ERK2-regulated TIMP1 Induces Hyperproliferation of K-RasG12D-Transformed Pancreatic Ductal Cells (original) (raw)
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2016
Pancreatic ductal adenocarcinomas (PDAC) are highly invasive and metastatic neoplasms commonly unre-sponsive to current drug therapy. Overwhelmingly, PDAC harbors early constitutive, oncogenic mutations in K-RasG12D that exist prior to invasion. Histologic and genetic analyses of human PDAC biopsies also exhibit increased expression of extracellular signal-regulated kinase (ERK) 1/2 and proinvasive matrix metalloproteinases (MMP), indicators of poor prognosis. However, the distinct molecular mechanisms necessary for K-Ras/ERK1/2 signaling and its influence on MMP-directed stromal invasion in primary human pancreatic ductal epithelial cells (PDEC) have yet to be elucidated in three-dimensions. Expression of oncogenic K-RasG12D alone in genetically defined PDECs reveals increased invadopodia and epithelial-to-mesenchymal transition markers, but only when cultured in a three-dimensional model incorporating a basement membrane analog. Activation of ERK2, but not ERK1, also occurs only i...
Activation of ERK 2 Specifically Regulates 3 D Invasion of Human Pancreatic Cancer Cells via MMP-1
2011
Constitutive K-Ras Activation of ERK2 Specifically Regulates 3D Invasion of Human Pancreatic Cancer Cells via MMP-1 Gregory P. Botta, Mauricio J. Reginato, Maximillian Reichert, Anil K. Rustgi, Peter I. Lelkes Department of Biochemistry and Molecular Biology, Drexel University College of Medicine, Philadelphia, PA 19102 Drexel University School of Biomedical Engineering, Philadelphia, PA, 19104 Division of Gastroenterology, University of Pennsylvania School of Medicine, Philadelphia, PA, 19104
2012
Pancreatic ductal adenocarcinomas (PDAC) are highly invasive and metastatic neoplasms commonly unresponsive to current drug therapy. Overwhelmingly, PDAC harbors early constitutive, oncogenic mutations in K-Ras that exist prior to invasion. Histologic and genetic analyses of human PDAC biopsies also exhibit increased expression of extracellular signal-regulated kinase (ERK) 1/2 and proinvasive matrix metalloproteinases (MMP), indicators of poor prognosis. However, the distinct molecular mechanisms necessary for K-Ras/ERK1/2 signaling and its influence on MMP-directed stromal invasion in primary human pancreatic ductal epithelial cells (PDEC) have yet to be elucidated in three-dimensions. Expression of oncogenic K-Ras alone in genetically defined PDECs reveals increased invadopodia and epithelial-to-mesenchymal transition markers, but only when cultured in a three-dimensional model incorporating a basement membrane analog. Activation of ERK2, but not ERK1, also occurs only inK-Ras–mu...
A central role for RAF��� MEK��� ERK signaling in the genesis of pancreatic ductal adenocarcinoma
2012
KRAS mutation is a hallmark of pancreatic ductal adenocarcinoma (PDA) but remains an intractable pharmacologic target. Consequently, defi ning RAS effector pathway(s) required for PDA initiation and maintenance is critical to improve treatment of this disease. Here, we show that expression of BRAF V600E , but not PIK3CA H1047R , in the mouse pancreas leads to pancreatic intraepithelial neoplasia (PanIN) lesions. Moreover, concomitant expression of BRAF V600E and TP53 R270H result in lethal PDA. We tested pharmacologic inhibitors of RAS effectors against multiple human PDA cell lines. Mitogen-activated protein (MAP)/extracellular signal-regulated (ERK) kinase (MEK) inhibition was highly effective both in vivo and in vitro and was synergistic with AKT inhibition in most cell lines tested. We show that RAF→MEK→ERK signaling is central to the initiation and maintenance of PDA and to rational combination strategies in this disease. These results emphasize the value of leveraging multiple complementary experimental systems to prioritize pathways for effective intervention strategies in PDA. SIGNIFICANCE: PDA is diffi cult to treat, in large part, due to recurrent mutations in the KRAS gene. Here, we defi ne rational treatment approaches for the disease achievable today with existing drug combinations by thorough genetic and pharmacologic dissection of the major KRAS effector pathways, RAF→MEK→ERK and phosphoinositide 3′-kinase (PI3′K)→AKT. Cancer Discov; 2(8); 685-93.
Gut, 2002
Background: Pancreatic stellate cells (PSCs) have been implicated in pancreatic fibrosis as they synthesise increased amounts of extracellular matrix proteins in response to activation by profibrogenic mediators such as cytokines. Aims: The purpose of this study was to analyse cytokine receptor stimulated signalling pathways involved in PSC activation. Using a rat culture model of PSCs, we have also tested the potential of the platelet derived growth factor (PDGF) antagonist trapidil and PD98059, a specific inhibitor of extracellular signal regulated kinase (ERK) activation, to suppress PSC growth. Methods: Cultured PSCs were stimulated with PDGF, and the signal transduction pathways activated in response to the mitogen were analysed by immunoblotting, kinase assays, and electrophoretic mobility shift assays. Furthermore, comparison of signalling cascades activated in PSCs before and after completing transdifferentiation to α-smooth muscle actin expressing myofibroblasts was performed. Biological effects of PDGF, trapidil, and PD98059 were analysed by proliferation assays and correlated with molecular effects of the substances. Results: PDGF induced rapid activation of Raf-1, ERKs 1 and 2, as well as AP-1 proteins. The transforming growth factor β activated transcription factor Smad2 was found to be constitutively phosphorylated in PSCs of different transdifferentiation grades. Furthermore, the results indicate a correlation between ERK activities and induction of PSC activation. Trapidil efficiently inhibited both PDGF induced ERK activation and, in common with PD98059, PSC proliferation. Conclusions: Our data suggest that ERKs play a key role in the regulation of PSC growth and that inhibition of the ERK signalling pathway may become a strategy to prevent activation of these cells.
Deciphering the Mechanisms of Tumorigenesis in Human Pancreatic Ductal Epithelial Cells
Clinical Cancer Research, 2013
Purpose: The most common genetic lesions in pancreatic ductal adenocarcinoma (PDAC) have been identified. However, significant gaps still exist in our understanding of how such genetic alterations act in concert to induce PDAC development. In this study, we investigated the mechanism of tumorigenic transformation in the immortalized human pancreatic ductal epithelial (HPDE) cell line by sequentially introducing PDAC signature alterations into this cell line. Experimental Design: The phenotype for stable expression of mutant K-ras, Her2, p16/p14shRNA, and Smad4shRNA in HPDE cells was examined by assays for cell proliferation, migration, invasion, soft agar, and orthotopic tumorigenesis. The mechanisms of tumorigenic transformation were further explored by gene expression profiling and pathway analyses. Results: The transformed cells exhibited enhanced proliferation, migration, and invasion, displayed anchorage-independent growth in soft agar, and grew orthotopic tumors with some hist...
KrasG12D induces EGFR-MYC cross signaling in murine primary pancreatic ductal epithelial cells
Oncogene, 2015
Epidermal growth factor receptor (EGFR) signaling has a critical role in oncogenic Kras-driven pancreatic carcinogenesis. However, the downstream targets of this signaling network are largely unknown. We developed a novel model system utilizing murine primary pancreatic ductal epithelial cells (PDECs), genetically engineered to allow time-specific expression of oncogenic Kras G12D from the endogenous promoter. We show that primary PDECs are susceptible to Kras G12D-driven transformation and form pancreatic ductal adenocarcinomas (PDAC) in vivo after Cdkn2a inactivation. In addition, we demonstrate that activation of Kras G12D induces an EGFR signaling loop to drive proliferation. Interestingly, pharmacological inhibition of EGFR fails to Users may view, print, copy, and download text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use: