ceruletide and Cell-Transformation--Neoplastic

The underlying molecular mechanisms of chronic pancreatitis (CP) developing into pancreatic ductal adenocarcinoma (PDAC) remain largely unknown. Here we show that the level of serotonin in mouse pancreatic tissues is upregulated in caerulein-induced CP mice. In vitro study demonstrates that serotonin promotes the formation of acinar-to-ductal metaplasia (ADM) and the activation of pancreatic stellate cells (PSCs), which results from the activation of RhoA/ROCK signaling cascade. Activation of this signaling cascade increases NF-κB nuclear translocation and α-SMA expression, which further enhance the inflammatory responses and fibrosis in pancreatic tissues. Intriguingly, quercetin inhibits both ADM lesion and PSCs activation in vitro and in vivo via its inhibitory effect on serotonin release. Our findings underscore the instrumental role of serotonin-mediated activation of RhoA/ROCK signaling pathway in development of PDAC from CP and highlight a potential to impede PDAC development by disrupting tumor-promoting functions of serotonin.

Topics: Acinar Cells; Animals; Biomarkers; Carcinoma, Pancreatic Ductal; Cell Transformation, Neoplastic; Ceruletide; Disease Models, Animal; Gene Expression Regulation; Immunohistochemistry; Metaplasia; Mice; Pancreatitis, Chronic; rho-Associated Kinases; rhoA GTP-Binding Protein; Serotonin; Signal Transduction

At the initial stage of carcinogenesis, when RasV12-transformed cells are surrounded by normal epithelial cells, RasV12 cells are apically extruded from epithelia through cell competition with the surrounding normal cells. In this study, we demonstrate that expression of cyclooxygenase (COX)-2 is upregulated in normal cells surrounding RasV12-transformed cells. Addition of COX inhibitor or COX-2-knockout promotes apical extrusion of RasV12 cells. Furthermore, production of Prostaglandin (PG) E

Topics: Animals; Anticarcinogenic Agents; Cell Line, Transformed; Cell Transformation, Neoplastic; Ceruletide; Cyclooxygenase 2; Cyclooxygenase Inhibitors; Dinoprostone; Disease Models, Animal; Dogs; Epithelial Cells; Female; Genes, ras; Ibuprofen; Madin Darby Canine Kidney Cells; Male; Mice, Inbred C57BL; Mice, Transgenic; Pancreatitis; Signal Transduction

Development of pancreatic ductal adenocarcinoma (PDA) involves acinar to ductal metaplasia and genesis of tuft cells. It has been a challenge to study these rare cells because of the lack of animal models. We investigated the role of tuft cells in pancreatic tumorigenesis.. We performed studies with LSL-Kras. Pancreata from KC mice had increased formation of tuft cells and higher levels of prostaglandin D. In mice with KRAS-induced pancreatic tumorigenesis, loss of tuft cells accelerates tumorigenesis and increases the severity of caerulein-induced pancreatic injury, via decreased production of prostaglandin D

Topics: Animals; Carcinoma, Pancreatic Ductal; Cell Transformation, Neoplastic; Ceruletide; Disease Models, Animal; Energy Metabolism; Fibrosis; Humans; Interleukins; Intramolecular Oxidoreductases; Mice, Transgenic; Mutation; Octamer Transcription Factors; Pancreas; Pancreatic Neoplasms; Pancreatitis; Prostaglandin D2; Proto-Oncogene Proteins p21(ras); Time Factors; Transcription Factors

PDA is a major cause of US cancer-related deaths. Oncogenic Kras presents in 90% of human PDAs. Kras mutations occur early in pre-neoplastic lesions but are insufficient to cause PDA. Other contributing factors early in disease progression include chronic pancreatitis, alterations in epigenetic regulators, and tumor suppressor gene mutation. GPCRs activate heterotrimeric G-proteins that stimulate intracellular calcium and oncogenic Kras signaling, thereby promoting pancreatitis and progression to PDA. By contrast, Rgs proteins inhibit Gi/q-coupled GPCRs to negatively regulate PDA progression. Rgs16::GFP is expressed in response to caerulein-induced acinar cell dedifferentiation, early neoplasia, and throughout PDA progression. In genetically engineered mouse models of PDA, Rgs16::GFP is useful for pre-clinical rapid in vivo validation of novel chemotherapeutics targeting early lesions in patients following successful resection or at high risk for progressing to PDA. Cultured primary PDA cells express Rgs16::GFP in response to cytotoxic drugs. A histone deacetylase inhibitor, TSA, stimulated Rgs16::GFP expression in PDA primary cells, potentiated gemcitabine and JQ1 cytotoxicity in cell culture, and Gem + TSA + JQ1 inhibited tumor initiation and progression in vivo. Here we establish the use of Rgs16::GFP expression for testing drug combinations in cell culture and validation of best candidates in our rapid in vivo screen.

Topics: Acinar Cells; Adenocarcinoma; Animals; Antineoplastic Agents; Calcium; Carcinogenesis; Carcinoma, Pancreatic Ductal; Cell Dedifferentiation; Cell Transformation, Neoplastic; Cells, Cultured; Ceruletide; Deoxycytidine; Disease Progression; Gemcitabine; GTP-Binding Proteins; Histone Deacetylase Inhibitors; Mice; Pancreatic Ducts; Pancreatic Neoplasms; Pancreatitis; Proto-Oncogene Proteins p21(ras); RGS Proteins; Signal Transduction

Pancreatitis is a major risk factor for the development of pancreatic cancer. In genetically engineered mouse models, induction of pancreatic inflammation dramatically accelerates oncogenic KRas-induced fibrosis, precancerous PanIN formation, and tumorigenesis. Here we describe simple methods of secretagogue-induced experimental acute and chronic pancreatitis, the most commonly used pancreatitis models, and their applications in pancreatic cancer research. Additionally, the preparation of primary pancreatic acinar cells is introduced. Primary acinar cells can be used to study the early events of pancreatic inflammation and pancreatic acinar-to-ductal (ADM) metaplasia.

Topics: Acinar Cells; Animals; Carcinogenesis; Cell Transformation, Neoplastic; Ceruletide; Disease Models, Animal; Humans; Metaplasia; Mice; Mice, Transgenic; Mutation; Pancreas; Pancreatic Ducts; Pancreatic Neoplasms; Pancreatitis; Primary Cell Culture; Proto-Oncogene Proteins p21(ras); Tumor Cells, Cultured

Acinar-to-ductal metaplasia (ADM) of the pancreas is a process that pancreatic acinar cells differentiate into ductal-like cells with ductal cell traits. The metaplasia of pancreatic acinar cells manifests their ability to adapt to the genetic and environmental pressure they encounter. However, with oncogenic genetic insults and/or sustained environmental stress, ADM may lead to pancreatic intraepithelial neoplasia (PanIN), which is a common precancerous lesion that precedes pancreatic cancer. Understanding the intermediate states of ADM and important molecules that regulate ADM formation may help the development of novel preventive strategies that could be translated to the clinic to benefit the people with high risk of pancreatic cancer. Mouse model is widely used in both in vivo and ex vivo studies of ADM. In this chapter, we describe detailed protocols of injury models of the adult mouse pancreas that can function as a tool to study mechanisms of ADM formation.

Topics: Acinar Cells; Animals; Carcinoma, Pancreatic Ductal; Cell Transdifferentiation; Cell Transformation, Neoplastic; Ceruletide; Disease Models, Animal; Humans; Metaplasia; Mice; Pancreatic Ducts; Pancreatic Neoplasms; Pancreatitis; Primary Cell Culture; Tumor Cells, Cultured

Topics: Animals; Antineoplastic Agents; Carcinoma, Pancreatic Ductal; Cell Transformation, Neoplastic; Ceruletide; Disease Models, Animal; Disease Progression; Gene Knockdown Techniques; Heterografts; Humans; Metaplasia; Mice; Mice, Inbred C57BL; Mice, Nude; Mice, Transgenic; Naphthoquinones; Pancreas; Pancreatic Neoplasms; Precancerous Conditions; Snail Family Transcription Factors; Tumor Cells, Cultured

Acinar-to-ductal metaplasia (ADM) is a reversible epithelial transdifferentiation process that occurs in the pancreas in response to acute inflammation. ADM can rapidly progress towards pre-malignant pancreatic intraepithelial neoplasia (PanIN) lesions in the presence of mutant KRas and ultimately pancreatic adenocarcinoma (PDAC). In the present work, we elucidate the role and related mechanism of glycogen synthase kinase-3beta (GSK-3β) in ADM development using in vitro 3D cultures and genetically engineered mouse models. We show that GSK-3β promotes TGF-α-induced ADM in 3D cultured primary acinar cells, whereas deletion of GSK-3β attenuates caerulein-induced ADM formation and PanIN progression in Kras

Topics: Acinar Cells; Animals; Carcinoma in Situ; Cell Proliferation; Cell Transdifferentiation; Cell Transformation, Neoplastic; Cells, Cultured; Ceruletide; Disease Models, Animal; Disease Progression; Genetic Predisposition to Disease; Glycogen Synthase Kinase 3 beta; Homeodomain Proteins; Male; Metaplasia; Mice, Knockout; Pancreas, Exocrine; Pancreatic Ducts; Pancreatic Neoplasms; Pancreatitis; Phenotype; Proto-Oncogene Proteins p21(ras); Ribosomal Protein S6 Kinases; Signal Transduction; Time Factors; TOR Serine-Threonine Kinases; Trans-Activators; Tumor Necrosis Factor-alpha

The ability of exocrine pancreatic cells to change the cellular phenotype is required for tissue regeneration upon injury, but also contributes to their malignant transformation and tumor progression. We investigated context-dependent signaling and transcription mechanisms that determine pancreatic cell fate decisions toward regeneration and malignancy. In particular, we studied the function and regulation of the inflammatory transcription factor nuclear factor of activated T cells 1 (NFATC1) in pancreatic cell plasticity and tissue adaptation.. We analyzed cell plasticity during pancreatic regeneration and transformation in mice with pancreas-specific expression of a constitutively active form of NFATC1, or depletion of enhancer of zeste 2 homologue 2 (EZH2), in the context of wild-type or constitutively activate Kras, respectively. Acute and chronic pancreatitis were induced by intraperitoneal injection of caerulein. EZH2-dependent regulation of NFATC1 expression was studied in mouse in human pancreatic tissue and cells by immunohistochemistry, immunoblotting, and quantitative reverse transcription polymerase chain reaction. We used genetic and pharmacologic approaches of EZH2 and NFATC1 inhibition to study the consequences of pathway disruption on pancreatic morphology and function. Epigenetic modifications on the NFATC1 gene were investigated by chromatin immunoprecipitation assays.. NFATC1 was rapidly and transiently induced in early adaptation to acinar cell injury in human samples and in mice, where it promoted acinar cell transdifferentiation and blocked proliferation of metaplastic pancreatic cells. However, in late stages of regeneration, Nfatc1 was epigenetically silenced by EZH2-dependent histone methylation, to enable acinar cell redifferentiation and prevent organ atrophy and exocrine insufficiency. In contrast, oncogenic activation of KRAS signaling in pancreatic ductal adenocarcinoma cells reversed the EZH2-dependent effects on the NFATC1 gene and was required for EZH2-mediated transcriptional activation of NFATC1.. In studies of human and mouse pancreatic cells and tissue, we identified context-specific epigenetic regulation of NFATc1 activity as an important mechanism of pancreatic cell plasticity. Inhibitors of EZH2 might therefore interfere with oncogenic activity of NFATC1 and be used in treatment of pancreatic ductal adenocarcinoma.

Topics: Acinar Cells; Animals; Carcinoma, Pancreatic Ductal; Cell Plasticity; Cell Proliferation; Cell Transdifferentiation; Cell Transformation, Neoplastic; Ceruletide; Cyclin-Dependent Kinase Inhibitor p16; Enhancer of Zeste Homolog 2 Protein; Gene Expression Regulation; Gene Silencing; Histones; Humans; Methylation; Mice; NFATC Transcription Factors; Pancreas; Pancreatic Neoplasms; Pancreatitis, Chronic; Promoter Regions, Genetic; Proto-Oncogene Proteins p21(ras); Regeneration; Signal Transduction; Transcription, Genetic

Determining signalling pathways that regulate pancreatic regeneration following pancreatitis is critical for implementing therapeutic interventions. In this study we elucidated the molecular mechanisms underlying the effects of transforming growth factor-β (TGFβ) in pancreatic epithelial cells during tissue regeneration. To this end, we conditionally inactivated TGFβ receptor II (TGFβ-RII) using a Cre-LoxP system under the control of pancreas transcription factor 1a (PTF1a) promoter, specific for the pancreatic epithelium, and evaluated the molecular and cellular changes in a mouse model of cerulein-induced pancreatitis. We show that TGFβ-RII signalling does not mediate the initial acinar cell damage observed at the onset of pancreatitis. However, TGFβ-RII signalling not only restricts acinar cell replication during the regenerative phase of the disease but also limits ADM formation in vivo and in vitro in a cell-autonomous manner. Analyses of molecular mechanisms underlying the observed phenotype revealed that TGFβ-RII signalling stimulates the expression of cyclin-dependent kinase inhibitors and intersects with the EGFR signalling axis. Finally, TGFβ-RII ablation in epithelial cells resulted in increased infiltration of inflammatory cells in the early phases of pancreatitis and increased activation of pancreatic stellate cells in the later stages of pancreatitis, thus highlighting a TGFβ-based crosstalk between epithelial and stromal cells regulating the development of pancreatic inflammation and fibrosis. Collectively, our data not only contribute to clarifying the cellular processes governing pancreatic tissue regeneration, but also emphasize the conserved role of TGFβ as a tumour suppressor, both in the regenerative process following pancreatitis and in the initial phases of pancreatic cancer.

Topics: Acinar Cells; Amylases; Animals; Carcinoma, Pancreatic Ductal; Cell Cycle Checkpoints; Cell Proliferation; Cell Transformation, Neoplastic; Cells, Cultured; Ceruletide; Epithelial Cells; Fibrosis; Irritants; Lipase; Male; Metaplasia; Mice, Knockout; Mice, Transgenic; Pancreas; Pancreatic Neoplasms; Pancreatitis; Protein Serine-Threonine Kinases; Receptor, Transforming Growth Factor-beta Type II; Receptors, Transforming Growth Factor beta

The current paradigm of pancreatic neoplastic transformation proposes an initial step whereby acinar cells convert into acinar-to-ductal metaplasias, followed by progression of these lesions into neoplasias under sustained oncogenic activity and inflammation. Understanding the molecular mechanisms driving these processes is crucial to the early diagnostic and prevention of pancreatic cancer. Emerging evidence indicates that transcription factors that control exocrine pancreatic development could have either, protective or facilitating roles in the formation of preneoplasias and neoplasias in the pancreas. We previously identified that the homeodomain transcription factor Prox1 is a novel regulator of mouse exocrine pancreas development. Here we investigated whether Prox1 function participates in early neoplastic transformation using in vivo, in vitro and in silico approaches. We found that Prox1 expression is transiently re-activated in acinar cells undergoing dedifferentiation and acinar-to-ductal metaplastic conversion. In contrast, Prox1 expression is largely absent in neoplasias and tumors in the pancreas of mice and humans. We also uncovered that Prox1-heterozygosis markedly increases the formation of acinar-to-ductal-metaplasias and early neoplasias, and enhances features associated with inflammation, in mouse pancreatic tissues expressing oncogenic Kras. Furthermore, we discovered that Prox1-heterozygosis increases tissue damage and delays recovery from inflammation in pancreata of mice injected with caerulein. These results are the first demonstration that Prox1 activity protects pancreatic cells from acute tissue damage and early neoplastic transformation. Additional data in our study indicate that this novel role of Prox1 involves suppression of pathways associated with inflammatory responses and cell invasiveness.

Topics: Acinar Cells; Animals; Cell Transformation, Neoplastic; Ceruletide; Heterozygote; Homeodomain Proteins; Humans; Inflammation; Metaplasia; Mice; Pancreas; Pancreatic Neoplasms; Proto-Oncogene Proteins p21(ras); Tumor Suppressor Proteins

Autophagy promotes tumor progression downstream of oncogenic KRAS, yet also restrains inflammation and dysplasia through mechanisms that remain incompletely characterized. Understanding the basis of this paradox has important implications for the optimal targeting of autophagy in cancer. Using a mouse model of cerulein-induced pancreatitis, we found that loss of autophagy by deletion of Atg5 enhanced activation of the IκB kinase (IKK)-related kinase TBK1 in vivo, associated with increased neutrophil and T-cell infiltration and PD-L1 upregulation. Consistent with this observation, pharmacologic or genetic inhibition of autophagy in pancreatic ductal adenocarcinoma cells, including suppression of the autophagy receptors NDP52 or p62, prolonged TBK1 activation and increased expression of CCL5, IL6, and several other T-cell and neutrophil chemotactic cytokines in vitro Defective autophagy also promoted PD-L1 upregulation, which is particularly pronounced downstream of IFNγ signaling and involves JAK pathway activation. Treatment with the TBK1/IKKε/JAK inhibitor CYT387 (also known as momelotinib) not only inhibits autophagy, but also suppresses this feedback inflammation and reduces PD-L1 expression, limiting KRAS-driven pancreatic dysplasia. These findings could contribute to the dual role of autophagy in oncogenesis and have important consequences for its therapeutic targeting. Cancer Immunol Res; 4(6); 520-30. ©2016 AACR.

Topics: Acute Disease; Adenocarcinoma; Animals; Autophagy; Autophagy-Related Protein 5; B7-H1 Antigen; Benzamides; Cell Transformation, Neoplastic; Ceruletide; Chemokine CCL5; Cytokines; Enzyme Activation; Gene Deletion; Mice; Pancreatic Neoplasms; Pancreatitis; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins p21(ras); Pyrimidines; Signal Transduction; Tumor Cells, Cultured

Chronic pancreatitis is a significant risk factor for pancreatic cancer. Previously, we demonstrated that the pancreatic cancer cells show enhanced expression of collapsin response mediator protein 4 (CRMP4) that strongly correlates with severe venous invasion, liver metastasis, and poor prognosis. However, involvement of CRMP4 in acute or chronic pancreatitis remains unknown.. Acute and chronic pancreatitis mice models were developed by periodic injection of caerulein. The expression levels of CRMP4 and its phosphorylation were examined.. Elevated CRMP4 levels were observed in the infiltrated lymphocytes as well as in the pancreas parenchyma of both acute and chronic pancreatitis. The expression pattern of phosphorylated CRMP4 was similar to that of CRMP4. Cdk5 partially co-localized with the phosphorylated CRMP4.. Pancreatitis induces CRMP4 expression in the pancreas parenchyma and in the infiltrated lymphocytes. Overlapping expression of CRMP4 and Cdk5 may suggest that the Cdk5 is at least, in part, responsible for the phosphorylation of CRMP4. The results suggest that CRMP4 is involved in the inflammatory response in pancreatitis. Understanding the mechanisms of CRMP4 would help us to develop novel therapeutic strategies against acute or chronic pancreatitis, and pancreatic cancer.

Topics: Acute Disease; Animals; Biopsy, Needle; Cell Transformation, Neoplastic; Ceruletide; Chronic Disease; Cyclin-Dependent Kinase 5; Disease Models, Animal; Gene Expression Regulation; Humans; Immunohistochemistry; Mice; Nerve Tissue Proteins; Pancreatic Neoplasms; Pancreatitis; Phosphorylation; Precancerous Conditions; RNA, Small Interfering

Oncogenic mutations in KRAS contribute to the development of pancreatic ductal adenocarcinoma, but are not sufficient to initiate carcinogenesis. Secondary events, such as inflammation-induced signaling via the epidermal growth factor receptor (EGFR) and expression of the SOX9 gene, are required for tumor formation. Herein we sought to identify the mechanisms that link EGFR signaling with activation of SOX9 during acinar-ductal metaplasia, a transdifferentiation process that precedes pancreatic carcinogenesis.. We analyzed pancreatic tissues from Kras(G12D);pdx1-Cre and Kras(G12D);NFATc1(Δ/Δ);pdx1-Cre mice after intraperitoneal administration of caerulein, vs cyclosporin A or dimethyl sulfoxide (controls). Induction of EGFR signaling and its effects on the expression of Nuclear factor of activated T cells c1 (NFATc1) or SOX9 were investigated by quantitative reverse-transcription polymerase chain reaction, immunoblot, and immunohistochemical analyses of mouse and human tissues and acinar cell explants. Interactions between NFATc1 and partner proteins and effects on DNA binding or chromatin modifications were studied using co-immunoprecipitation and chromatin immunoprecipitation assays in acinar cell explants and mouse tissue.. EGFR activation induced expression of NFATc1 in metaplastic areas from patients with chronic pancreatitis and in pancreatic tissue from Kras(G12D) mice. EGFR signaling also promoted formation of a complex between NFATc1 and C-JUN in dedifferentiating mouse acinar cells, leading to activation of Sox9 transcription and induction of acinar-ductal metaplasia. Pharmacologic inhibition of NFATc1 or disruption of the Nfatc1 gene inhibited EGFR-mediated induction of Sox9 transcription and blocked acinar-ductal transdifferentiation and pancreatic cancer initiation in mice.. EGFR signaling induces expression of NFATc1 and Sox9, leading to acinar cell transdifferentiation and initiation of pancreatic cancer. Strategies designed to disrupt this pathway might be developed to prevent pancreatic cancer initiation in high-risk patients with chronic pancreatitis.

Topics: Animals; Carcinoma, Pancreatic Ductal; Cell Line; Cell Transdifferentiation; Cell Transformation, Neoplastic; Ceruletide; Cyclosporine; Disease Models, Animal; ErbB Receptors; Gene Expression Regulation; Humans; Male; Metaplasia; Mice, Inbred C57BL; Mice, Knockout; Mutation; NFATC Transcription Factors; Pancreas, Exocrine; Pancreatic Ducts; Pancreatic Neoplasms; Pancreatitis; Precancerous Conditions; Proto-Oncogene Proteins p21(ras); Signal Transduction; SOX9 Transcription Factor; Tissue Culture Techniques; Transcriptional Activation

Pancreatic ductal adenocarcinoma (PDAC) is the fourth leading cause of cancer death in the Western world. The disease has the worst prognosis in the gastrointestinal malignancies with an overall 5-year survival rate of less than 5 %. Therefore, in the search for novel therapeutic targets, biomarkers for early detection and particularly adequate methods to develop and validate therapeutic strategies for this disease are still in urgent demand. Although significant progress has been achieved in understanding the genetic and molecular mechanisms, most approaches have not yet translated sufficiently for better outcome of the patients. In part, this situation is due to inappropriate or insufficient methods in modeling PDAC in laboratory settings. In the past several years, there has been an explosion of genetically engineered mouse models (GEMM) and patient-derived xenografts (PDX) that recapitulate both genetic and morphological alterations that lead to the development of PDAC. Both models are increasingly used for characterization and validation of diagnostic and therapeutic strategies. In this chapter we will discuss state-of-the-art models to consider when selecting an appropriate in vivo system to study disease etiology, cell signaling, and drug development.

Topics: Animals; Cell Line, Tumor; Cell Separation; Cell Transformation, Neoplastic; Ceruletide; Disease Models, Animal; Female; Humans; Mice; Mice, Transgenic; Pancreatic Neoplasms; Pancreatitis

We describe the first mouse model of pancreatic intraepithelial neoplasia (PanIN) lesions induced by alcohol in the presence and absence of chronic pancreatitis.. Pdx1-Cre;LSL-K-ras mice were exposed to Lieber-DeCarli alcohol diet for 6 weeks with cerulein injections. The PanIN lesions and markers of fibrosis, inflammation, histone deacetylation, epithelial-to-mesenchymal transition (EMT), and cancer stemness were measured by immunohistochemistry and Western.. Exposure of Pdx1-Cre;LSL-K-ras mice to an alcohol diet significantly stimulated fibrosis and slightly but not significantly increased the level of PanIN lesions associated with an increase in tumor-promoting M2 macrophages. Importantly, the alcohol diet did not increase activation of stellate cells. Alcohol diet and cerulein injections resulted in synergistic and additive effects on PanIN lesion and M2 macrophage phenotype induction, respectively. Cerulein pancreatitis caused stellate cell activation, EMT, and cancer stemness in the pancreas. Pancreatitis caused histone deacetylation, which was promoted by the alcohol diet. Pancreatitis increased EMT and cancer stemness markers, which were not further affected by the alcohol diet.. The results suggest that alcohol has independent effects on promotion of PDAC associated with fibrosis formed through a stellate cell-independent mechanism and that it further promotes early PDAC and M2 macrophage induction in the context of chronic pancreatitis.

Topics: Acetylation; Acute Disease; Animals; Carcinoma in Situ; Cell Transformation, Neoplastic; Ceruletide; Disease Models, Animal; Epithelial-Mesenchymal Transition; Ethanol; Fibrosis; Histones; Macrophages; Mice, Transgenic; Neoplastic Stem Cells; Pancreas; Pancreatic Neoplasms; Pancreatic Stellate Cells; Pancreatitis; Pancreatitis, Alcoholic; Pancreatitis, Chronic; Time Factors

Emerging evidence from mouse models suggests that mutant Kras can drive the development of pancreatic ductal adenocarcinoma (PDA) precursors from acinar cells by enforcing ductal de-differentiation at the expense of acinar identity. Recently, human genome-wide association studies have identified NR5A2, a key regulator of acinar function, as a susceptibility locus for human PDA. We investigated the role of Nr5a2 in exocrine maintenance, regeneration and Kras driven neoplasia.. To investigate the function of Nr5a2 in the pancreas, we generated mice with conditional pancreatic Nr5a2 deletion (PdxCre(late); Nr5a2(c/c)). Using this model, we evaluated acinar differentiation, regeneration after caerulein pancreatitis and Kras driven pancreatic neoplasia in the setting of Nr5a2 deletion.. We show that Nr5a2 is not required for the development of the pancreatic acinar lineage but is important for maintenance of acinar identity. Nr5a2 deletion leads to destabilisation of the mature acinar differentiation state, acinar to ductal metaplasia and loss of regenerative capacity following acute caerulein pancreatitis. Loss of Nr5a2 also dramatically accelerates the development of oncogenic Kras driven acinar to ductal metaplasia and PDA precursor lesions.. Nr5a2 is a key regulator of acinar plasticity. It is required for maintenance of acinar identity and re-establishing acinar fate during regeneration. Nr5a2 also constrains pancreatic neoplasia driven by oncogenic Kras, providing functional evidence supporting a potential role as a susceptibility gene for human PDA.

Topics: Animals; Carcinoma, Acinar Cell; Carcinoma, Pancreatic Ductal; Cell Differentiation; Cell Line; Cell Transformation, Neoplastic; Ceruletide; Mice; Pancreatic Neoplasms; Pancreatitis; Proto-Oncogene Proteins p21(ras); Real-Time Polymerase Chain Reaction; Receptors, Cytoplasmic and Nuclear

Genetic mutations that give rise to active mutant forms of Ras are oncogenic and found in several types of tumor. However, such mutations are not clear biomarkers for disease, since they are frequently detected in healthy individuals. Instead, it has become clear that elevated levels of Ras activity are critical for Ras-induced tumorigenesis. However, the mechanisms underlying the production of pathological levels of Ras activity are unclear. Here, we show that in the presence of oncogenic Ras, inflammatory stimuli initiate a positive feedback loop involving NF-κB that further amplifies Ras activity to pathological levels. Stimulation of Ras signaling by typical inflammatory stimuli was transient and had no long-term sequelae in wild-type mice. In contrast, these stimuli generated prolonged Ras signaling and led to chronic inflammation and precancerous pancreatic lesions (PanINs) in mice expressing physiological levels of oncogenic K-Ras. These effects of inflammatory stimuli were disrupted by deletion of inhibitor of NF-κB kinase 2 (IKK2) or inhibition of Cox-2. Likewise, expression of active IKK2 or Cox-2 or treatment with LPS generated chronic inflammation and PanINs only in mice expressing oncogenic K-Ras. The data support the hypothesis that in the presence of oncogenic Ras, inflammatory stimuli trigger an NF-κB-mediated positive feedback mechanism involving Cox-2 that amplifies Ras activity to pathological levels. Because a large proportion of the adult human population possesses Ras mutations in tissues including colon, pancreas, and lung, disruption of this positive feedback loop may be an important strategy for cancer prevention.

Topics: Animals; Carcinoma, Pancreatic Ductal; Cell Transformation, Neoplastic; Ceruletide; Cyclooxygenase 2; Cyclooxygenase 2 Inhibitors; Enzyme Induction; Esters; Feedback, Physiological; Gabexate; Gene Expression Regulation, Neoplastic; Gene Knock-In Techniques; Genes, ras; Guanidines; Humans; I-kappa B Kinase; Inflammation; Inflammation Mediators; Lipopolysaccharides; Mice; Mice, Transgenic; Neoplasm Proteins; NF-kappa B; Pancreas; Pancreatic Neoplasms; Pancreatitis, Chronic; Precancerous Conditions; Proto-Oncogene Proteins p21(ras); Sincalide

Pancreatic ductal adenocarcinoma (PDAC) is a fatal disease without effective chemopreventive or therapeutic approaches. Although the role of oncogenic Kras in initiating development of PDAC is well established, downstream targets of aberrant Ras signaling are poorly understood. Acinar-ductal metaplasia (ADM) appears to be an important prerequisite for development of pancreatic intraepithelial neoplasia (PanIN), a common precursor to PDAC. RAS-related C3 botulinum substrate 1 (Rac1), which controls actin reorganization, can be activated by Ras, is up-regulated in several human cancers, and is required for cerulein-induced morphologic changes in acini. We investigated effects of loss of Rac1 in Kras-induced pancreatic carcinogenesis in mice.. Using a Cre/lox approach, we deleted Rac1 from pancreatic progenitor cells in different mouse models of PDAC and in mice with cerulein-induced acute pancreatitis. Acinar epithelial explants of mutant mice were used to investigate the role of Rac1 in vitro.. Rac1 expression increased in mouse and human pancreatic tumors, particularly in the stroma. Deletion of Rac1 in Kras(G12D)-induced PDAC in mice reduced formation of ADM, PanIN, and tumors and significantly prolonged survival. Pancreatic epithelial metaplasia was accompanied by apical-basolateral redistribution of F-actin, along with basal expression of Rac1. Acinar epithelial explants that lacked Rac1 or that were incubated with inhibitors of actin polymerization had a reduced ability to undergo ADM in 3-dimensional cultures.. In mice, Rac1 is required for early metaplastic changes and neoplasia-associated actin rearrangements in development of pancreatic cancer. Rac1 might be developed as a diagnostic marker or therapeutic target for PDAC.

Topics: Actins; Animals; Carcinoma in Situ; Carcinoma, Pancreatic Ductal; Cell Transformation, Neoplastic; Ceruletide; Genes, ras; Humans; Kaplan-Meier Estimate; Keratin-19; Metaplasia; Mice; Models, Animal; Pancreas; Pancreatic Neoplasms; Pancreatitis; rac1 GTP-Binding Protein; Signal Transduction; Survival Rate

Capsaicin is a major biologically active ingredient of chili peppers. Extensive studies indicate that capsaicin is a cancer-suppressing agent via blocking the activities of several signal transduction pathways including nuclear factor-kappaB, activator protein-1 and signal transducer and activator of transcription 3. However, there is little study on the effect of capsaicin on pancreatic carcinogenesis. In the present study, the effect of capsaicin on pancreatitis and pancreatic intraepithelial neoplasia (PanIN) was determined in a mutant Kras-driven and caerulein-induced pancreatitis-associated carcinogenesis in LSL-Kras(G12D)/Pdx1-Cre mice. Forty-five LSL-Kras(G12D)/Pdx1-Cre mice and 10 wild-type mice were subjected to one dose of caerulein (250 μg/kg body wt, intraperitoneally) at age 4 weeks to induce and synchronize the development of chronic pancreatitis and PanIN lesions. One week after caerulein induction, animals were randomly distributed into three groups and fed with either AIN-76A diet, AIN-76A diet containing 10 p.p.m. capsaicin or 20 p.p.m. capsaicin for a total of 8 weeks. The results showed that capsaicin significantly reduced the severity of chronic pancreatitis, as determined by evaluating the loss of acini, inflammatory cell infiltration and stromal fibrosis. PanIN formation was frequently observed in the LSL-Kras(G12D)/Pdx1-Cre mice. The progression of PanIN-1 to high-grade PanIN-2 and -3 were significantly inhibited by capsaicin. Further immunochemical studies revealed that treatment with 10 and 20 p.p.m. capsaicin significantly reduced proliferating cell nuclear antigen-labeled cell proliferation and suppressed phosphorylation of extracellular signal-regulated kinase (ERK) and c-Jun as well blocked Hedgehog/GLI pathway activation. These results indicate that capsaicin could be a promising agent for the chemoprevention of pancreatic carcinogenesis, possibly via inhibiting pancreatitis and mutant Kras-led ERK activation.

Topics: Animals; Blotting, Western; Capsaicin; Capsicum; Carcinoma in Situ; Cell Transformation, Neoplastic; Ceruletide; Extracellular Signal-Regulated MAP Kinases; Female; Homeodomain Proteins; Immunoenzyme Techniques; Integrases; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Pancreatic Neoplasms; Pancreatitis, Chronic; Proto-Oncogene Proteins p21(ras); Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Sensory System Agents; Trans-Activators; Transcription Factor AP-1

Although both endocrine and the exocrine pancreas display a significant capacity for tissue regeneration and renewal, the existence of progenitor cells in the adult pancreas remains uncertain. Using a model of cerulein-mediated injury and repair, we demonstrate that mature exocrine cells, defined by expression of an Elastase1 promoter, actively contribute to regenerating pancreatic epithelium through formation of metaplastic ductal intermediates. Acinar cell regeneration is associated with activation of Hedgehog (Hh) signaling, as assessed by up-regulated expression of multiple pathway components, as well as activation of a Ptch-lacZ reporter allele. Using both pharmacologic and genetic techniques, we also show that the ability of mature exocrine cells to accomplish pancreatic regeneration is impaired by blockade of Hh signaling. Specifically, attenuated regeneration in the absence of an intact Hh pathway is characterized by persistence of metaplastic epithelium expressing markers of pancreatic progenitor cells, suggesting an inhibition of redifferentiation into mature exocrine cells. Given the known role of Hh signaling in exocrine pancreatic cancer, these findings may provide a mechanistic link between injury-induced activation of pancreatic progenitors and subsequent pancreatic neoplasia.

Topics: Animals; Cell Differentiation; Cell Proliferation; Cell Transformation, Neoplastic; Ceruletide; Disease Models, Animal; Epithelial Cells; Genes, Reporter; Hedgehog Proteins; Intermediate Filament Proteins; Metaplasia; Mice; Mice, Inbred C57BL; Mice, Transgenic; Nerve Tissue Proteins; Nestin; Pancreas, Exocrine; Pancreatic Ducts; Pancreatic Elastase; Pancreatitis; Receptors, G-Protein-Coupled; Regeneration; Signal Transduction; Smoothened Receptor; Stem Cells; Time Factors; Veratrum Alkaloids

In this issue of Cancer Cell, Guerra and colleagues provide important new insights regarding the ability of specific pancreatic cell types to generate invasive pancreatic cancer. First, they demonstrate that classical pancreatic "ductal" neoplasia can be induced by activation of oncogenic Kras in nonductal exocrine cells. Second, they show that, while Kras activation in immature acinar and centroacinar cells is readily able to induce ductal neoplasia, Kras-mediated tumorigenesis in mature exocrine pancreas requires the induction of chronic epithelial injury. The results shed new light on the "cell of origin" of pancreatic ductal cancer and demonstrate that chronic pancreatitis provides a permissive environment for Kras-induced pancreatic neoplasia.

Topics: Animals; Carcinoma in Situ; Carcinoma, Pancreatic Ductal; Cell Lineage; Cell Transformation, Neoplastic; Ceruletide; Genes, ras; Humans; Mice; Mutation; Neoplasm Invasiveness; Pancreatic Neoplasms; Pancreatitis, Chronic

Pancreatic ductal adenocarcinoma (PDA), one of the deadliest human cancers, often involves somatic activation of K-Ras oncogenes. We report that selective expression of an endogenous K-Ras(G12V) oncogene in embryonic cells of acinar/centroacinar lineage results in pancreatic intraepithelial neoplasias (PanINs) and invasive PDA, suggesting that PDA originates by differentiation of acinar/centroacinar cells or their precursors into ductal-like cells. Surprisingly, adult mice become refractory to K-Ras(G12V)-induced PanINs and PDA. However, if these mice are challenged with a mild form of chronic pancreatitis, they develop the full spectrum of PanINs and invasive PDA. These observations suggest that, during adulthood, PDA stems from a combination of genetic (e.g., somatic K-Ras mutations) and nongenetic (e.g., tissue damage) events.

Topics: Animals; Carcinoma in Situ; Carcinoma, Pancreatic Ductal; Cell Lineage; Cell Transformation, Neoplastic; Ceruletide; Doxycycline; Genes, ras; Liver Neoplasms; Lung Neoplasms; Mice; Mice, Mutant Strains; Mutation; Neoplasm Invasiveness; Pancreas; Pancreatic Neoplasms; Pancreatitis, Chronic; Signal Transduction