ARID1A Maintains Differentiation of Pancreatic Ductal Cells and Inhibits Development of Pancreatic Ductal Adenocarcinoma in Mice

ABSTRACTPancreatic cancer (PC) is acquired postnatally; to mimic this scenario, we developed an inducible KrasG12D; Ptf1a-CreER™ (iKC) mouse model, in which Kras is activated postnatally at week 16 upon tamoxifen (TAM) administration. Upon TAM treatment, iKC mice develop pancreatic intraepithelial neoplasia (PanIN) lesions and PC with metastasis at the fourth and fortieth weeks, respectively, and exhibited acinar-to-ductal metaplasia (ADM) and transdifferentiation. Kras activation upregulated the transcription factors Ncoa3, p-cJun and FoxM1, which in turn upregulated expression of transmembrane mucins (Muc1, Muc4 and Muc16) and secretory mucin (Muc5Ac). Interestingly, knockdown of KrasG12D in multiple PC cell lines resulted in downregulation of MUC1, MUC4, MUC5AC and MUC16. In addition, iKC mice exhibited ADM and transdifferentiation. Our results show that the iKC mouse more closely mimics human PC development and can be used to investigate pancreatic ductal adenocarcinoma (PDAC) biomarkers, early onset of PDAC, and ADM. The iKC model can also be used for preclinical strategies such as targeting mucin axis alone or in combination with neo-adjuvant, immunotherapeutic approaches and to monitor chemotherapy response.

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Expression of the axon guidance protein Robo1 in pancreatic ductal adenocarcinoma from smokers compared to nonsmokers.

Journal of Clinical Oncology ◽

10.1200/jco.2015.33.3_suppl.305 ◽

2015 ◽

Vol 33 (3_suppl) ◽

pp. 305-305

Author(s):

Lorraine A. Chantrill ◽

Andreia Pinho ◽

Jiamin Wu ◽

Marc Giry-Latierre ◽

Amanda Mawson ◽

...

Keyword(s):

Pancreatic Cancer ◽

Risk Factor ◽

Protein Expression ◽

Axon Guidance ◽

Pancreatic Ductal Adenocarcinoma ◽

Interaction Analysis ◽

Ductal Adenocarcinoma ◽

Ductal Cells ◽

Patient Consent ◽

Never Smokers

305 Background: The most important modifiable risk factor for the development of pancreas cancer is smoking, which accounts for up to 25% of pancreatic ductal adenocarcinomas (PDAC; Maisonneuve P, Lowenfels AB: Epidemiology of pancreatic cancer: an update. Digestive Diseases 28:645-656, 2010), and the incidence of PDAC correlates with smoking prevalence (Weiss W, Benarde MA: The temporal relation between cigarette smoking and pancreatic cancer. American journal of public health 73:1403-1404, 1983). A recently published Ingenuity Pathway Analysis of GWAS genotype and risk factor data from the Pancreatic Cancer Case Control Consortium demonstrated that axon guidance signalling genes were significantly overrepresented in smokers (Tang H, Wei P, Duell EJ, et al: Axonal guidance signaling pathway interacting with smoking in modifying the risk of pancreatic cancer: A gene and pathway-based interaction analysis of GWAS data. Carcinogenesis:bgu010, 2014) and we aimed to investigate this link in our cohort. Methods: Tissue from resected PDAC were obtained from 200 patients via the Australian Pancreatic Cancer Genome Initiative with patient consent and ethical approval. Immunohistochemistry was performed on TMAs with the anti-Robo1 antibody (ab7279). The sections were scored on a 4-point scale of 0, 1, 2 and 3 intensity. All sections were scored blindly by two independent reviewers. Results: Robo1 protein expression is found in the normal pancreas, predominantly in acinar cells. In PDAC, Robo1 expression is predominantly in epithelial ductal cells. Most PDACs have Robo1 expression with an overall mean score of 1.65±0.05. 20 out of 200 patients were current smokers at the time of their pancreatectomy, 97 were never smokers, the remainder were ex-smokers. The mean score for smokers was 2.1± 0.1 and for never smokers 1.6±0.1 (p = 0.0003). Genomic analysis did not demonstrate any mutations in the Robo1 gene. There were 7 cases of loss of heterozygosity for Robo1; none of these were current smokers and they had an average score of 1.25±0.17. Conclusions: In addition to our genomic (Biankin AV, Waddell N, Kassahn KS, et al: Pancreatic cancer genomes reveal aberrations in axon guidance pathway genes. Nature 491:399-405, 2012) and methylation data (Nones K, Waddell N, Song S, et al: Genome‐wide DNA methylation patterns in pancreatic ductal adenocarcinoma reveal epigenetic deregulation of SLIT‐ROBO, ITGA2 and MET signaling. International Journal of Cancer, 2014), we provide the results of protein expression of Robo1 in a clinically annotated cohort of 200 cases of PDAC. We demonstrate that patients who are currently smoking have enhanced Robo1 expression. Preliminary results indicate that this may confer a poorer prognosis when coupled with high SLIT2 expression.

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Expression of GRP78 protein in pancreatic ductal adenocarcinoma and intraductal papillary mucinous neoplasm.

Journal of Clinical Oncology ◽

10.1200/jco.2017.35.4_suppl.323 ◽

2017 ◽

Vol 35 (4_suppl) ◽

pp. 323-323

Author(s):

Asha R. Dhanarajan ◽

Reginald Hill ◽

Sam G. Pappas ◽

Gerard J. Abood ◽

Jennifer Lynn Gnerlich ◽

...

Keyword(s):

Pancreatic Ductal Adenocarcinoma ◽

Intraductal Papillary Mucinous Neoplasm ◽

Protein Quality ◽

Expression Level ◽

Ductal Adenocarcinoma ◽

Pancreatic Ducts ◽

Low Grade ◽

Ductal Cells ◽

Mucinous Neoplasm ◽

Significant Difference

323 Background: Pancreatic ductal adenocarcinoma (PDAC) has a poor prognosis due to late detection, aggressive biology and resistance to chemotherapy. Markers for early detection of PDAC and mediators of chemoresistance are poorly understood. GRP78 is a major endoplasmic reticulum (ER) chaperone protein critical for protein quality control of the ER. Increased GRP78 expression promotes in vitro cancer cell survival, progression, and chemoresistance. This study aims to examine expression of GRP78 protein in PDAC and intraductal papillary mucinous neoplasm (IPMN) compared to normal pancreatic ducts using immunohistochemistry (IHC). Methods: The expression of GRP78 was assessed using IHC in 93 sections of formalin-fixed paraffin-embedded tissue: normal ducts (38), low grade IPMN (IPMN-LG, 26), high grade IPMN (IPMN-HG, 9), and PDAC (20). Immunostaining intensity of GRP78 protein was categorized as no or weak staining (0-1+) and strong staining (2-3+). Fisher’s exact test with two tails was performed using the GraphPad statistical software. Results: GRP78 expression was identified in the cytoplasm of normal pancreatic ducts, IPMN and PDAC with a fine granular pattern. All PDAC and IPMN-HG showed strong expression of GRP78 while the normal ductal cells showed only minimal expression of GRP78. 73% of IPMN-LG expresses GRP78 strongly. Statistical analysis revealed a significant difference in GRP78 expression level between normal ductal cells and all three pathological conditions including PDCA, IPMN-HG and IPMN-LG (all p < 0.0001). Conclusions: This study shows that the expression level of GRP78 is significantly increased in PDAC, IPMN-HG and IPMN-LG compared to normal ductal cells. There appears to be a progressive increase in GRP78 expression from IPMN-LG to IPMN-HG and invasive PDAC. Increased GRP78 expression may be a marker for those at risk of developing PDAC. Changes in GRP78 expression over time and its role in PDAC chemoresistance should be further assessed. [Table: see text]

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MicroRNA-21 Is Induced Early in Pancreatic Ductal Adenocarcinoma Precursor Lesions

Clinical Chemistry ◽

10.1373/clinchem.2009.137364 ◽

2010 ◽

Vol 56 (4) ◽

pp. 603-612 ◽

Cited By ~ 142

Author(s):

Maël Chalret du Rieu ◽

Jérôme Torrisani ◽

Janick Selves ◽

Talal Al Saati ◽

Anny Souque ◽

...

Keyword(s):

Early Detection ◽

Mouse Model ◽

Pancreatic Ductal Adenocarcinoma ◽

Early Stage ◽

Survival Rates ◽

Ductal Adenocarcinoma ◽

Early Event ◽

Precursor Lesions ◽

Tissue Samples ◽

Ductal Cells

Abstract Background: Pancreatic ductal adenocarcinoma (PDAC) has the poorest overall prognosis among gastrointestinal cancers; however, curative resection in early-stage PDAC greatly improves survival rates, indicating the importance of early detection. Because abnormal microRNA production is commonly detected in cancer, we investigated noninvasive precursor pancreatic intraepithelial neoplasia (PanIN) lesions for microRNA production as a potential early biomarker of PDAC. Methods: Pathologists identified and classified ductal lesions. We extracted total RNA from laser-capture microdissected PanIN tissue samples from a conditional KRAS(G12D) mouse model (n = 29) or of human origin (n = 38) (KRAS is v-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog). MicroRNA production was quantified by quantitative real-time PCR. Internal controls included 5S and U6 RNAs. Results: Production of microRNAs miR-21, miR-205, and miR-200 paralleled PanIN progression in the KRAS(G12D) mouse model, compared with microRNA production in samples of nonpathologic ducts. miR-21 demonstrated the highest relative concentrations in the precursor lesions. Interestingly, miR-205 and miR-21 up-regulation preceded phenotypic changes in the ducts. The production of microRNAs miR-21, miR-221, miR-222, and let-7a increased with human PanIN grade, with peak production occurring in hyperplastic PanIN-2/3 lesions. In situ hybridization analysis indicated miR-21 production to be concentrated in pathologic ductal cells. miR-21 production was regulated by KRAS(G12D) and epidermal growth factor receptor in PDAC-derived cell lines. Conclusions: Aberrant microRNA production is an early event in the development of PanIN. Our findings indicate that miR-21 warrants further investigation as a marker for early detection of PDAC.

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CCAAT/Enhancer-Binding Protein Delta (C/EBPδ): A Previously Unrecognized Tumor Suppressor that Limits the Oncogenic Potential of Pancreatic Ductal Adenocarcinoma Cells

Cancers ◽

10.3390/cancers12092546 ◽

2020 ◽

Vol 12 (9) ◽

pp. 2546

Author(s):

Leonie Hartl ◽

JanWillem Duitman ◽

Hella L. Aberson ◽

Kan Chen ◽

Frederike Dijk ◽

...

Keyword(s):

Pancreatic Cancer ◽

Tumor Suppressor ◽

Pancreatic Ductal Adenocarcinoma ◽

Binding Protein ◽

Ductal Adenocarcinoma ◽

Ductal Cells ◽

Enhancer Binding Protein ◽

Adenocarcinoma Cells ◽

Ccaat Enhancer Binding Protein

CCAAT/enhancer-binding protein δ (C/EBPδ) is a transcription factor involved in growth arrest and differentiation, which has consequently been suggested to harbor tumor suppressive activities. However, C/EBPδ over-expression correlates with poor prognosis in glioblastoma and promotes genomic instability in cervical cancer, hinting at an oncogenic role of C/EBPδ in these contexts. Here, we explore the role of C/EBPδ in pancreatic cancer. We determined C/EBPδ expression in biopsies from pancreatic cancer patients using public gene-expression datasets and in-house tissue microarrays. We found that C/EBPδ is highly expressed in healthy pancreatic ductal cells but lost in pancreatic ductal adenocarcinoma. Furthermore, loss of C/EBPδ correlated with increased lymph node involvement and shorter overall survival in pancreatic ductal adenocarcinoma patients. In accordance with this, in vitro experiments showed reduced clonogenic capacity and proliferation of pancreatic ductal adenocarcinoma cells following C/EBPδ re-expression, concurrent with decreased sphere formation capacity in soft agar assays. We thus report a previously unrecognized but important tumor suppressor role of C/EBPδ in pancreatic ductal adenocarcinoma. This is of particular interest since only few tumor suppressors have been identified in the context of pancreatic cancer. Moreover, our findings suggest that restoration of C/EBPδ activity could hold therapeutic value in pancreatic ductal adenocarcinoma, although the latter claim needs to be substantiated in future studies.

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Cell Type of Pancreatic Ductal Adenocarcinoma Origin: Implications for Prognosis and Clinical Outcomes

Visceral Medicine ◽

10.1159/000520946 ◽

2021 ◽

pp. 1-6

Author(s):

Shilpa Patil ◽

Yan Dou ◽

Janel L. Kopp

Keyword(s):

Pancreatic Ductal Adenocarcinoma ◽

Mouse Models ◽

Pancreatic Injury ◽

Genetically Engineered ◽

Ductal Adenocarcinoma ◽

Cell Of Origin ◽

Cell Type ◽

Genetically Engineered Mouse Models ◽

Cellular Origin ◽

Ductal Cells

Background: Pancreatic ductal adenocarcinoma (PDAC) is a devastating disease that has no effective early detection method or treatment to date. Summary: The normal cell type that initiates PDAC, or its cellular origin, is still unknown. To investigate the contribution of distinct normal epithelial cell types to PDAC tumorigenesis, genetically engineered mouse models were used to show that both acinar and ductal cells are capable of giving rise to PDAC. These studies indicated that genetic mutations and pancreatic injury interact differently with each cellular origin to affect their predilection and process for forming PDAC. In this review, we summarize recent findings using various genetically engineered mouse models in the identification and characterization of the PDAC cell of origin. We also discuss potential implications for cellular origin on tumor development, PDAC transcriptional subtype, and disease prognosis of patients. Key Message: Although it is clear that both ductal and acinar cells have the potential to form PDAC, whether cellular origin can indeed influence patient prognosis and whether knowledge of cellular origin will aid in the diagnosis or treatment of patients in the future will need further study.

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Snail-Overexpression Induces Epithelial-mesenchymal Transition and Metabolic Reprogramming in Human Pancreatic Ductal Adenocarcinoma and Non-tumorigenic Ductal Cells

Journal of Clinical Medicine ◽

10.3390/jcm8060822 ◽

2019 ◽

Vol 8 (6) ◽

pp. 822 ◽

Cited By ~ 9

Author(s):

Menghan Liu ◽

Sarah E. Hancock ◽

Ghazal Sultani ◽

Brendan P. Wilkins ◽

Eileen Ding ◽

...

Keyword(s):

Pancreatic Ductal Adenocarcinoma ◽

Cell Lines ◽

Epithelial Mesenchymal Transition ◽

Lactate Production ◽

Metabolic Reprogramming ◽

Ductal Adenocarcinoma ◽

Pancreatic Cell ◽

Mesenchymal Transition ◽

Atp Production ◽

Ductal Cells

The zinc finger transcription factor Snail is a known effector of epithelial-to-mesenchymal transition (EMT), a process that underlies the enhanced invasiveness and chemoresistance of common to cancerous cells. Induction of Snail-driven EMT has also been shown to drive a range of pro-survival metabolic adaptations in different cancers. In the present study, we sought to determine the specific role that Snail has in driving EMT and adaptive metabolic programming in pancreatic ductal adenocarcinoma (PDAC) by overexpressing Snail in a PDAC cell line, Panc1, and in immortalized, non-tumorigenic human pancreatic ductal epithelial (HPDE) cells. Snail overexpression was able to induce EMT in both pancreatic cell lines through suppression of epithelial markers and upregulation of mesenchymal markers alongside changes in cell morphology and enhanced migratory capacity. Snail-overexpressed pancreatic cells additionally displayed increased glucose uptake and lactate production with concomitant reduction in oxidative metabolism measurements. Snail overexpression reduced maximal respiration in both Panc1 and HPDE cells, with further reductions seen in ATP production, spare respiratory capacity and non-mitochondrial respiration in Snail overexpressing Panc1 cells. Accordingly, lower expression of mitochondrial electron transport chain proteins was observed with Snail overexpression, particularly within Panc1 cells. Modelling of 13C metabolite flux within both cell lines revealed decreased carbon flux from glucose in the TCA cycle in snai1-overexpressing Panc1 cells only. This work further highlights the role that Snail plays in EMT and demonstrates its specific effects on metabolic reprogramming of glucose metabolism in PDAC.

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Shared extracellular vesicle miRNA profiles of matched ductal pancreatic adenocarcinoma organoids and blood plasma samples show the power of organoid technology

Cellular and Molecular Life Sciences ◽

10.1007/s00018-020-03703-8 ◽

2020 ◽

Author(s):

Anikó Zeöld ◽

Gyöngyvér Orsolya Sándor ◽

Anna Kiss ◽

András Áron Soós ◽

Tamás Tölgyes ◽

...

Keyword(s):

Blood Plasma ◽

Cellular Heterogeneity ◽

Ductal Adenocarcinoma ◽

Driver Mutations ◽

Characteristic Change ◽

Plasma Samples ◽

Pdac Patient ◽

Ductal Cells ◽

Pancreatic Ductal Cells ◽

Ductal Pancreatic Adenocarcinoma

AbstractExtracellular vesicles (EV) are considered as a promising diagnostic tool for pancreatic ductal adenocarcinoma (PDAC), a disease with a poor 5-year survival that has not improved in the past years. PDAC patient-derived 3D organoids maintain the intratumoral cellular heterogeneity, characteristic for the tumor in vivo.Thus, they represent an ideal in vitro model system to study human cancers. Here we show that the miRNA cargo of EVs from PDAC organoids largely differs among patients. However, we detected a common set of EV miRNAs that were present in matched organoids and blood plasma samples of individual patients. Importantly, the levels of EV miR-21 and miR-195 were higher in PDAC blood EV preparations than in healthy controls, albeit we found no difference compared to chronic pancreatitis (CP) samples. In addition, here we report that the accumulation of collagen I, a characteristic change in the extracellular matrix (ECM) in both CP and PDAC, largely increases EV release from pancreatic ductal organoids. This provides a possible explanation why both CP and PDAC patient-derived plasma samples have an elevated amount of CD63 + EVs. Collectively, we show that PDAC patient-derived organoids represent a highly relevant model to analyze the cargo of tumor cell-derived EVs. Furthermore, we provide evidence that not only driver mutations, but also changes in the ECM may critically modify EV release from pancreatic ductal cells.

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Cell of origin affects tumour development and phenotype in pancreatic ductal adenocarcinoma

Gut ◽

10.1136/gutjnl-2017-314426 ◽

2018 ◽

Vol 68 (3) ◽

pp. 487-498 ◽

Cited By ~ 19

Author(s):

Alex Y L Lee ◽

Claire L Dubois ◽

Karnjit Sarai ◽

Soheila Zarei ◽

David F Schaeffer ◽

...

Keyword(s):

Pancreatic Ductal Adenocarcinoma ◽

Acinar Cells ◽

Ductal Adenocarcinoma ◽

Low Grade ◽

High Grade ◽

Cell Of Origin ◽

Ductal Cells ◽

Tumour Development ◽

Cellular Context ◽

The Impact

ObjectivePancreatic ductal adenocarcinoma (PDAC) is a highly aggressive tumour thought to arise from ductal cells via pancreatic intraepithelial neoplasia (PanIN) precursor lesions. Modelling of different genetic events in mice suggests both ductal and acinar cells can give rise to PDAC. However, the impact of cellular context alone on tumour development and phenotype is unknown.DesignWe examined the contribution of cellular origin to PDAC development by inducing PDAC-associated mutations, KrasG12D expression and Trp53 loss, specifically in ductal cells (Sox9CreER;KrasLSL-G12D;Trp53flox/flox (‘Duct:KPcKO’)) or acinar cells (Ptf1aCreER;KrasLSL-G12D;Trp53flox/flox (‘Acinar:KPcKO’)) in mice. We then performed a thorough analysis of the resulting histopathological changes.ResultsBoth mouse models developed PDAC, but Duct:KPcKO mice developed PDAC earlier than Acinar:KPcKO mice. Tumour development was more rapid and associated with high-grade murine PanIN (mPanIN) lesions in Duct:KPcKO mice. In contrast, Acinar:KPcKO mice exhibited widespread metaplasia and low-grade as well as high-grade mPanINs with delayed progression to PDAC. Acinar-cell-derived tumours also had a higher prevalence of mucinous glandular features reminiscent of early mPanIN lesions.ConclusionThese findings indicate that ductal cells are primed to form carcinoma in situ that become invasive PDAC in the presence of oncogenic Kras and Trp53 deletion, while acinar cells with the same mutations appear to require a prolonged period of transition or reprogramming to initiate PDAC. Our findings illustrate that PDAC can develop in multiple ways and the cellular context in which mutations are acquired has significant impact on precursor lesion initiation, disease progression and tumour phenotype.

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