ceruletide and Carcinoma--Pancreatic-Ductal

Polo-like kinase 1 (Plk1) plays an important role in cell-cycle regulation. Recent work has suggested that Plk1 could be a biomarker of gemcitabine response in pancreatic ductal adenocarcinoma (PDAC). Although targeting Plk1 to treat PDAC has been attempted in clinical trials, the results were not promising, and the mechanisms of resistance to Plk1 inhibition is poorly understood. In addition, the role of Plk1 in PDAC progression requires further elucidation. Here, we showed that Plk1 was associated with poor outcomes in patients with PDAC. In an inducible transgenic mouse line with specific expression of Plk1 in the pancreas, Plk1 overexpression significantly inhibited caerulein-induced acute pancreatitis and delayed development of acinar-to-ductal metaplasia and pancreatic intraepithelial neoplasia. Bioinformatics analyses identified the regulatory networks in which Plk1 is involved in PDAC disease progression, including multiple inflammation-related pathways. Unexpectedly, inhibition or depletion of Plk1 resulted in upregulation of PD-L1 via activation of the NF-κB pathway. Mechanistically, Plk1-mediated phosphorylation of RB at S758 inhibited the translocation of NF-κB to nucleus, inactivating the pathway. Inhibition of Plk1 sensitized PDAC to immune checkpoint blockade therapy through activation of an antitumor immune response. Together, Plk1 suppresses PDAC progression and inhibits NF-κB activity, and targeting Plk1 can potentiate the efficacy of immunotherapy in PDAC.. Inhibition of Plk1 induces upregulation of PD-L1 expression in pancreatic ductal adenocarcinoma, stimulating antitumor immunity and sensitizing tumors to immunotherapy.

Topics: Acute Disease; Animals; B7-H1 Antigen; Carcinoma, Pancreatic Ductal; Cell Cycle Proteins; Ceruletide; Humans; Immune Checkpoint Inhibitors; Mice; NF-kappa B; Pancreatic Neoplasms; Pancreatitis; Polo-Like Kinase 1; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins

Topics: Alcohol Drinking; Animals; Carcinogens; Carcinoma, Pancreatic Ductal; Central Nervous System Depressants; Ceruletide; Cytokines; Diet, Western; Ethanol; Hepatocytes; Homeodomain Proteins; Liver Neoplasms; Mice; Mutation; Nicotine; Pancreatic Neoplasms; Proto-Oncogene Proteins p21(ras); Trans-Activators

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

Topics: Animals; Carcinogenesis; Carcinoma, Pancreatic Ductal; Cells, Cultured; Ceruletide; Homeodomain Proteins; Metaplasia; Mice; Mice, Transgenic; NF-kappa B; Pancreatitis; Proto-Oncogene Proteins p21(ras); Resveratrol; Signal Transduction; Trans-Activators

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

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

Pancreatitis is a debilitating disease of the exocrine pancreas that, under chronic conditions, is a major susceptibility factor for pancreatic ductal adenocarcinoma (PDAC). Although down-regulation of genes that promote the mature acinar cell fate is required to reduce injury associated with pancreatitis, the factors that promote this repression are unknown. Activating transcription factor 3 (ATF3) is a key mediator of the unfolded protein response, a pathway rapidly activated during pancreatic insult. Using chromatin immunoprecipitation followed by next-generation sequencing, we show that ATF3 is bound to the transcriptional regulatory regions of >30% of differentially expressed genes during the initiation of pancreatitis. Of importance, ATF3-dependent regulation of these genes was observed only upon induction of pancreatitis, with pathways involved in inflammation, acinar cell differentiation, and cell junctions being specifically targeted. Characterizing expression of transcription factors that affect acinar cell differentiation suggested that acinar cells lacking ATF3 maintain a mature cell phenotype during pancreatitis, a finding supported by maintenance of junctional proteins and polarity markers. As a result,

Topics: Acinar Cells; Activating Transcription Factor 3; Animals; Carcinoma, Pancreatic Ductal; Cell Differentiation; Ceruletide; Down-Regulation; Male; Mice; Mice, Knockout; Pancreatic Neoplasms; Pancreatitis; Phenotype

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

Pancreatic ductal adenocarcinoma (PDAC) is characterized by an exuberant inflammatory desmoplastic response. The PDAC microenvironment is complex, containing both pro- and antitumorigenic elements, and remains to be fully characterized. Here, we show that sensory neurons, an under-studied cohort of the pancreas tumor stroma, play a significant role in the initiation and progression of the early stages of PDAC. Using a well-established autochthonous model of PDAC (PKC), we show that inflammation and neuronal damage in the peripheral and central nervous system (CNS) occurs as early as the pancreatic intraepithelial neoplasia (PanIN) 2 stage. Also at the PanIN2 stage, pancreas acinar-derived cells frequently invade along sensory neurons into the spinal cord and migrate caudally to the lower thoracic and upper lumbar regions. Sensory neuron ablation by neonatal capsaicin injection prevented perineural invasion (PNI), astrocyte activation, and neuronal damage, suggesting that sensory neurons convey inflammatory signals from Kras-induced pancreatic neoplasia to the CNS. Neuron ablation in PKC mice also significantly delayed PanIN formation and ultimately prolonged survival compared with vehicle-treated controls (median survival, 7.8 vs. 4.5 mo; P = 0.001). These data establish a reciprocal signaling loop between the pancreas and nervous system, including the CNS, that supports inflammation associated with oncogenic Kras-induced neoplasia. Thus, pancreatic sensory neurons comprise an important stromal cell population that supports the initiation and progression of PDAC and may represent a potential target for prevention in high-risk populations.

Topics: Adenocarcinoma in Situ; Afferent Pathways; Animals; Animals, Newborn; Capsaicin; Carcinoma, Pancreatic Ductal; Ceruletide; Denervation; Disease Progression; Female; Ganglia, Sympathetic; Genes, ras; Humans; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Myelitis; Neoplasm Invasiveness; Pancreas; Pancreatic Neoplasms; Pancreatitis; Precancerous Conditions; Sensory Receptor Cells; Spinal Cord; Spinothalamic Tracts; Thoracic Vertebrae

Surface CD24 has previously been described, together with CD44 and ESA, for the characterization of putative cancer stem cells in pancreatic ductal adenocarcinoma (PDAC), the most fatal of all solid tumors. CD24 has a variety of biological functions including the regulation of invasiveness and cell proliferation, depending on the tumor entity and subcellular localization. Genetically engineered mouse models (GEMM) expressing oncogenic KrasG12D recapitulate the human disease and develop PDAC. In this study we investigate the function of CD24 using GEMM of endogenous PDAC and a model of cerulein-induced acute pancreatitis. We found that (i) CD24 expression was upregulated in murine and human PDAC and during acute pancreatitis (ii) CD24 was expressed exclusively in differentiated PDAC, whereas CD24 absence was associated with undifferentiated tumors and (iii) membranous CD24 expression determines tumor subpopulations with an epithelial phenotype in grafted models. In addition, we show that CD24 protein is stabilized in response to WNT activation and that overexpression of CD24 in pancreatic cancer cells upregulated β-catenin expression augmenting an epithelial, non-metastatic signature. Our results support a positive feedback model according to which (i) WNT activation and subsequent β-catenin dephosphorylation stabilize CD24 protein expression, and (ii) sustained CD24 expression upregulates β-catenin expression. Eventually, membranous CD24 augments the epithelial phenotype of pancreatic tumors. Thus we link the WNT/β-catenin pathway with the regulation of CD24 in the context of PDAC differentiation.

Topics: Animals; Carcinoma, Pancreatic Ductal; CD24 Antigen; Cell Differentiation; Cell Membrane; Cell Proliferation; Ceruletide; Epithelial-Mesenchymal Transition; Epithelium; Gene Expression Regulation, Neoplastic; Humans; Mice; Mice, Knockout; Mice, SCID; Neoplasm Transplantation; Pancreatic Neoplasms; Pancreatitis; Phenotype; Phosphorylation; Proto-Oncogene Proteins p21(ras); Up-Regulation

Perturbation of pancreatic acinar cell state can lead to acinar-to-ductal metaplasia (ADM), a precursor lesion to the development of pancreatic ductal adenocarcinoma (PDAC). In the pancreas, Notch signaling is active both during development and in adult cellular differentiation processes. Hes1, a key downstream target of the Notch signaling pathway, is expressed in the centroacinar compartment of the adult pancreas as well as in both preneoplastic and malignant lesions. In this study, we used a murine genetic in vivo approach to ablate Hes1 in pancreatic progenitor cells (Ptf1a

Topics: Acinar Cells; Animals; Carcinogenesis; Carcinoma, Pancreatic Ductal; Cell Differentiation; Cell Plasticity; Ceruletide; Disease Models, Animal; Female; Humans; Male; Metaplasia; Mice; Pancreas; Pancreas, Exocrine; Pancreatic Neoplasms; Pancreatitis; Regeneration; Signal Transduction; Stem Cells; Transcription Factor HES-1

Pancreatic cancer is one of the most aggressive cancers and has an extremely poor prognosis. Despite recent progress in both basic and clinical research, most pancreatic cancers are detected at an incurable stage owing to the absence of disease-specific symptoms. Thus, developing novel approaches for detecting pancreatic cancer at an early stage is imperative. Our in silico and immunohistochemical analyses showed that KIAA1199 is specifically expressed in human pancreatic cancer cells and pancreatic intraepithelial neoplasia, the early lesion of pancreatic cancer, in a genetically engineered mouse model and in human patient samples. We also detected secreted KIAA1199 protein in blood samples obtained from pancreatic cancer mouse models, but not in normal mice. Furthermore, we found that assessing KIAA1199 autoantibody increased the sensitivity of detecting pancreatic cancer. These results indicate the potential benefits of using KIAA1199 as a biomarker for early-stage pancreatic cancer.

Topics: Acute Disease; Animals; Autoantibodies; Biomarkers, Tumor; Carcinoma, Pancreatic Ductal; Cell Line, Tumor; Ceruletide; Databases, Genetic; Disease Models, Animal; Early Diagnosis; Gene Expression Profiling; Gene Expression Regulation, Neoplastic; Humans; Hyaluronoglucosaminidase; Male; Mice; Mice, Inbred C57BL; Pancreatic Neoplasms; Pancreatitis; Proteins; Tissue Array Analysis

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

Recent development of genetically engineered mouse models (GEMs) for pancreatic cancer (PC) that recapitulates human disease progression has helped to identify new strategies to delay/inhibit PC development. We first found that expression of the pancreatic tumor-initiating/cancer stem cells (CSC) marker DclK1 occurs in early stage PC and in both early and late pancreatic intraepithelial neoplasia (PanIN) and that it increases as disease progresses in GEM and also in human PC. Genome-wide next generation sequencing of pancreatic ductal adenocarcinoma (PDAC) from GEM mice revealed significantly increased DclK1 along with inflammatory genes. Genetic ablation of cyclo-oxygenase-2 (COX-2) decreased DclK1 in GEM. Induction of inflammation/pancreatitis with cerulein in GEM mice increased DclK1, and the novel dual COX/5-lipoxygenase (5-LOX) inhibitor licofelone reduced it. Dietary licofelone significantly inhibited the incidence of PDAC and carcinoma in situ with significant inhibition of pancreatic CSCs. Licofelone suppressed pancreatic tumor COX-2 and 5-LOX activities and modulated miRNAs characteristic of CSC and inflammation in correlation with PDAC inhibition. These results offer a preclinical proof of concept to target the inflammation initiation to inhibit cancer stem cells early for improving the treatment of pancreatic cancers, with immediate clinical implications for repositioning dual COX/5-LOX inhibitors in human trials for high risk patients.

Topics: Animals; Anti-Inflammatory Agents; Apoptosis; Arachidonic Acid; Carcinoma in Situ; Carcinoma, Pancreatic Ductal; Cell Proliferation; Ceruletide; Cyclooxygenase 2; Disease Models, Animal; Disease Progression; Doublecortin-Like Kinases; Lipoxygenase Inhibitors; Mice; Mice, Knockout; MicroRNAs; Neoplastic Stem Cells; Pancreatic Neoplasms; Pancreatitis; Protein Serine-Threonine Kinases; Pyrroles

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal human malignancies, with a poor long-term prognosis, and effective therapeutic options are lacking. Observing the dynamics of the pathogenesis of pancreatic intraepithelial neoplasia (PanIN) and PDAC in tumor models can facilitate understanding of the molecular mechanisms involved in early PDAC. Furthermore, it can compensate for the research limitations associated with analyzing clinical specimens of late-stage PDAC. In this study, we orthotopically treated the pancreas with dimethylbenzanthracene (DMBA) combined with caerulein in wild-type C57BL/6 J mice to induce inflammation-related pancreatic carcinogenesis. We observed that DMBA and caerulein treatment induced a chronic consumptive disease, which caused a decrease in the relative body and pancreas weights, diminishing the health status of the mice and enhancing the inflammation-related histological changes. Moreover, mid-dose and high-frequency treatment with caerulein caused prolonged inflammatory damage to the pancreas and contributed to a permissive environment for the development of PDAC. CXCL12/CXCR4, CCL2/CCR2, and several cytokines, such as interleukin (IL)-1β, IL-6, and tumor necrosis factor (TNF)-α were upregulated in the tumor tissue of DMBA and caerulein-induced PDAC mice. This orthotopic mouse pancreatic carcinogenesis model mimic human disease because it reproduces a spectrum of pathological changes observed in human PDAC, ranging from inflammatory lesions to pancreatic intraepithelial neoplasia. Thus, this mouse model may improve the understanding of molecular mechanisms underlying the injury-inflammation-cancer pathway in the early stages of pancreatic carcinogenesis.

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; Carcinogenesis; Carcinoma, Pancreatic Ductal; Ceruletide; Female; Inflammation; Interleukin-1beta; Interleukin-6; Mice; Mice, Inbred C57BL; Pancreatic Neoplasms; Tumor Necrosis Factor-alpha; Up-Regulation

Nr5a2 participates in biliary acid metabolism and is a major regulator of the pancreatic exocrine programme. Single nucleotide polymorphisms in the vicinity of NR5A2 are associated with the risk of pancreatic ductal adenocarcinoma (PDAC).. To determine the role of Nr5a2 in pancreatic homeostasis, damage-induced regeneration and mutant KRas-driven pancreatic tumourigenesis.. Nr5a2+/- and KRas(G12V);Ptf1a-Cre;Nr5a2+/- mice were used to investigate whether a full dose of Nr5a2 is required for normal pancreas development, recovery from caerulein-induced pancreatitis, and protection from tumour development.. Adult Nr5a2+/- mice did not display histological abnormalities in the pancreas but showed a more severe acute pancreatitis, increased acino-ductal metaplasia and impaired recovery from damage. This was accompanied by increased myeloid cell infiltration and proinflammatory cytokine gene expression, and hyperactivation of nuclear factor κb and signal transducer and activator of transcription 3 signalling pathways. Induction of multiple episodes of acute pancreatitis was associated with more severe damage and delayed regeneration. Inactivation of one Nr5a2 allele selectively in pancreatic epithelial cells was sufficient to cause impaired recovery from pancreatitis. In comparison with Nr5a2+/+ mice, KRas(G12V);Ptf1a(Cre/+);Nr5a2+/- mice showed a non-statistically significant increase in the area affected by preneoplastic lesions. However, a single episode of acute pancreatitis cooperated with loss of one Nr5a2 allele to accelerate KRas(G12V)-driven development of preneoplastic lesions.. A full Nr5a2 dose is required to restore pancreatic homeostasis upon damage and to suppress the KRas(G12V)-driven mouse pancreatic intraepithelial neoplasia progression, indicating that Nr5a2 is a novel pancreatic tumour suppressor. Nr5a2 could contribute to PDAC through a role in the recovery from pancreatitis-induced damage.

Topics: Animals; Blotting, Western; Carcinoma, Pancreatic Ductal; Ceruletide; Heterozygote; Mice; Mice, Inbred C57BL; Mice, Inbred Strains; Mice, Knockout; NF-kappa B; Pancreatic Neoplasms; Pancreatitis; Polymerase Chain Reaction; Proto-Oncogene Proteins p21(ras); Receptors, Cytoplasmic and Nuclear; Signal Transduction; STAT3 Transcription Factor

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

Gene expression is affected by modifications to histone core proteins within chromatin. Changes in these modifications, or epigenetic reprogramming, can dictate cell fate and promote susceptibility to disease. The goal of this study was to determine the extent of epigenetic reprogramming in response to chronic stress that occurs following ablation of MIST1 (Mist1(-/-) ), which is repressed in pancreatic disease. Chromatin immunoprecipitation for trimethylation of lysine residue 4 on histone 3 (H3K4Me3) in purified acinar cells from wild type and Mist1(-/-) mice was followed by Next Generation sequencing (ChIP-seq) or ChIP-qPCR. H3K4Me3-enriched genes were assessed for expression by qRT-PCR in pancreatic tissue before and after induction of cerulein-induced pancreatitis. While most of H3K4Me3-enrichment is restricted to transcriptional start sites, >25% of enrichment sites are found within, downstream or between annotated genes. Less than 10% of these sites were altered in Mist1(-/-) acini, with most changes in H3K4Me3 enrichment not reflecting altered gene expression. Ingenuity Pathway Analysis of genes differentially-enriched for H3K4Me3 revealed an association with pancreatitis and pancreatic ductal adenocarcinoma in Mist1(-/-) tissue. Most of these genes were not differentially expressed but several were readily induced by acute experimental pancreatitis, with significantly increased expression in Mist1(-/-) tissue relative to wild type mice. We suggest that the chronic cell stress observed in the absence of MIST1 results in epigenetic reprogramming of genes involved in promoting pancreatitis to a poised state, thereby increasing the sensitivity to events that promote disease.

Topics: Acinar Cells; Animals; Basic Helix-Loop-Helix Transcription Factors; Carcinoma, Pancreatic Ductal; Ceruletide; Chromatin; Epigenesis, Genetic; Histones; Male; Metabolic Networks and Pathways; Methylation; Mice; Mice, Knockout; Pancreas; Pancreatic Neoplasms; Pancreatitis

Pancreas cancer, or pancreatic ductal adenocarcinoma, is the deadliest of solid tumors, with a five-year survival rate of <5%. Detection of resectable disease improves survival rates, but access to tissue and other biospecimens that could be used to develop early detection markers is confounded by the insidious nature of pancreas cancer. Mouse models that accurately recapitulate the human condition allow disease tracking from inception to invasion and can therefore be useful for studying early disease stages in which surgical resection is possible. Using a highly faithful mouse model of pancreas cancer in conjunction with a high-density antibody microarray containing ∼2500 antibodies, we interrogated the pancreatic tissue proteome at preinvasive and invasive stages of disease. The goal was to discover early stage tissue markers of pancreas cancer and follow them through histologically defined stages of disease using cohorts of mice lacking overt clinical signs and symptoms and those with end-stage metastatic disease, respectively. A panel of seven up-regulated proteins distinguishing pancreas cancer from normal pancreas was validated, and their levels were assessed in tissues collected at preinvasive, early invasive, and moribund stages of disease. Six of the seven markers also differentiated pancreas cancer from an experimental model of chronic pancreatitis. The levels of serine/threonine stress kinase 4 (STK4) increased between preinvasive and invasive stages, suggesting its potential as a tissue biomarker, and perhaps its involvement in progression from precursor pancreatic intraepithelial neoplasia to pancreatic ductal adenocarcinoma. Immunohistochemistry of STK4 at different stages of disease revealed a dynamic expression pattern further implicating it in early tumorigenic events. Immunohistochemistry of a panel of human pancreas cancers confirmed that STK4 levels were increased in tumor epithelia relative to normal tissue. Overall, this integrated approach yielded several tissue markers that could serve as signatures of disease stage, including early (resectable), and therefore clinically meaningful, stages.

Topics: Animals; Antibodies; Biomarkers, Tumor; Carcinoma, Pancreatic Ductal; Ceruletide; Diagnosis, Differential; Disease Models, Animal; Disease Progression; Early Diagnosis; Gene Expression Profiling; Gene Expression Regulation, Neoplastic; Humans; Mice; Molecular Sequence Annotation; Neoplasm Proteins; Pancreatic Neoplasms; Pancreatitis, Chronic; Protein Array Analysis; Protein Serine-Threonine Kinases; Proteome; Signal Transduction; Time Factors

Diabetes mellitus type 2 and chronic pancreatitis are regarded as risk factors for pancreatic cancer. Pancreatic duct glands (PDGs) were recently described as a new compartment of the major duct in humans and mice. To evaluate the influence of diabetes and chronic pancreatitis on PDGs, cerulein was injected i.p., repetitively over 10 weeks, in mice exhibiting obesity and a type 2 diabetes-like syndrome (B6.V-Lep(ob/ob)) and in lean littermates. By using 5-bromo-2'-deoxyuridine (BrdU), a label-retaining cell population was characterized in PDGs. Cerulein administration led to more BrdU(+) cells in PDGs of obese mice compared with lean mice. The observed increase was specific to PDGs, because BrdU incorporation in cells of the pancreatic duct was not increased. In addition, the expression of distinct tumor markers in PDGs was characterized by Muc5ac, S100P, regenerating islet-derived 3β, 14-3-3 σ, and prostate stem cell antigen immunochemistry. Type 2 diabetes-like syndrome, accompanied by chronic pancreatitis, enhanced nuclear localization of S100P. Both risk factors for pancreatic cancer also induced the production of Muc5ac and the nuclear localization of S100P [corrected]. These results demonstrate that diabetes and chronic pancreatitis jointly enhance BrdU incorporation and production of pancreatic cancer-specific proteins in PDGs. The observed alterations suggest that pancreatic tumors might originate from the newly discovered histomorphological structures, called PDGs, which could represent a target for future anticancer therapies.

Topics: Animals; Bromodeoxyuridine; Carcinoma, Pancreatic Ductal; Ceruletide; Diabetes Mellitus, Type 2; Humans; Male; Metaplasia; Mice; Mice, Obese; Mucins; Neoplasm Proteins; Pancreatic Ducts; Pancreatic Neoplasms; Pancreatitis, Chronic; Risk Factors

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

Targeting of oncogenic Kras to the pancreatic Nestin-expressing embryonic progenitor cells and subsequently to the adult acinar compartment and Nestin-expressing cells is sufficient for the development of low grade pancreatic intraepithelial neoplasia (PanIN) between 2 and 4 months. The mice die around 6 month-old of unrelated causes, and it is therefore not possible to assess whether the lesions will progress to carcinoma. We now report that two brief episodes of caerulein-induced acute pancreatitis in 2 month-old mice causes rapid PanIN progression and pancreatic ductal adenocarcinoma (PDAC) development by 4 months of age. These events occur with similar frequency as observed in animals where the oncogene is targeted during embryogenesis to all pancreatic cell types. Thus, these data show that oncogenic Kras-driven PanIN originating in a non-ductal compartment can rapidly progress to PDAC when subjected to a brief inflammatory insult.

Topics: Animals; Carcinoma in Situ; Carcinoma, Pancreatic Ductal; Cell Lineage; Ceruletide; Disease Progression; Gene Targeting; Humans; Integrases; Intermediate Filament Proteins; Mice; Mice, Transgenic; Nerve Tissue Proteins; Nestin; Pancreatic Ducts; Pancreatic Neoplasms; Pancreatitis; Precancerous Conditions; Proto-Oncogene Proteins p21(ras); STAT3 Transcription Factor; Stem Cells; Transgenes

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