogues (H-M. Koo, et al., Cancer Res., 56: 5211-5216, 1996). Human tumor cell lines with or without a ras oncogene as well as a pair of isogenic cell lines with one containing an activated ras oncogene were used to study the basis for differential sensitivity. We found that human tumor cells containing ras oncogenes upon entry into the S phase of the cell cycle underwent apoptosis in response to Ara-C treatment. By contrast, human tumor cells harboring wild-type ras alleles were only delayed in the S phase when exposed to Ara-C. Thus, the ras oncogene specifically renders human cells more sensitive to Ara-C by preventing S-phase arrest. This may occur by the ras oncogene compromising an S-phase checkpoint.
  Cancer Research 01/2000; 59(24):6057-62. · 7.86 Impact Factor
• Article: Alteration of Met protooncogene product expression and prognosis in breast carcinomas.

  I Tsarfaty, W G Alvord, J H Resau, R T Altstock, R Lidereau, I Bieche, F Bertrand, J Horev, R L Klabansky, I Keydar, G F Vande Woude
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  ABSTRACT: To objectively quantify the expression and prognostic implications of the met protooncogene product (Met) in human breast cancer. One hundred eighty-two cases of primary human breast cancer were collected. Both the normal and tumor portions of the original surgical pathology specimen were immunostained for Met and imaged using laser scanning confocal microscopy. Then the cases were ranked according to relative concentrations of normal and tumor Met expression. Subsequently, they were quantified using image analysis and the results correlated with clinical outcome to determine the prognostic value of relative levels of Met. Using a quantitative index to evaluate the relative levels of Met expression, high levels of Met expression in the tumor as compared with the adjacent normal ducts predicted poor prognosis for overall survival and metastasis-free survival. The risk ratio for elevated Met expression was 3.94 (P = .0009). This new method also allows determination of the clinical relevance of low levels of Met in the tumor. The overall survival between the patient population with higher, lower and unchanged levels of Met in normal tissue as compared to tumor were significantly different (P = .0020). Our studies suggest that in a subpopulation of node-negative breast cancer patients, either high or low levels of Met in tumor tissue relative to normal tissue is an indicator of poor overall survival (P = .0068). Thus, Met expression could be useful for identifying node-negative patients who could benefit from adjuvant therapy.
  Analytical and quantitative cytology and histology / the International Academy of Cytology [and] American Society of Cytology 11/1999; 21(5):397-408. · 0.41 Impact Factor
• Article: Anthrax toxins.

  N S Duesbery, G F Vande Woude
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  ABSTRACT: Though its lethal effects were ascribed to an exotoxin almost half a century ago, the pathogenesis of anthrax has yet to be satisfactorily explained. Subsequent work has led to the molecular identification and enzymatic characterization of three proteins that constitute two anthrax toxins. Protective antigen binds an as yet unknown cell receptor and mediates the entry of the other two components to the cytoplasm via the endosomal pathway. Edema factor, so named for its ability to induce edema, is a Ca2+/calmodulin-dependent adenylate cyclase. Lethal factor, the dominant virulence factor associated with the toxin, proteolytically inactivates mitogen-activated protein kinase kinases, key players in signal transduction. We describe the fascinating work that has led to these discoveries and discuss their relevance to our understanding of the pathogenesis of anthrax.
  Cellular and Molecular Life Sciences CMLS 10/1999; 55(12):1599-609. · 6.57 Impact Factor
• Article: The von Hippel-Lindau tumor suppressor gene inhibits hepatocyte growth factor/scatter factor-induced invasion and branching morphogenesis in renal carcinoma cells.

  S Koochekpour, M Jeffers, P H Wang, C Gong, G A Taylor, L M Roessler, R Stearman, J R Vasselli, W G Stetler-Stevenson, W G Kaelin, W M Linehan, R D Klausner, J R Gnarra, G F Vande Woude
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  ABSTRACT: Loss of function in the von Hippel-Lindau (VHL) tumor suppressor gene occurs in familial and most sporadic renal cell carcinomas (RCCs). VHL has been linked to the regulation of cell cycle cessation (G(0)) and to control of expression of various mRNAs such as for vascular endothelial growth factor. RCC cells express the Met receptor tyrosine kinase, and Met mediates invasion and branching morphogenesis in many cell types in response to hepatocyte growth factor/scatter factor (HGF/SF). We examined the HGF/SF responsiveness of RCC cells containing endogenous mutated (mut) forms of the VHL protein (VHL-negative RCC) with that of isogenic cells expressing exogenous wild-type (wt) VHL (VHL-positive RCC). We found that VHL-negative 786-0 and UOK-101 RCC cells were highly invasive through growth factor-reduced (GFR) Matrigel-coated filters and exhibited an extensive branching morphogenesis phenotype in response to HGF/SF in the three-dimensional (3D) GFR Matrigel cultures. In contrast, the phenotypes of A498 VHL-negative RCC cells were weaker, and isogenic RCC cells ectopically expressing wt VHL did not respond at all. We found that all VHL-negative RCC cells expressed reduced levels of tissue inhibitor of metalloproteinase 2 (TIMP-2) relative to the wt VHL-positive cells, implicating VHL in the regulation of this molecule. However, consistent with the more invasive phenotype of the 786-0 and UOK-101 VHL-negative RCC cells, the levels of TIMP-1 and TIMP-2 were reduced and levels of the matrix metalloproteinases 2 and 9 were elevated compared to the noninvasive VHL-positive RCC cells. Moreover, recombinant TIMPs completely blocked HGF/SF-mediated branching morphogenesis, while neutralizing antibodies to the TIMPs stimulated HGF/SF-mediated invasion in vitro. Thus, the loss of the VHL tumor suppressor gene is central to changes that control tissue invasiveness, and a more invasive phenotype requires additional genetic changes seen in some but not all RCC lines. These studies also demonstrate a synergy between the loss of VHL function and Met signaling.
  Molecular and Cellular Biology 10/1999; 19(9):5902-12. · 5.53 Impact Factor
• Article: Anthrax lethal factor causes proteolytic inactivation of mitogen-activated protein kinase kinase.

  N S Duesbery, G F Vande Woude
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  ABSTRACT: A search of the National Cancer Institute's Anti-Neoplastic Drug Screen for compounds with an inhibitory profile similar to that of the mitogen-activated protein kinase kinase (MAPKK) inhibitor PD098059 yielded anthrax lethal toxin. Anthrax lethal factor was found to inhibit progesterone-induced meiotic maturation of frog oocytes by preventing the phosphorylation and activation of mitogen-activated protein kinase (MAPK). Similarly, lethal toxin prevented the activation of MAPK in serum stimulated, ras-transformed NIH3T3 cells. In vitro analyses using recombinant proteins indicated that lethal factor proteolytically modified the NH2-terminus of both MAPKK1 and 2, rendering them inactive and hence incapable of activating MAPK. The consequences of this inactivation upon meiosis and transformed cells are also discussed.
  Journal of Applied Microbiology 09/1999; 87(2):289-93. · 2.34 Impact Factor
• Article: MEK wars, a new front in the battle against cancer.

  N S Duesbery, C P Webb, G F Vande Woude
  Nature Medicine 08/1999; 5(7):736-7. · 22.46 Impact Factor
• Article: Activation mechanisms of the urokinase-type plasminogen activator promoter by hepatocyte growth factor/scatter factor.

  S Ried, C Jäger, M Jeffers, G F Vande Woude, H Graeff, M Schmitt, E Lengyel
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  ABSTRACT: Hepatocyte growth factor/scatter factor (HGF/SF) is a pleiotropic effector inducing invasion and metastasis of tumor cells that express the Met tyrosine kinase receptor. One of the effectors of HGF/SF is the urokinase-type plasminogen activator, a serine protease that facilitates tumor progression and metastasis by controlling the synthesis of the extracellular matrix degrading plasmin. Stimulation of NIH 3T3 cells that were stably transfected with the human Met receptor (NIH 3T3-Methum) with HGF/SF induced a trans-activation of the urokinase promoter and urokinase secretion. Induction of the urokinase promoter by HGF/SF via the Met receptor was blocked by co-expression of a dominant-negative Grb2 and Sos1 expression construct. Further, the expression of the catalytically inactive mutants of Ha-Ras, RhoA, c-Raf, and Erk2 or addition of the Mek1-specific inhibitor PD 098059 abrogated the stimulation of the urokinase promoter by HGF/SF. A sequence residing between -2109 and -1870 base pairs (bp) was critical for stimulation of the urokinase gene by HGF/SF. Mobility shift assays with oligonucleotides spanning an AP-1 site at -1880 bp or a combined PEA3/AP-1 site at -1967 bp showed binding of nuclear factors from NIH 3T3-Methum cells. Expression of an expression plasmid that inhibits DNA binding of AP-1 proteins (A-Fos) abrogated inducible and basal activation of the urokinase promoter. Nuclear extract from unstimulated NIH 3T3-Methum cells contained more JunD and showed a stronger JunD supershift with the AP-1 oligonucleotides, compared with HGF/SF-stimulated cells. Consistent with the levels of JunD expression being functionally important for basal expression of the urokinase promoter, we found that overexpression of wild type JunD inhibited the induction of the urokinase promoter by HGF/SF. These data suggest that the induction of urokinase by HGF/SF is regulated by a Grb2/Sos1/Ha-Ras/c-Raf/RhoA/Mek1/Erk2/c-++ +Jun-dependent mitogen-activated protein kinase pathway.
  Journal of Biological Chemistry 07/1999; 274(23):16377-86. · 4.77 Impact Factor