Pancreatic ductal adenocarcinoma (PDAC) is composed of stromal, immune, and cancerous epithelial cells. Transcriptomic analysis of the epithelial compartment allows classification into different ...phenotypic subtypes as classical and basal‐like. However, little is known about the intra‐tumor heterogeneity particularly in the epithelial compartment. Growing evidences suggest that this phenotypic segregation is not so precise and different cancerous cell types may coexist in a single tumor. To test this hypothesis, we performed single‐cell transcriptomic analyses using combinational barcoding exclusively on epithelial cells from six different classical PDAC patients obtained by Endoscopic Ultrasound (EUS) with Fine Needle Aspiration (FNA). To purify the epithelial compartment, PDAC were grown as biopsy‐derived pancreatic cancer organoids. Single‐cell transcriptomic analysis allowed the identification of four main cell clusters present in different proportions in all tumors. Remarkably, although all these tumors were classified as classical, one cluster present in all corresponded to a basal‐like phenotype. These results reveal an unanticipated high heterogeneity of pancreatic cancers and demonstrate that basal‐like cells, which have a highly aggressive phenotype, are more widespread than expected.
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.
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.
The malignant progression of pancreatic ductal adenocarcinoma (PDAC) is accompanied by a profound desmoplasia, which forces proliferating tumor cells to metabolically adapt to this new ...microenvironment. We established the PDAC metabolic signature to highlight the main activated tumor metabolic pathways. Comparative transcriptomic analysis identified lipid-related metabolic pathways as being the most highly enriched in PDAC, compared with a normal pancreas. Our study revealed that lipoprotein metabolic processes, in particular cholesterol uptake, are drastically activated in the tumor. This process results in an increase in the amount of cholesterol and an overexpression of the low-density lipoprotein receptor (LDLR) in pancreatic tumor cells. These findings identify LDLR as a novel metabolic target to limit PDAC progression. Here, we demonstrate that shRNA silencing of LDLR, in pancreatic tumor cells, profoundly reduces uptake of cholesterol and alters its distribution, decreases tumor cell proliferation, and limits activation of ERK1/2 survival pathway. Moreover, blocking cholesterol uptake sensitizes cells to chemotherapeutic drugs and potentiates the effect of chemotherapy on PDAC regression. Clinically, high PDAC Ldlr expression is not restricted to a specific tumor stage but is correlated to a higher risk of disease recurrence. This study provides a precise overview of lipid metabolic pathways that are disturbed in PDAC. We also highlight the high dependence of pancreatic cancer cells upon cholesterol uptake, and identify LDLR as a promising metabolic target for combined therapy, to limit PDAC progression and disease patient relapse.
Significance Pancreatic ductal adenocarcinoma (PDAC) is projected to become the second deadliest cancer by 2030. Advances in therapeutic treatments are urgently required to fight against this fatal disease. Here, elucidation of the metabolic signature of PDAC has identified the low-density lipoprotein receptor (LDLR), which facilitates cholesterol uptake, as a promising therapeutic target. Blocking of LDLR reduces the proliferative and clonogenic potential of PDAC cells and decreases activation of the ERK1/2 survival pathway. Moreover, LDLR silencing sensitizes PDAC cells to chemotherapeutic drugs and potentiates the tumoral regression promoted by chemotherapy. Finally, Ldlr is highly expressed at all stages of human PDAC and expression is associated with an increased risk of PDAC recurrence.
Abstract
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.
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.
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•Patient xenografts are relevant models of pancreatic cancers and of their stroma•Xenografts allow the genomic analysis of unresectable pancreatic cancers•The epigenome, transcriptome, and stroma of pancreatic cancer define two subtypes•Pancreatic cancer is sensitive to inhibition of NPC1L1 by Ezetimibe
Nicolle et al. present a genomic analysis of pancreatic cancer xenografts showing that tumor subtypes are defined by specific epigenetic, transcriptional, and stromal landscapes. They reveal potential therapeutic targets through analysis of signaling cross-talk between tumor and stromal cells.
A significant gap in pancreatic ductal adenocarcinoma (PDAC) patient's care is the lack of molecular parameters characterizing tumours and allowing a personalized treatment.
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.
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).
PAMG unify all PDAC pre-existing classifications inducing a shift in the actual paradigm of binary classifications towards a better characterization in a gradient.
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.
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.
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.
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.
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.
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.
Pancreatic ductal adenocarcinoma (PDAC) is a heterogeneous disease, therefore stratification of patients is essential to predict their responses to therapies and to choose the best treatment. ...PDAC-derived organoids were produced from PDTX and Endoscopic Ultrasound-Guided Fine-Needle Aspiration (EUS-FNA) biopsies. A signature based on 16 genes targets of the c-MYC oncogene was applied to classify samples into two sub-groups with distinctive phenotypes named MYC-high and MYC-low. The analysis of 9 PDTXs and the corresponding derived organoids revealed that this signature which was previously designed from PDTX is transferable to the organoid model. Primary organoids from 24 PDAC patients were treated with NHWD-870 or JQ1, two inhibitors of c-MYC transcription. Notably, the comparison of their effect between the two sub-groups showed that both compounds are more efficient in MYC-high than in MYC-low samples, being NHWD-870 the more potent treatment. In conclusion, this study shows that the molecular signatures could be applied to organoids obtained directly from PDAC patients to predict the treatment response and could help to take the more appropriate therapeutic decision for each patient in a clinical timeframe.
Although significant advances have been made recently to characterize the biology of pancreatic ductal adenocarcinoma (PDAC), more efforts are needed to improve our understanding and to face ...challenges related to the aggressiveness, high mortality rate and chemoresistance of this disease.
In this study, we perform the metabolomics profiling of 77 PDAC patient-derived tumor xenografts (PDTX) to investigate the relationship of metabolic profiles with overall survival (OS) in PDAC patients, tumor phenotypes and resistance to five anticancer drugs (gemcitabine, oxaliplatin, docetaxel, SN-38 and 5-Fluorouracil).
We identified a metabolic signature that was able to predict the clinical outcome of PDAC patients (p < 0.001, HR=2.68 95% CI: 1.5–4.9). The correlation analysis showed that this metabolomic signature was significantly correlated with the PDAC molecular gradient (PAMG) (R = 0.44 and p < 0.001) indicating significant association to the transcriptomic phenotypes of tumors. Resistance score established, based on growth rate inhibition metrics using 35 PDTX-derived primary cells, allowed to identify several metabolites related to drug resistance which was globally accompanied by accumulation of several diacy-phospholipids and decrease in lysophospholipids. Interestingly, targeting glycerophospholipid synthesis improved sensitivity to the three tested cytotoxic drugs indicating that interfering with metabolism could be a promising therapeutic strategy to overcome the challenging resistance of PDAC.
In conclusion, this study shows that the metabolomic profile of pancreatic PDTX models is strongly associated to clinical outcome, transcriptomic phenotypes and drug resistance. We also showed that targeting the lipidomic profile could be used in combinatory therapies against chemoresistance in PDAC.
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