ABSTRACT
Pancreatic ductal adenocarcinoma (PDA) tumor cells are deprived of oxygen and nutrients and therefore must adapt their metabolism to ensure proliferation. In some physiological states, cells rely on ketone bodies to satisfy their metabolic needs, especially during nutrient stress. Here, we show that PDA cells can activate ketone body metabolism and that ß-hydroxybutyrate (ßOHB) is an alternative cell-intrinsic or systemic fuel that can promote PDA growth and progression. PDA cells activate enzymes required for ketogenesis, utilizing various nutrients as carbon sources for ketone body formation. By assessing metabolic gene expression from spontaneously arising PDA tumors in mice, we find HMG-CoA lyase (HMGCL), involved in ketogenesis, to be among the most deregulated metabolic enzymes in PDA compared to normal pancreas. In vitro depletion of HMGCL impedes migration, tumor cell invasiveness, and anchorage-independent tumor sphere compaction. Moreover, disrupting HMGCL drastically decreases PDA tumor growth in vivo, while ßOHB stimulates metastatic dissemination to the liver. These findings suggest that ßOHB increases PDA aggressiveness and identify HMGCL and ketogenesis as metabolic targets for limiting PDA progression.
Subject(s)
Ketone Bodies , Pancreatic Neoplasms , 3-Hydroxybutyric Acid/metabolism , Animals , Ketone Bodies/metabolism , Mice , Oxo-Acid-Lyases , Pancreas/metabolismABSTRACT
Non-alcoholic fatty liver (NAFL) and related syndromes affect one-third of the adult population in industrialized and developing countries. Lifestyle and caloric oversupply are the main causes of such array of disorders, but the molecular mechanisms underlying their etiology remain elusive. Nuclear Protein 1 (NUPR1) expression increases upon cell injury in all organs including liver. Recently, we reported NUPR1 actively participates in the activation of the Unfolded Protein Response (UPR). The UPR typically maintains protein homeostasis, but downstream mediators of the pathway regulate metabolic functions including lipid metabolism. As increases in UPR and NUPR1 in obesity and liver disease have been well documented, the goal of this study was to investigate the roles of NUPR1 in this context. To establish whether NUPR1 is involved in these liver conditions we used patient-derived liver biopsies and in vitro and in vivo NUPR1 loss of functions models. First, we analyzed NUPR1 expression in a cohort of morbidly obese patients (MOPs), with simple fatty liver (NAFL) or more severe steatohepatitis (NASH). Next, we explored the metabolic roles of NUPR1 in wild-type (Nupr1+/+ ) or Nupr1 knockout mice (Nupr1-/- ) fed with a high-fat diet (HFD) for 15 weeks. Immunohistochemical and mRNA analysis revealed NUPR1 expression is inversely correlated to hepatic steatosis progression. Mechanistically, we found NUPR1 participates in the activation of PPAR-α signaling via UPR. As PPAR-α signaling is controlled by UPR, collectively, these findings suggest a novel function for NUPR1 in protecting liver from metabolic distress by controlling lipid homeostasis, possibly through the UPR.
Subject(s)
Basic Helix-Loop-Helix Transcription Factors/physiology , Endoplasmic Reticulum Stress , Lipid Metabolism , Liver/metabolism , Neoplasm Proteins/physiology , Animals , Cell Line, Tumor , Diet, High-Fat , Homeostasis , Humans , Mice , Unfolded Protein ResponseABSTRACT
BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) is characterised by an extensive tissue invasion and an early formation of metastasis. Alterations in the expression of cadherins have been reported in PDAC. Yet, how these changes contribute to tumour progression is poorly understood. Here, we investigated the relationship between cadherins expression and PDAC development. METHODS: Cadherins expression was assessed by immunostaining in both human and murine tissue specimens. We have generated pancreatic cancer cell lines expressing both cadherin-1 and cadherin-3 or only one of these cadherins. Functional implications of such genetic alterations were analysed both in vitro and in vivo. RESULTS: Cadherin-3 is detected early at the plasma membrane during progression of pancreatic intraepithelial neoplasia 1 (PanIN-1) to PDAC. Despite tumoural cells turn on cadherin-3, a significant amount of cadherin-1 remains expressed at the cell surface during tumourigenesis. We found that cadherin-3 regulates tumour growth, while cadherin-1 drives type I collagen organisation in the tumour. In vitro assays showed that cadherins differentially participate to PDAC aggressiveness. Cadherin-3 regulates cell migration, whereas cadherin-1 takes part in the invadopodia activity. CONCLUSIONS: Our results show differential, but complementary, roles for cadherins during PDAC carcinogenesis and illustrate how their expression conditions the PDAC aggressiveness.
Subject(s)
Antigens, CD/metabolism , Cadherins/metabolism , Carcinoma, Pancreatic Ductal/metabolism , Pancreatic Neoplasms/metabolism , Animals , Antigens, CD/genetics , Cadherins/genetics , Carcinoma, Pancreatic Ductal/genetics , Cell Line, Tumor , Cell Membrane/metabolism , Cell Movement , Cell Proliferation , Collagen Type I/metabolism , Disease Progression , Humans , Mice , Neoplasm Transplantation , Pancreatic Neoplasms/geneticsABSTRACT
Endothelial-mesenchymal transition (EndMT) is an important source of cancer-associated fibroblasts (CAFs), which facilitates tumour progression. PDAC is characterised by abundant CAFs and tumour necrosis factor-α (TNF-α). Here, we show that TNF-α strongly induces human endothelial cells to undergo EndMT. Interestingly, TNF-α strongly downregulates the expression of the endothelial receptor TIE1, and reciprocally TIE1 overexpression partially prevents TNF-α-induced EndMT, suggesting that TNF-α acts, at least partially, through TIE1 regulation in this process. We also show that TNF-α-induced EndMT is reversible. Furthermore, TNF-α treatment of orthotopic mice resulted in an important increase in the stroma, including CAFs. Finally, secretome analysis identified TNFSF12, as a regulator that is also present in PDAC patients. With the aim of restoring normal angiogenesis and better access to drugs, our results support the development of therapies targeting CAFs or inducing the EndMT reversion process in PDAC.
Subject(s)
Cancer-Associated Fibroblasts/drug effects , Carcinoma, Pancreatic Ductal/pathology , Endothelial Cells/drug effects , Epithelial-Mesenchymal Transition/drug effects , Pancreatic Neoplasms/pathology , Tumor Necrosis Factor-alpha/pharmacology , Animals , Cancer-Associated Fibroblasts/metabolism , Cancer-Associated Fibroblasts/pathology , Carcinoma, Pancreatic Ductal/genetics , Carcinoma, Pancreatic Ductal/metabolism , Cells, Cultured , Cytokine TWEAK/genetics , Cytokine TWEAK/metabolism , Endothelial Cells/metabolism , Endothelial Cells/pathology , Humans , Male , Mice, Transgenic , Pancreatic Neoplasms/genetics , Pancreatic Neoplasms/metabolism , Receptor, TIE-1/genetics , Receptor, TIE-1/metabolism , Snail Family Transcription Factors/genetics , Snail Family Transcription Factors/metabolism , Zinc Finger E-box Binding Homeobox 2/genetics , Zinc Finger E-box Binding Homeobox 2/metabolismABSTRACT
BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) is characterized by an important heterogeneity, reflected by different clinical outcomes and chemoresistance. During carcinogenesis, tumor cells display aberrant glycosylated structures, synthetized by deregulated glycosyltransferases, supporting the tumor progression. In this study, we aimed to determine whether PDAC could be stratified through their glycosyltransferase expression profiles better than the current binary classification (basal-like and classical) in order to improve detection of patients with poor prognosis. METHODS: Bioinformatic analysis of 169 glycosyltransferase RNA sequencing data were performed for 74 patient-derived xenografts (PDX) of resected and unresectable tumors. The Australian cohort of International Cancer Genome Consortium and the microarray dataset from Puleo patient's cohort were used as independent validation datasets. FINDINGS: New PDAC stratification based on glycosyltransferase expression profile allowed to distinguish different groups of patients with distinct clinical outcome (p-value = 0.007). A combination of 19 glycosyltransferases differentially expressed in PDX defined a glyco-signature, whose prognostic value was validated on datasets including resected whole tumor tissues. The glyco-signature was able to discriminate three clusters of PDAC patients on the validation cohorts, two clusters displaying a short overall survival compared to one cluster having a better prognosis. Both poor prognostic clusters having different glyco-profiles in Puleo patient's cohort were correlated with stroma activated or desmoplastic subtypes corresponding to distinct microenvironment features (p-value < 0.0001). Besides, differential expression and enrichment analyses revealed deregulated functional pathways specific to different clusters. INTERPRETATION: This study identifies a glyco-signature relevant for a prognostic use, potentially applicable to resected and unresectable PDAC. Furthermore, it provides new potential therapeutic targets. FUNDING: This work was supported by INCa (Grants number 2018-078 and 2018-079), Fondation ARC (Grant number ARCPJA32020070002326), Cancéropôle PACA, DGOS (labelization SIRIC, Grant number 6038), Amidex Foundation and Ligue Nationale Contre le Cancer and by institutional fundings from INSERM and the Aix-Marseille Université.
Subject(s)
Adenocarcinoma/genetics , Glycosyltransferases/genetics , Pancreatic Neoplasms/genetics , Adenocarcinoma/metabolism , Adenocarcinoma/pathology , Glycosylation , Glycosyltransferases/metabolism , Humans , Pancreatic Neoplasms/metabolism , Pancreatic Neoplasms/pathology , Survival Analysis , Transcriptome , Tumor MicroenvironmentABSTRACT
Despite clinical advances in diagnosis and treatment, pancreatic ductal adenocarcinoma (PDAC) remains the third leading cause of cancer death, and is still associated with poor prognosis and dismal survival rates. Identifying novel PDAC-targeted tools to tackle these unmet clinical needs is thus an urgent requirement. Here we use a peptide conjugate that specifically targets PDAC through low-density lipoprotein receptor (LDLR). We demonstrate by using near-infrared fluorescence imaging the potential of this conjugate to specifically detect and discriminate primary PDAC from healthy organs including pancreas and from benign mass-forming chronic pancreatitis, as well as detect metastatic pancreatic cancer cells in healthy liver. This work paves the way towards clinical applications in which safe LDLR-targeting peptide conjugate promotes tumor-specific delivery of imaging and/or therapeutic agents, thereby leading to substantial improvements of the PDAC patient's outcome.
Subject(s)
Carcinoma, Pancreatic Ductal/genetics , Pancreatic Neoplasms/genetics , Receptors, LDL/genetics , Carcinoma, Pancreatic Ductal/drug therapy , Carcinoma, Pancreatic Ductal/pathology , Cell Line, Tumor , Humans , Pancreatic Neoplasms/drug therapy , Pancreatic Neoplasms/pathology , Receptors, LDL/metabolismABSTRACT
BACKGROUND: A significant gap in pancreatic ductal adenocarcinoma (PDAC) patient's care is the lack of molecular parameters characterizing tumours and allowing a personalized treatment. METHODS: Patient-derived xenografts (PDX) were obtained from 76 consecutive PDAC and classified according to their histology into five groups. A PDAC molecular gradient (PAMG) was constructed from PDX transcriptomes recapitulating the five histological groups along a continuous gradient. The prognostic and predictive value for PMAG was evaluated in: i/ two independent series (n = 598) of resected tumours; ii/ 60 advanced tumours obtained by diagnostic EUS-guided biopsy needle flushing and iii/ on 28 biopsies from mFOLFIRINOX treated metastatic tumours. FINDINGS: A unique transcriptomic signature (PAGM) was generated with significant and independent prognostic value. PAMG significantly improves the characterization of PDAC heterogeneity compared to non-overlapping classifications as validated in 4 independent series of tumours (e.g. 308 consecutive resected PDAC, uHR=0.321 95% CI [0.207-0.5] and 60 locally-advanced or metastatic PDAC, uHR=0.308 95% CI [0.113-0.836]). The PAMG signature is also associated with progression under mFOLFIRINOX treatment (Pearson correlation to tumour response: -0.67, p-value < 0.001). INTERPRETATION: PAMG unify all PDAC pre-existing classifications inducing a shift in the actual paradigm of binary classifications towards a better characterization in a gradient. FUNDING: Project funding was provided by INCa (Grants number 2018-078 and 2018-079, BACAP BCB INCa_6294), Canceropole PACA, DGOS (labellisation SIRIC), Amidex Foundation, Fondation de France, INSERM and Ligue Contre le Cancer.
Subject(s)
Adenocarcinoma/diagnosis , Neoplasm Proteins/genetics , Pancreatic Neoplasms/diagnosis , Transcriptome/genetics , Adenocarcinoma/genetics , Adenocarcinoma/pathology , Adolescent , Adult , Aged , Animals , Antineoplastic Combined Chemotherapy Protocols/adverse effects , Antineoplastic Combined Chemotherapy Protocols/pharmacology , Biomarkers, Tumor/genetics , Cell Line, Tumor , Clinical Trials as Topic , Disease-Free Survival , Drug Resistance, Neoplasm/genetics , Endoscopic Ultrasound-Guided Fine Needle Aspiration , Female , Fluorouracil/adverse effects , Fluorouracil/pharmacology , Gene Expression Regulation, Neoplastic/drug effects , Heterografts , Humans , Irinotecan/adverse effects , Irinotecan/pharmacology , Leucovorin/adverse effects , Leucovorin/pharmacology , Male , Mice , Middle Aged , Neoplasm Metastasis , Oxaliplatin/adverse effects , Oxaliplatin/pharmacology , Pancreatic Neoplasms/genetics , Pancreatic Neoplasms/pathology , Precision Medicine , Prognosis , Young Adult , Pancreatic NeoplasmsABSTRACT
Tissue architecture contributes to pancreatic ductal adenocarcinoma (PDAC) phenotypes. Cancer cells within PDAC form gland-like structures embedded in a collagen-rich meshwork where nutrients and oxygen are scarce. Altered metabolism is needed for tumour cells to survive in this environment, but the metabolic modifications that allow PDAC cells to endure these conditions are incompletely understood. Here we demonstrate that collagen serves as a proline reservoir for PDAC cells to use as a nutrient source when other fuels are limited. We show PDAC cells are able to take up collagen fragments, which can promote PDAC cell survival under nutrient limited conditions, and that collagen-derived proline contributes to PDAC cell metabolism. Finally, we show that proline oxidase (PRODH1) is required for PDAC cell proliferation in vitro and in vivo. Collectively, our results indicate that PDAC extracellular matrix represents a nutrient reservoir for tumour cells highlighting the metabolic flexibility of this cancer.
Subject(s)
Carcinoma, Pancreatic Ductal/genetics , Collagen/metabolism , Gene Expression Regulation, Neoplastic , Pancreatic Neoplasms/genetics , Proline Oxidase/genetics , Proline/metabolism , Animals , Biological Transport , Carcinoma, Pancreatic Ductal/metabolism , Carcinoma, Pancreatic Ductal/pathology , Cell Line, Tumor , Cell Proliferation , Cell Survival , Collagen/chemistry , Extracellular Matrix/chemistry , Extracellular Matrix/metabolism , Humans , Male , Matrix Metalloproteinase 13/genetics , Matrix Metalloproteinase 13/metabolism , Matrix Metalloproteinase 9/genetics , Matrix Metalloproteinase 9/metabolism , Mice , Mice, Transgenic , Neoplasm Transplantation , Pancreatic Neoplasms/metabolism , Pancreatic Neoplasms/pathology , Procollagen-Proline Dioxygenase/genetics , Procollagen-Proline Dioxygenase/metabolism , Proline Oxidase/metabolism , Signal TransductionABSTRACT
c-MYC controls more than 15% of genes responsible for proliferation, differentiation, and cellular metabolism in pancreatic as well as other cancers making this transcription factor a prime target for treating patients. The transcriptome of 55 patient-derived xenografts show that 30% of them share an exacerbated expression profile of MYC transcriptional targets (MYC-high). This cohort is characterized by a high level of Ki67 staining, a lower differentiation state, and a shorter survival time compared to the MYC-low subgroup. To define classifier expression signature, we selected a group of 10 MYC target transcripts which expression is increased in the MYC-high group and six transcripts increased in the MYC-low group. We validated the ability of these markers panel to identify MYC-high patient-derived xenografts from both: discovery and validation cohorts as well as primary cell cultures from the same patients. We then showed that cells from MYC-high patients are more sensitive to JQ1 treatment compared to MYC-low cells, in monolayer, 3D cultured spheroids and in vivo xenografted tumors, due to cell cycle arrest followed by apoptosis. Therefore, these results provide new markers and potentially novel therapeutic modalities for distinct subgroups of pancreatic tumors and may find application to the future management of these patients within the setting of individualized medicine clinics.
Subject(s)
Antineoplastic Agents/metabolism , Azepines/metabolism , Gene Expression Profiling , Heterografts , Pancreatic Neoplasms/pathology , Triazoles/metabolism , Adult , Aged , Aged, 80 and over , Animals , Cell Survival/drug effects , Cells, Cultured , Female , Humans , Male , Mice , Middle Aged , Precision Medicine/methodsABSTRACT
Preclinical models based on patient-derived xenografts have remarkable specificity in distinguishing transformed human tumor cells from non-transformed murine stromal cells computationally. We obtained 29 pancreatic ductal adenocarcinoma (PDAC) xenografts from either resectable or non-resectable patients (surgery and endoscopic ultrasound-guided fine-needle aspirate, respectively). Extensive multiomic profiling revealed two subtypes with distinct clinical outcomes. These subtypes uncovered specific alterations in DNA methylation and transcription as well as in signaling pathways involved in tumor-stromal cross-talk. The analysis of these pathways indicates therapeutic opportunities for targeting both compartments and their interactions. In particular, we show that inhibiting NPC1L1 with Ezetimibe, a clinically available drug, might be an efficient approach for treating pancreatic cancers. These findings uncover the complex and diverse interplay between PDAC tumors and the stroma and demonstrate the pivotal role of xenografts for drug discovery and relevance to PDAC.
Subject(s)
Pancreatic Neoplasms/drug therapy , Animals , Carcinoma, Pancreatic Ductal , Cell Transformation, Neoplastic/drug effects , Datasets as Topic , Ezetimibe/pharmacology , Ezetimibe/therapeutic use , Humans , Male , Mice , Pancreatic Neoplasms/metabolism , Spheroids, Cellular/drug effects , Xenograft Model Antitumor Assays , Pancreatic NeoplasmsABSTRACT
The intratumoral microenvironment, or stroma, is of major importance in the pathobiology of pancreatic ductal adenocarcinoma (PDA), and specific conditions in the stroma may promote increased cancer aggressiveness. We hypothesized that this heterogeneous and evolving compartment drastically influences tumor cell abilities, which in turn influences PDA aggressiveness through crosstalk that is mediated by extracellular vesicles (EVs). Here, we have analyzed the PDA proteomic stromal signature and identified a contribution of the annexin A6/LDL receptor-related protein 1/thrombospondin 1 (ANXA6/LRP1/TSP1) complex in tumor cell crosstalk. Formation of the ANXA6/LRP1/TSP1 complex was restricted to cancer-associated fibroblasts (CAFs) and required physiopathologic culture conditions that improved tumor cell survival and migration. Increased PDA aggressiveness was dependent on tumor cell-mediated uptake of CAF-derived ANXA6+ EVs carrying the ANXA6/LRP1/TSP1 complex. Depletion of ANXA6 in CAFs impaired complex formation and subsequently impaired PDA and metastasis occurrence, while injection of CAF-derived ANXA6+ EVs enhanced tumorigenesis. We found that the presence of ANXA6+ EVs in serum was restricted to PDA patients and represents a potential biomarker for PDA grade. These findings suggest that CAF-tumor cell crosstalk supported by ANXA6+ EVs is predictive of PDA aggressiveness, highlighting a therapeutic target and potential biomarker for PDA.