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Neural invasion in pancreatic cancer is characterized by beta-1-Integrin- and L1-CAM-dependent heterotypic cell adhesion between pancreatic cancer cells and neural Schwann cells
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Abstracts / Pancreatology 13 (2013) S2–S98
Conclusion: Our study uncovers an cell-autonomous EGFR-NFATc1 signaling loop which is required for pancreatic cancer initiation.
O-42 Abstract id: 318. Nuclear factor of activated T-cells c1 integrates STAT3 signals to link chronic inflammation and carcinogenesis in the pancreas Sandra Baumgart 1, Nai-ming Chen 1, Marius Brunner 1, Reinecke €nig 1, Jinsan Johanna 1, Julius Nikorowitsch 1, Shiv Singh 1, Alexander Ko Zhang 2, Elmar Wolf 3, Marek Bartkuhn 4, Jens Siveke 5, Irene Esposito 5, Elisabeth Glesel 1, Bettina Geisel 1, Kristina Reutlinger 1, Garima Singh 1, Martin Eilers 3, Thomas Gress 1, Daniel Billadeau 2, Volker Ellenrieder 1. 1
Philipps University of Marburg, Germany Mayo Clinic, United States 3 €rzburg, Germany University of Wu 4 Justus-Liebig University of Giessen, Germany 5 Technical University of Munich, Germany 2
Introduction: Chronic pancreatitis along with activating mutations of the oncogene Kras is a central risk factor for pancreatic cancer development. Inflammation-induced signaling involves nuclear factor of activated T cells (NFAT) pathways. NFATc1 is overexpressed and activated in Krasmutated human pancreatic cancers where it mediates cancer growth stimulation. Aims: This study aims to determine the in vivo role of NFATc1 in KrasG12D-dependent carcinogenesis. Materials & methods: Transgenic mice expressing KrasG12D and NFATc1 in the pancreas and NFATc1 knockout mice were engineered and analyzed in terms of (inflammation-induced) carcinogenesis. Immunohistochemistry, Western blot, qPCR analyses, chromatin studies, ChIP-Seq and genome-wide expression profiling were performed to investigate the mechanisms of NFATc1-KrasG12D cooperation in murine and human tissues and tumor cells. Results: Activation of NFATc1 - upon inflammation and in KrasG12D;NFATc1 mice - dramatically accelerated carcinogenesis and reduced survival. Mechanistically, nuclear NFATc1 activated expression of oncogenic STAT3 transcription factor. Accordingly, high correlative nuclear expression levels of NFATc1 and active STAT3 were detected in the majority of human and murine cancer tissues. In turn, NFATc1/STAT3 complexes, which regulated genome-wide NFATc1 binding to distal chromatin enhancer sites and subsequently enforced recruitment of transcriptional co-activators, were identified. Likewise, corresponding enhancer-promoter communications that stimulated NFATc1-dependent transcription of newly-identified target genes were epigenetically promoted. Pharmacologic and genetic depletion of the NFATc1/STAT3-axis significantly arrested carcinogenesis in mouse models and confirmed the requirement of NFATc1 in Kras-driven pancreatic carcinogenesis. Conclusion: We identified a previously unknown NFATc1-STAT3 complex formation as a driving epigenetic regulatory force in inflammation-linked pancreatic carcinogenesis, which defines this novel pathway a possible point of therapeutic intervention.
O-43 Abstract id: 236. Therapeutic interest to target the Phosphoinositide-3-Kinase (PI3K) – mTOR pathway in pancreatic Cancer-Associated Fibroblasts Camille Duluc 1, Siham Moatassim-Billah 2, Herbert Schmid 3, Marieephane Pyronnet 1, Julie Bernadette Delisle 4, Barbara Garmy-Susini 1, St Guillermet-Guibert 1, Corinne Bousquet 1. 1
INSERM U1037, France Faculty of Sciences Rabat, University Mohammed V - Agdal, Morocco 3 Novartis, Bales, Switzerland 4 Pathology Toulouse Hospital, France 2
Introduction: Pancreatic ductal adenocarcinoma (PDAC) is one of the most stroma-rich cancers. Cancer-associated fibroblasts (CAFs), the most abundant cells in this stroma, orchestrate the secretion of growth factors that engage, in cancer and other stromal cells, survival and angiogenic signals redundant to those targeted by therapies, and of fibrillar components that constitute a barrier to drug delivery. Aims: Therapeutic pitfalls observed in PDAC may in part be explained by the under-estimation of the influences exerted by the microenvironment on cancer cells. Targeting CAFs may therefore constitute a promising strategy. Patients & methods: Primary cultures of CAFs have been isolated from human pancreatic tumor resections and used for in vitro and in vivo studies. Results: Isolated CAFs present, and maintain during the in vitro passages (< 10), an activated phenotype ( aSMA-positive). CAFs promote survival, invasion and chemoprotection (against gemcitabine) of pancreatic cancer cells, either in co-cultures or using CAF conditioned media. Interestingly, CAFs present a high intrinsic activation of the PI3K-mTOR pathway. It results in elevated protein synthesis and secretion specifically of pro-tumoral factors and extracellular matrix proteins. Strikingly, drugs that inhibit the PI3K-mTOR pathway abrogate CAF pro-tumoral and chemoprotective effects on cancer cells both in vitro and in vivo, in co-xenografted mice with CAF and pancreatic cancer cells. Conclusion: Because CAFs are present both in the tumor and in the periphery where drug delivery is still feasible, therapeutic targeting of CAFs using inhibitors of protein synthesis may be of utmost interest for PDAC.
O-44 Abstract id: 187. Neural invasion in pancreatic cancer is characterized by beta-1-Integrin- and L1-CAM-dependent heterotypic cell adhesion between pancreatic cancer cells and neural Schwann cells € ralp O. Ihsan Ekin Demir, Natascha Klose, Eva Brunner, Helmut Friess, Gu Ceyhan. € nchen, Department of Surgery, Klinikum rechts der Isar, TU Mu Germany Introduction: Neural invasion is one of the most frequent modes of tumor spread in pancreatic cancer (PCa). However, how PCa cells can attach to and migrate along Schwann-cell-covered axons in nerves remains unclear. Aims: The aim of this study was to investigate the potential heterotypic cell adhesion between PCa cells and Schwann cells (SC) and to identify the surface molecules that enable this interaction. Materials & methods: Human SC, PCa cells, human PCa tissue, normal human pancreas and intrapancreatic nerves were investigated for the expression of the cell adhesion molecules beta-1-Integrin, NCAM, L1-CAM and NrCAM via immunoblotting, QRT-PCR and immunohistochemistry. Heterotypic cell adhesion and mutual migration between SC and PCa cells were quantified via a recently established adhesion assay and 3D migration assay by applying neutralizing antibodies against these adhesion molecules. Results: Expression of beta1-Integrin and L1-CAM were more prevalent in PCa tissues, human SC, intrapancreatic nerves and PCa cells than NCAM and NrCAM. Confrontation of SC with PCa cells led to rapid heterotypic cell adhesion, which could be inhibited by neutralizing antibodies against beta-1-Integrin and L1-CAM, but not against NCAM and NrCAM on PCa cells. Similarly, blockade of beta-1-Integrin on PCa cells diminished the carcinotropic migration of SC. Interestingly, inhibition of these molecues on SC did not influence the adhesion or carcinotropic migration of SC. Conclusion: Neural invasion harbors heterotypic cell adhesion and mutual migration between SC and PCa cells. Therefore, cell-cell-adhesion represents a key pathophysiological mechanism in neural invasion and for the associated local tumor recurrence and neuropathic pain in PCa.
Abstracts / Pancreatology 13 (2013) S2–S98
Conclusion: Our study uncovers an cell-autonomous EGFR-NFATc1 signaling loop which is required for pancreatic cancer initiation.
O-42 Abstract id: 318. Nuclear factor of activated T-cells c1 integrates STAT3 signals to link chronic inflammation and carcinogenesis in the pancreas Sandra Baumgart 1, Nai-ming Chen 1, Marius Brunner 1, Reinecke €nig 1, Jinsan Johanna 1, Julius Nikorowitsch 1, Shiv Singh 1, Alexander Ko Zhang 2, Elmar Wolf 3, Marek Bartkuhn 4, Jens Siveke 5, Irene Esposito 5, Elisabeth Glesel 1, Bettina Geisel 1, Kristina Reutlinger 1, Garima Singh 1, Martin Eilers 3, Thomas Gress 1, Daniel Billadeau 2, Volker Ellenrieder 1. 1
Philipps University of Marburg, Germany Mayo Clinic, United States 3 €rzburg, Germany University of Wu 4 Justus-Liebig University of Giessen, Germany 5 Technical University of Munich, Germany 2
Introduction: Chronic pancreatitis along with activating mutations of the oncogene Kras is a central risk factor for pancreatic cancer development. Inflammation-induced signaling involves nuclear factor of activated T cells (NFAT) pathways. NFATc1 is overexpressed and activated in Krasmutated human pancreatic cancers where it mediates cancer growth stimulation. Aims: This study aims to determine the in vivo role of NFATc1 in KrasG12D-dependent carcinogenesis. Materials & methods: Transgenic mice expressing KrasG12D and NFATc1 in the pancreas and NFATc1 knockout mice were engineered and analyzed in terms of (inflammation-induced) carcinogenesis. Immunohistochemistry, Western blot, qPCR analyses, chromatin studies, ChIP-Seq and genome-wide expression profiling were performed to investigate the mechanisms of NFATc1-KrasG12D cooperation in murine and human tissues and tumor cells. Results: Activation of NFATc1 - upon inflammation and in KrasG12D;NFATc1 mice - dramatically accelerated carcinogenesis and reduced survival. Mechanistically, nuclear NFATc1 activated expression of oncogenic STAT3 transcription factor. Accordingly, high correlative nuclear expression levels of NFATc1 and active STAT3 were detected in the majority of human and murine cancer tissues. In turn, NFATc1/STAT3 complexes, which regulated genome-wide NFATc1 binding to distal chromatin enhancer sites and subsequently enforced recruitment of transcriptional co-activators, were identified. Likewise, corresponding enhancer-promoter communications that stimulated NFATc1-dependent transcription of newly-identified target genes were epigenetically promoted. Pharmacologic and genetic depletion of the NFATc1/STAT3-axis significantly arrested carcinogenesis in mouse models and confirmed the requirement of NFATc1 in Kras-driven pancreatic carcinogenesis. Conclusion: We identified a previously unknown NFATc1-STAT3 complex formation as a driving epigenetic regulatory force in inflammation-linked pancreatic carcinogenesis, which defines this novel pathway a possible point of therapeutic intervention.
O-43 Abstract id: 236. Therapeutic interest to target the Phosphoinositide-3-Kinase (PI3K) – mTOR pathway in pancreatic Cancer-Associated Fibroblasts Camille Duluc 1, Siham Moatassim-Billah 2, Herbert Schmid 3, Marieephane Pyronnet 1, Julie Bernadette Delisle 4, Barbara Garmy-Susini 1, St Guillermet-Guibert 1, Corinne Bousquet 1. 1
INSERM U1037, France Faculty of Sciences Rabat, University Mohammed V - Agdal, Morocco 3 Novartis, Bales, Switzerland 4 Pathology Toulouse Hospital, France 2
Introduction: Pancreatic ductal adenocarcinoma (PDAC) is one of the most stroma-rich cancers. Cancer-associated fibroblasts (CAFs), the most abundant cells in this stroma, orchestrate the secretion of growth factors that engage, in cancer and other stromal cells, survival and angiogenic signals redundant to those targeted by therapies, and of fibrillar components that constitute a barrier to drug delivery. Aims: Therapeutic pitfalls observed in PDAC may in part be explained by the under-estimation of the influences exerted by the microenvironment on cancer cells. Targeting CAFs may therefore constitute a promising strategy. Patients & methods: Primary cultures of CAFs have been isolated from human pancreatic tumor resections and used for in vitro and in vivo studies. Results: Isolated CAFs present, and maintain during the in vitro passages (< 10), an activated phenotype ( aSMA-positive). CAFs promote survival, invasion and chemoprotection (against gemcitabine) of pancreatic cancer cells, either in co-cultures or using CAF conditioned media. Interestingly, CAFs present a high intrinsic activation of the PI3K-mTOR pathway. It results in elevated protein synthesis and secretion specifically of pro-tumoral factors and extracellular matrix proteins. Strikingly, drugs that inhibit the PI3K-mTOR pathway abrogate CAF pro-tumoral and chemoprotective effects on cancer cells both in vitro and in vivo, in co-xenografted mice with CAF and pancreatic cancer cells. Conclusion: Because CAFs are present both in the tumor and in the periphery where drug delivery is still feasible, therapeutic targeting of CAFs using inhibitors of protein synthesis may be of utmost interest for PDAC.
O-44 Abstract id: 187. Neural invasion in pancreatic cancer is characterized by beta-1-Integrin- and L1-CAM-dependent heterotypic cell adhesion between pancreatic cancer cells and neural Schwann cells € ralp O. Ihsan Ekin Demir, Natascha Klose, Eva Brunner, Helmut Friess, Gu Ceyhan. € nchen, Department of Surgery, Klinikum rechts der Isar, TU Mu Germany Introduction: Neural invasion is one of the most frequent modes of tumor spread in pancreatic cancer (PCa). However, how PCa cells can attach to and migrate along Schwann-cell-covered axons in nerves remains unclear. Aims: The aim of this study was to investigate the potential heterotypic cell adhesion between PCa cells and Schwann cells (SC) and to identify the surface molecules that enable this interaction. Materials & methods: Human SC, PCa cells, human PCa tissue, normal human pancreas and intrapancreatic nerves were investigated for the expression of the cell adhesion molecules beta-1-Integrin, NCAM, L1-CAM and NrCAM via immunoblotting, QRT-PCR and immunohistochemistry. Heterotypic cell adhesion and mutual migration between SC and PCa cells were quantified via a recently established adhesion assay and 3D migration assay by applying neutralizing antibodies against these adhesion molecules. Results: Expression of beta1-Integrin and L1-CAM were more prevalent in PCa tissues, human SC, intrapancreatic nerves and PCa cells than NCAM and NrCAM. Confrontation of SC with PCa cells led to rapid heterotypic cell adhesion, which could be inhibited by neutralizing antibodies against beta-1-Integrin and L1-CAM, but not against NCAM and NrCAM on PCa cells. Similarly, blockade of beta-1-Integrin on PCa cells diminished the carcinotropic migration of SC. Interestingly, inhibition of these molecues on SC did not influence the adhesion or carcinotropic migration of SC. Conclusion: Neural invasion harbors heterotypic cell adhesion and mutual migration between SC and PCa cells. Therefore, cell-cell-adhesion represents a key pathophysiological mechanism in neural invasion and for the associated local tumor recurrence and neuropathic pain in PCa.