ABSTRACT
The diagnosis of pancreatic neuroendocrine tumours (PanNETs) is increasing owing to more sensitive detection methods, and this increase is creating challenges for clinical management. We performed whole-genome sequencing of 102 primary PanNETs and defined the genomic events that characterize their pathogenesis. Here we describe the mutational signatures they harbour, including a deficiency in G:C > T:A base excision repair due to inactivation of MUTYH, which encodes a DNA glycosylase. Clinically sporadic PanNETs contain a larger-than-expected proportion of germline mutations, including previously unreported mutations in the DNA repair genes MUTYH, CHEK2 and BRCA2. Together with mutations in MEN1 and VHL, these mutations occur in 17% of patients. Somatic mutations, including point mutations and gene fusions, were commonly found in genes involved in four main pathways: chromatin remodelling, DNA damage repair, activation of mTOR signalling (including previously undescribed EWSR1 gene fusions), and telomere maintenance. In addition, our gene expression analyses identified a subgroup of tumours associated with hypoxia and HIF signalling.
Subject(s)
Carcinoma, Neuroendocrine/genetics , Genome, Human/genetics , Genomics , Pancreatic Neoplasms/genetics , Base Sequence , Calmodulin-Binding Proteins/genetics , Chromatin Assembly and Disassembly/genetics , Chromosome Aberrations , DNA Copy Number Variations/genetics , DNA Glycosylases/genetics , DNA Mutational Analysis , DNA Repair/genetics , Female , Germ-Line Mutation/genetics , Humans , Male , RNA-Binding Protein EWS , RNA-Binding Proteins/genetics , TOR Serine-Threonine Kinases/metabolism , Telomere/genetics , Telomere/metabolismABSTRACT
This corrects the article DOI: 10.1038/nature21063.
ABSTRACT
Cystic neoplasms of the pancreas are an increasingly important public health problem. The majority of these lesions are benign but some progress to invasive pancreatic ductal adenocarcinoma (PDAC). There is a dearth of mouse models of these conditions. The orphan nuclear receptor NR5A2 regulates development, differentiation, and inflammation. Germline Nr5a2 heterozygosity sensitizes mice to the oncogenic effects of mutant Kras in the pancreas. Here, we show that - unlike constitutive Nr5a2+/- mice - conditional Nr5a2 heterozygosity in pancreatic epithelial cells, combined with mutant Kras (KPN+/- ), leads to a dramatic replacement of the pancreatic parenchyma with cystic structures and an accelerated development of high-grade PanINs and PDAC. Timed histopathological analyses indicated that in KPN+/- mice PanINs precede the formation of cystic lesions and the latter precede PDAC. A single episode of acute caerulein pancreatitis is sufficient to accelerate the development of cystic lesions in KPN+/- mice. Epithelial cells of cystic lesions of KPN+/- mice express MUC1, MUC5AC, and MUC6, but lack expression of MUC2, CDX2, and acinar markers, indicative of a pancreato-biliary/gastric phenotype. In accordance with this, in human samples we found a non-significantly decreased expression of NR5A2 in mucinous tumours, compared with conventional PDAC. These results highlight that the effects of loss of one Nr5a2 allele are time- and cell context-dependent. KPN+/- mice represent a new model to study the formation of cystic pancreatic lesions and their relationship with PanINs and classical PDAC. Our findings suggest that pancreatitis could also contribute to acceleration of cystic tumour progression in patients. © 2020 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
Subject(s)
Carcinoma, Pancreatic Ductal/pathology , Pancreatic Neoplasms/pathology , Receptors, Cytoplasmic and Nuclear/metabolism , Adult , Aged , Aged, 80 and over , Animals , Carcinoma, Pancreatic Ductal/genetics , Disease Progression , Epithelial Cells/pathology , Female , Heterozygote , Humans , Male , Mice , Middle Aged , Pancreatic Cyst/pathology , Pancreatic Ducts/pathology , Pancreatic Neoplasms/genetics , Proto-Oncogene Proteins p21(ras)/genetics , Proto-Oncogene Proteins p21(ras)/metabolism , Receptors, Cytoplasmic and Nuclear/geneticsABSTRACT
Early onset pancreatic cancer (EOPC) is a rare disease with a very high mortality rate. Almost nothing is known on the genetic susceptibility of EOPC, therefore, we performed a genome-wide association study (GWAS) to identify novel genetic variants specific for patients diagnosed with pancreatic ductal adenocarcinoma (PDAC) at younger ages. In the first phase, conducted on 821 cases with age of onset ≤60 years, of whom 198 with age of onset ≤50, and 3227 controls from PanScan I-II, we observed four SNPs (rs7155613, rs2328991, rs4891017 and rs12610094) showing an association with EOPC risk (P < 1 × 10-4 ). We replicated these SNPs in the PANcreatic Disease ReseArch (PANDoRA) consortium and used additional in silico data from PanScan III and PanC4. Among these four variants rs2328991 was significant in an independent set of 855 cases with age of onset ≤60 years, of whom 265 with age of onset ≤50, and 4142 controls from the PANDoRA consortium while in the in silico data, we observed no statistically significant association. However, the resulting meta-analysis supported the association (P = 1.15 × 10-4 ). In conclusion, we propose a novel variant rs2328991 to be involved in EOPC risk. Even though it was not possible to find a mechanistic link between the variant and the function, the association is supported by a solid statistical significance obtained in the largest study on EOPC genetics present so far in the literature.
Subject(s)
Genetic Predisposition to Disease/genetics , Pancreatic Neoplasms/genetics , Carcinoma, Pancreatic Ductal/genetics , Carcinoma, Pancreatic Ductal/pathology , Case-Control Studies , Female , Genome-Wide Association Study/methods , Humans , Male , Middle Aged , Pancreas/pathology , Pancreatic Neoplasms/pathology , Polymorphism, Single Nucleotide/genetics , Risk Factors , Pancreatic NeoplasmsABSTRACT
Solid pseudopapillary neoplasms (SPN) of the pancreas are rare, low-grade malignant neoplasms that metastasise to the liver or peritoneum in 10-15% of cases. They almost invariably present somatic activating mutations of CTNNB1. No comprehensive molecular characterisation of metastatic disease has been conducted to date. We performed whole-exome sequencing and copy-number variation (CNV) analysis of 10 primary SPN and comparative sequencing of five matched primary/metastatic tumour specimens by high-coverage targeted sequencing of 409 genes. In addition to CTNNB1-activating mutations, we found inactivating mutations of epigenetic regulators (KDM6A, TET1, BAP1) associated with metastatic disease. Most of these alterations were shared between primary and metastatic lesions, suggesting that they occurred before dissemination. Differently from mutations, the majority of CNVs were not shared among lesions from the same patients and affected genes involved in metabolic and pro-proliferative pathways. Immunostaining of 27 SPNs showed that loss or reduction of KDM6A and BAP1 expression was significantly enriched in metastatic SPNs. Consistent with an increased transcriptional response to hypoxia in pancreatic adenocarcinomas bearing KDM6A inactivation, we showed that mutation or reduced KDM6A expression in SPNs is associated with increased expression of the HIF1α-regulated protein GLUT1 at both primary and metastatic sites. Our results suggest that BAP1 and KDM6A function is a barrier to the development of metastasis in a subset of SPNs, which might open novel avenues for the treatment of this disease. © 2018 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.
Subject(s)
Biomarkers, Tumor/genetics , Carcinoma, Papillary/genetics , Carcinoma, Papillary/secondary , DNA Copy Number Variations , Gene Dosage , Mutation , Pancreatic Neoplasms/genetics , Pancreatic Neoplasms/pathology , Adolescent , Adult , Biomarkers, Tumor/analysis , Carcinoma, Papillary/chemistry , Child , Epigenesis, Genetic , Female , Gene Expression Regulation, Neoplastic , Genetic Predisposition to Disease , Glucose Transporter Type 1/genetics , Histone Demethylases/genetics , Humans , Male , Middle Aged , Mixed Function Oxygenases/genetics , Pancreatic Neoplasms/chemistry , Phenotype , Proto-Oncogene Proteins/genetics , Tumor Suppressor Proteins/genetics , Ubiquitin Thiolesterase/genetics , Young Adult , beta Catenin/geneticsABSTRACT
The intraductal oncocytic papillary neoplasm (IOPN) of the pancreas has been recognized by WHO classification as a unique intraductal papillary mucinous neoplasm (IPMN) category. IOPN is composed of oxyphil cells, usually expressing MUC5AC, MUC6, and Hep Par-1, and harboring PRKACA/B fusion genes as their genetic hallmark. Although IOPNs are associated with an infiltrative adenocarcinoma in up to 30% of cases, the survival rate after surgical resection approaches 100%. This highlights the importance of the correct IOPN diagnosis, above all in cases with an associated invasive component. In this study, the immunohistochemical expression of CD117 was investigated in 111 IPMNs, including 17 oncocytic, 45 gastric, 20 pancreatico-biliary, and 29 intestinal IPMNs. We also tested the expression of MUC5AC, MUC6, and Hep Par-1 in the IOPN cohort. CD117 positivity was significantly more frequent in IOPNs compared to the other IPMN subtypes (p < 0.0001). Furthermore, within IOPN, a lower or absent CD117, MUC5AC, MUC6, and Hep Par-1 expression tended to be associated with the presence of an infiltrative component. Our findings shed light into the biology of these complex lesions, which are confirmed to be a distinctive IPMN subtype; notably, CD117 emerged as a potential, additional tool in the differential diagnosis of IPMNs.
Subject(s)
Adenocarcinoma, Mucinous/pathology , Biomarkers, Tumor/metabolism , Carcinoma, Papillary/pathology , Oxyphil Cells/pathology , Pancreatic Intraductal Neoplasms/pathology , Proto-Oncogene Proteins c-kit/metabolism , Adenocarcinoma, Mucinous/metabolism , Carcinoma, Papillary/metabolism , Cell Line, Tumor , Cohort Studies , Humans , Mucins/metabolism , Pancreatic Intraductal Neoplasms/metabolismABSTRACT
OBJECTIVE: First, to assess the impact of the number of examined lymph nodes (ELNs) on staging and survival after distal pancreatectomy (DP) for pancreatic adenocarcinoma (PDAC). Second, to identify the minimum number of ELNs (MNELNs) ensuring an accurate detection of nodal involvement. Third, to reappraise the role of lymph node (LN) parameters, including N-status and lymph node ratio (LNR). BACKGROUND: In contrast with pancreatoduodenectomy, information on LN staging and the MNELN required in DP is lacking. METHODS: Patients undergoing DP for PDAC at 2 academic hospitals from 2000 through 2013 were retrospectively analyzed. The eighth edition of the American Joint Committee on Cancer staging system was used. The MNELN was estimated using the binomial probability law. Survival analyses were performed separately for node-negative and node-positive patients using univariable and multivariable models. RESULTS: The study population consisted of 240 patients. The median number of ELN was 21, significantly lower in node-negative patients as compared with node-positive patients (18.5 vs 24.0; P = 0.001). The proportion of node-positive patients increased with increasing numbers of ELNs, whereas LNR showed an inverse trend. The estimated MNELN was 20. The number of ELN (≥ or <20) was an independent prognostic factor only in node-negative patients [odds ratio (OR) 3.23 for ELN <20), suggesting a stage migration effect. In node-positive patients, N2-class, but not LNR, was a significant predictor of survival at multivariable analysis (OR 1.68). CONCLUSION: The number of ELN affects nodal staging in body/tail PDAC. At least 20 LNs are required for correct staging. N-status is superior to LNR in predicting survival of node-positive patients.
Subject(s)
Adenocarcinoma/pathology , Adenocarcinoma/surgery , Lymph Node Excision , Pancreatectomy , Pancreatic Neoplasms/pathology , Pancreatic Neoplasms/surgery , Adenocarcinoma/mortality , Adult , Aged , Female , Humans , Male , Middle Aged , Neoplasm Staging , Pancreatic Neoplasms/mortality , Pancreaticoduodenectomy , Retrospective Studies , Survival AnalysisABSTRACT
BACKGROUND: Two recent studies based on multi-omics data analysis identified distinct subtypes of bile-duct cancers (BDC) with important implications in terms of disease classification and patients' treatment. METHODS: Patients with mutations in KRAS, NRAS, TP53, and ARID1A genes were classified in KRAS/TP53 group while patients with mutations in IDH1-2, BAP1, and PBRM1 were classified in IDH1-2/BAP1/PBRM1 group. The aim of this study was to define long-term outcomes among patients stratified by patterns of genes mutated. RESULTS: Among 105 patients who underwent surgical resection for BDCs, 71 (68%) patients were classified in two groups based on patterns of genes mutated. While in IDH1-2/BAP1/PBRM1 group there were 58%, 22%, and 10% of patients with intrahepatic-cholangiocarcinoma (ICC), perihilar-cholangiocarcinoma (PHCC), and gallbladder cancer (GBC), in KRAS/TP53 group there were 42%, 78%, and 90% of patients with ICC, PHCC, and GBC (p = 0.003), respectively. Patients in IDH1-2/BAP1/PBRM1 group had a 5-year OS of 40% compared with 13% for KRAS/TP53 group (p = 0.032). In a multivariable model adjusted for margins, lymph-node status, microvascular invasion, and tumor grade, patients in KRAS/TP53 group had a 2.1-fold increased risk of death compared with patients in IDH1-2/BAP1/PBRM1 group (p = 0.028). CONCLUSIONS: Genetic data were able to overcome the clinical based staging system in predicting patients' prognosis.
Subject(s)
Bile Duct Neoplasms/genetics , Bile Duct Neoplasms/mortality , Aged , Bile Duct Neoplasms/pathology , Bile Duct Neoplasms/surgery , Cholangiocarcinoma/genetics , Cholangiocarcinoma/mortality , Cholangiocarcinoma/pathology , Cholangiocarcinoma/surgery , DNA-Binding Proteins/genetics , Female , Follow-Up Studies , GTP Phosphohydrolases/genetics , Gallbladder Neoplasms/genetics , Gallbladder Neoplasms/mortality , Gallbladder Neoplasms/pathology , Gallbladder Neoplasms/surgery , Humans , Isocitrate Dehydrogenase/genetics , Klatskin Tumor/genetics , Klatskin Tumor/mortality , Klatskin Tumor/pathology , Klatskin Tumor/surgery , Lymphatic Metastasis , Male , Margins of Excision , Membrane Proteins/genetics , Middle Aged , Multivariate Analysis , Mutation , Neoplasm Invasiveness , Prognosis , Proto-Oncogene Proteins p21(ras)/genetics , Retrospective Studies , Transcription Factors/genetics , Tumor Suppressor Protein p53/genetics , Tumor Suppressor Proteins/genetics , Ubiquitin Thiolesterase/geneticsABSTRACT
OBJECTIVE: To identify molecular prognostic factors and potentially actionable mutations in ampulla of Vater cancer (AVC). BACKGROUND: The largely variable outcomes of AVCs make clinical decisions difficult regarding the need of postsurgical therapy, which is based on morphological and immunohistochemical classification that do not adequately consider the varying degrees of heterogeneity present in many AVCs. No approved targeted therapies for AVC exist, but some show promising results requiring better molecular characterization to identify potential responders. METHODS: We assessed 80 AVCs for the prognostic value of mutations of kirsten rat sarcoma (KRAS), neuroblastoma RAS (NRAS), B rapidly accelerated fibrosarcoma (BRAF), TP53, and 4 membrane erythroblastosis oncogene B (ERBB) receptor tyrosine kinases (EGFR-ERBB1, HER2-ERBB2, HER3-ERBB3, HER4-ERBB4) amenable to pharmacological inhibition. Moreover, we evaluated mutations in 16 key components of rat sarcoma (RAS), phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K), protein 53 (P53), transforming growth factor beta (TGF-ß), and wingless/integrated (WNT) pathways, recently associated to AVC by whole-exome sequencing. RESULTS: TP53 and KRAS were mutated in 41% and 35% of cases, respectively, and emerged as independent prognostic factors together with tumor stage and regardless of the histotype (TP53: P = 0.0006; KRAS: P = 0.0018; stage IIB: P = 0.0117; stage III-IV: P = 0.0020). ERBB, WNT and PI3K pathway genes were mutated in 37.5% of cases. CONCLUSIONS: KRAS and TP53 mutations are negative predictors of survival in AVCs, regardless of histotype. Potentially actionable mutations in ERBB, WNT, and PI3K signaling pathway genes are present in 37.5% of all cases. These might be amenable to target therapy using available drugs like Everolimus in PI3K-mutated cases or compounds under active screening against ERBB and WNT signaling.
Subject(s)
Ampulla of Vater , Common Bile Duct Neoplasms/genetics , Mutation , Phosphatidylinositol 3-Kinases/genetics , Receptor, ErbB-3/genetics , Tumor Suppressor Protein p53/genetics , Wnt Signaling Pathway/genetics , Biomarkers, Tumor/genetics , Biomarkers, Tumor/metabolism , Common Bile Duct Neoplasms/diagnosis , Common Bile Duct Neoplasms/metabolism , DNA Mutational Analysis , DNA, Neoplasm/genetics , Female , Humans , Male , Middle Aged , Phosphatidylinositol 3-Kinases/metabolism , Prognosis , Receptor, ErbB-3/metabolism , Tumor Suppressor Protein p53/metabolismABSTRACT
Next-generation sequencing (NGS) was applied to 148 lung neuroendocrine tumours (LNETs) comprising the four World Health Organization classification categories: 53 typical carcinoid (TCs), 35 atypical carcinoid (ACs), 27 large-cell neuroendocrine carcinomas, and 33 small-cell lung carcinomas. A discovery screen was conducted on 46 samples by the use of whole-exome sequencing and high-coverage targeted sequencing of 418 genes. Eighty-eight recurrently mutated genes from both the discovery screen and current literature were verified in the 46 cases of the discovery screen, and validated on additional 102 LNETs by targeted NGS; their prevalence was then evaluated on the whole series. Thirteen of these 88 genes were also evaluated for copy number alterations (CNAs). Carcinoids and carcinomas shared most of the altered genes but with different prevalence rates. When mutations and copy number changes were combined, MEN1 alterations were almost exclusive to carcinoids, whereas alterations of TP53 and RB1 cell cycle regulation genes and PI3K/AKT/mTOR pathway genes were significantly enriched in carcinomas. Conversely, mutations in chromatin-remodelling genes, including those encoding histone modifiers and members of SWI-SNF complexes, were found at similar rates in carcinoids (45.5%) and carcinomas (55.0%), suggesting a major role in LNET pathogenesis. One AC and one TC showed a hypermutated profile associated with a POLQ damaging mutation. There were fewer CNAs in carcinoids than in carcinomas; however ACs showed a hybrid pattern, whereby gains of TERT, SDHA, RICTOR, PIK3CA, MYCL and SRC were found at rates similar to those in carcinomas, whereas the MEN1 loss rate mirrored that of TCs. Multivariate survival analysis revealed RB1 mutation (p = 0.0005) and TERT copy gain (p = 0.016) as independent predictors of poorer prognosis. MEN1 mutation was associated with poor prognosis in AC (p = 0.0045), whereas KMT2D mutation correlated with longer survival in SCLC (p = 0.0022). In conclusion, molecular profiling may complement histology for better diagnostic definition and prognostic stratification of LNETs. © 2016 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.
Subject(s)
Carcinoid Tumor/genetics , Carcinoma, Neuroendocrine/genetics , Chromatin Assembly and Disassembly/genetics , Lung Neoplasms/genetics , Neuroendocrine Tumors/genetics , Small Cell Lung Carcinoma/genetics , Adult , Aged , Carcinoid Tumor/pathology , Carcinoma, Neuroendocrine/pathology , Cohort Studies , DNA Copy Number Variations , DNA-Binding Proteins/genetics , Female , High-Throughput Nucleotide Sequencing , Humans , Italy , Lung Neoplasms/pathology , Male , Middle Aged , Mutation , Neoplasm Proteins/genetics , Neuroendocrine Tumors/pathology , Prognosis , Proto-Oncogene Proteins/genetics , Retinoblastoma Binding Proteins/genetics , Sequence Analysis, DNA , Small Cell Lung Carcinoma/pathology , Telomerase/genetics , Ubiquitin-Protein Ligases/geneticsABSTRACT
PURPOSE: SMAD4 mutational status correlates with pancreatic ductal adenocarcinoma (PDAC) failure pattern. We investigated in a subset of locally advanced patients submitted to radiofrequency ablation (RFA) whether the assessment of SMAD4 status is a useful way to select the patients. METHODS: Clinical, radiological, and follow-up details of patients submitted to RFA for locally advanced pancreatic cancer (LAPC), in whom cytohistological material was available at our institution, were retrospectively retrieved. SMAD4 expression was evaluated by immunohistochemistry (IHC) and considered "negative" or "positive." The survival analysis was conducted using Kaplan-Meier and Cox proportional hazards models. RESULTS: The study population consisted of 30 patients. Thirteen patients (43.3%) received RFA upfront, whereas 17 (56.7%) after induction treatments. SMAD4 was mutant in 18 out of 30 patients (60%). The overall estimated post-RFA disease-specific survival (DSS) was 15 months (95% CI 11.64-18.35). The estimated post-RFA DSS of patients with wild-type and mutant SMAD4 was 22 and 12 months, respectively (log-rank p < 0.05). At the multivariate analysis, SMAD4 was the only independent predictor of survival (p = 0.05). The pattern of failure was not associated with SMAD4 status (p = 0.4). CONCLUSIONS: Within patients undergoing RFA for LAPC, SMAD4 analysis could segregate a subgroup of subjects with improved survival, who likely benefited from tumor ablation.
Subject(s)
Carcinoma, Pancreatic Ductal/genetics , Carcinoma, Pancreatic Ductal/surgery , Catheter Ablation/methods , Pancreatic Neoplasms/genetics , Pancreatic Neoplasms/surgery , Smad4 Protein/genetics , Adult , Aged , Aged, 80 and over , Analysis of Variance , Carcinoma, Pancreatic Ductal/mortality , Carcinoma, Pancreatic Ductal/pathology , Catheter Ablation/mortality , Cohort Studies , Disease-Free Survival , Female , Follow-Up Studies , Gene Expression Regulation, Neoplastic , Hospitals, University , Humans , Italy , Kaplan-Meier Estimate , Male , Middle Aged , Mutation , Neoplasm Invasiveness/pathology , Neoplasm Staging , Pancreatic Neoplasms/mortality , Pancreatic Neoplasms/pathology , Retrospective Studies , Risk Assessment , Survival Analysis , Treatment OutcomeABSTRACT
Intraductal neoplasms are important precursors to invasive pancreatic cancer and provide an opportunity to detect and treat pancreatic neoplasia before an invasive carcinoma develops. The diagnostic evaluation of these lesions is challenging, as diagnostic imaging and cytological sampling do not provide accurate information on lesion classification, the grade of dysplasia or the presence of invasion. Moreover, the molecular driver gene mutations of these precursor lesions have yet to be fully characterized. Fifty-two intraductal papillary neoplasms, including 48 intraductal papillary mucinous neoplasms (IPMNs) and four intraductal tubulopapillary neoplasms (ITPNs), were subjected to the mutation assessment in 51 cancer-associated genes, using ion torrent semiconductor-based next-generation sequencing. P16 and Smad4 immunohistochemistry was performed on 34 IPMNs and 17 IPMN-associated carcinomas. At least one somatic mutation was observed in 46/48 (96%) IPMNs; 29 (60%) had multiple gene alterations. GNAS and/or KRAS mutations were found in 44/48 (92%) of IPMNs. GNAS was mutated in 38/48 (79%) IPMNs, KRAS in 24/48 (50%) and these mutations coexisted in 18/48 (37.5%) of IPMNs. RNF43 was the third most commonly mutated gene and was always associated with GNAS and/or KRAS mutations, as were virtually all the low-frequency mutations found in other genes. Mutations in TP53 and BRAF genes (10% and 6%) were only observed in high-grade IPMNs. P16 was lost in 7/34 IPMNs and 9/17 IPMN-associated carcinomas; Smad4 was lost in 1/34 IPMNs and 5/17 IPMN-associated carcinomas. In contrast to IPMNs, only one of four ITPNs had detectable driver gene (GNAS and NRAS) mutations. Deep sequencing DNA from seven cyst fluid aspirates identified 10 of the 13 mutations detected in their associated IPMN. Using next-generation sequencing to detect cyst fluid mutations has the potential to improve the diagnostic and prognostic stratification of pancreatic cystic neoplasms.
Subject(s)
Biomarkers, Tumor/genetics , Carcinoma, Pancreatic Ductal/genetics , Carcinoma, Papillary/genetics , DNA Mutational Analysis/methods , High-Throughput Nucleotide Sequencing , Mutation , Neoplasms, Cystic, Mucinous, and Serous/genetics , Pancreatic Neoplasms/genetics , Adult , Aged , Aged, 80 and over , Biomarkers, Tumor/analysis , Carcinoma, Pancreatic Ductal/chemistry , Carcinoma, Pancreatic Ductal/pathology , Carcinoma, Papillary/chemistry , Carcinoma, Papillary/pathology , Female , Genetic Predisposition to Disease , Humans , Immunohistochemistry , Male , Middle Aged , Multiplex Polymerase Chain Reaction , Neoplasm Grading , Neoplasms, Cystic, Mucinous, and Serous/chemistry , Neoplasms, Cystic, Mucinous, and Serous/pathology , Pancreatic Neoplasms/chemistry , Pancreatic Neoplasms/pathology , Phenotype , Retrospective StudiesABSTRACT
BACKGROUND: Mixed adenoneuroendocrine carcinomas (MANECs) of the gastrointestinal tract are rare neoplasms characterized by coexisting exocrine and neuroendocrine neoplastic components. MANECs' histogenetic classification and molecular characterization remain unclear, significantly affecting the identification of innovative therapeutic options for these tumors. METHODS: The exocrine and neuroendocrine components of 6 gastrointestinal MANECs were microdissected and subjected to the simultaneous mutation assessment in selected regions of 54 cancer-associated genes using Ion Torrent semiconductor-based next-generation sequencing. Sanger sequencing and immunohistochemistry were used as validation of the mutational status. RESULTS: A total of 20 driver gene somatic mutations were observed among the 12 neoplastic components investigated. In 11 of 12 (91.7%) samples, at least one mutation was detected; 7 samples (58.3%) were found to have multiple mutations. TP53 gene mutations were the most frequent genetic alterations observed in the series, occurring in 11/12 samples (91.7%). Somatic mutations in other genes were detected at lower frequencies: ATM, CTNNB1, ERBB4, JAK3, KDR, KRAS, RB1. CONCLUSIONS: Five of the 6 MANECs presented an overlapping mutational profile in both components, suggesting a monoclonal origin of the two MANEC components.
Subject(s)
Carcinoma, Neuroendocrine/genetics , DNA Mutational Analysis , Gastrointestinal Neoplasms/genetics , Aged , Aged, 80 and over , Carcinoma, Neuroendocrine/pathology , Female , Gastrointestinal Neoplasms/pathology , Humans , Male , Middle Aged , Mutation , Tumor Suppressor Protein p53/geneticsSubject(s)
Endoscopic Ultrasound-Guided Fine Needle Aspiration , Pancreatic Cyst/diagnosis , Adult , Diagnosis, Differential , Equipment Design , Female , Humans , Middle Aged , Ovary/cytology , Pancreatic Cyst/diagnostic imaging , Pancreatic Cyst/pathology , Stromal Cells/pathology , Surgical InstrumentsABSTRACT
Gastroenteropancreatic neuroendocrine tumors (GEP-NETs) are rare and highly heterogeneous neoplasms whose incidence has markedly increased over the last decades. A grading system based on the tumor cells' proliferation index predicts high-risk for G3 NETs. However, low-to-intermediate grade (G1/G2) NETs have an unpredictable clinical course that varies from indolent to highly malignant. Cultures of human cancer cells enable to perform functional perturbation analyses that are instrumental to enhance our understanding of cancer biology. To date, no tractable and reliable long-term culture of G1/G2 NET has been reported to permit disease modeling and pharmacological screens. Here, we report of the first long-term culture of a G2 metastatic small intestinal NET that preserves the main genetic drivers of the tumor and retains expression patterns of the endocrine cell lineage. Replicating the tissue, this long-term culture showed a low proliferation index, and yet it could be propagated continuously without dramatic changes in the karyotype. The model was readily available for pharmacological screens using targeted agents and as expected, showed low tumorigenic capacity in vivo. Overall, this is the first long-term culture of NETs to faithfully recapitulate many aspects of the original neuroendocrine tumor.
Subject(s)
Neuroendocrine Tumors , Humans , Neuroendocrine Tumors/pathology , Prognosis , Neoplasm Grading , Ki-67 Antigen/metabolism , Receptor Protein-Tyrosine KinasesABSTRACT
Myeloid cells can restrain antitumor immunity by metabolic pathways, such as the degradation of l-arginine, whose concentrations are regulated by the arginase 1 (ARG1) enzyme. Results from preclinical studies indicate the important role of arginine metabolism in pancreatic ductal adenocarcinoma (PDAC) progression, suggesting a potential for clinical application; however, divergent evolution in ARG1 expression and function in rodents and humans has restricted clinical translation. To overcome this dichotomy, here, we show that neutrophil extracellular traps (NETs), released by spontaneously activated neutrophils isolated from patients with PDAC, create a microdomain where cathepsin S (CTSS) cleaves human (h)ARG1 into different molecular forms endowed with enhanced enzymatic activity at physiological pH. NET-associated hARG1 suppresses T lymphocytes whose proliferation is restored by either adding a hARG1-specific monoclonal antibody (mAb) or preventing CTSS-mediated cleavage, whereas small-molecule inhibitors are not effective. We show that ARG1 blockade, combined with immune checkpoint inhibitors, can restore CD8+ T cell function in ex vivo PDAC tumors. Furthermore, anti-hARG1 mAbs increase the frequency of adoptively transferred tumor-specific CD8+ T cells in tumor and enhance the effectiveness of immune checkpoint therapy in humanized mice. Thus, this study shows that extracellular ARG1, released by activated myeloid cells, localizes in NETs, where it interacts with CTSS that in turn cleaves ARG1, producing major molecular forms endowed with different enzymatic activity at physiological pH. Once exocytosed, ARG1 activity can be targeted by mAbs, which bear potential for clinical application for the treatment of PDAC and require further exploration.
Subject(s)
Extracellular Traps , Pancreatic Neoplasms , Humans , Animals , Mice , CD8-Positive T-Lymphocytes , Extracellular Traps/metabolism , Arginase/metabolism , Immunotherapy , Pancreatic Neoplasms/therapy , Antibodies, Monoclonal/pharmacology , Tumor Microenvironment , Pancreatic NeoplasmsABSTRACT
Transcriptomic analyses of pancreatic ductal adenocarcinoma (PDAC) have identified two major epithelial subtypes with distinct biology and clinical behaviours. Here, we aimed to clarify the role of FGFR1 and FGFR4 in the definition of aggressive PDAC phenotypes. We found that the expression of FGFR4 is exclusively detected in epithelial cells, significantly elevated in the classical PDAC subtype, and associates with better outcomes. In highly aggressive basal-like/squamous PDAC, reduced FGFR4 expression aligns with hypermethylation of the gene and lower levels of histone marks associated with active transcription in its regulatory regions. Conversely, FGFR1 has more promiscuous expression in both normal and malignant pancreatic tissues and is strongly associated with the EMT phenotype but not with the basal-like cell lineage. Regardless of the genetic background, the increased proliferation of FGFR4-depleted PDAC cells correlates with hyperactivation of the mTORC1 pathway both in vitro and in vivo. Downregulation of FGFR4 in classical cell lines invariably leads to the enrichment of basal-like/squamous gene programs and is associated with either partial or full switch of phenotype. In sum, we show that endogenous levels of FGFR4 limit the malignant phenotype of PDAC cells. Finally, we propose FGFR4 as a valuable marker for the stratification of PDAC patients.
Subject(s)
Carcinoma, Pancreatic Ductal , Carcinoma, Squamous Cell , Pancreatic Neoplasms , Carcinoma, Pancreatic Ductal/pathology , Humans , Mechanistic Target of Rapamycin Complex 1/genetics , Pancreatic Neoplasms/pathology , Phenotype , Receptor, Fibroblast Growth Factor, Type 4/genetics , Pancreatic NeoplasmsABSTRACT
BACKGROUND: Extrahepatic cholangiocarcinoma (ECC) is classified into two subtypes based on anatomic origin: distal extrahepatic (DECC) and perihilar (PHCC) cholangiocarcinoma. This study aimed to shed light on its genomic and transcriptomic profiles. RESEARCH DESIGN AND METHODS: The genomic alterations of 99 ECC (47 PHCC and 52 DECC) were investigated by next-generation sequencing of 96 genes. A subgroup of cases, representative of each subtype, was further investigated using transcriptomic analysis. Bioinformatics tools were applied for clustering and pathway analysis and defining the immune composition of the tumor microenvironment. RESULTS: PHCC had more frequent KRAS mutations (p = 0.0047), whereas TP53 mutations were more common in DECC (p = 0.006). Potentially actionable alterations included high-tumor mutational burden and/or microsatellite instability (7.1%), PI3KCA mutations (8.1%), and MYC (10.1%) and ERBB2 amplification (5.1%). The transcriptomic profiles showed the presence of three distinct clusters, which followed the anatomic origin and differed in immune microenvironment. DECC appeared to contain two distinct tumor subgroups, one enriched for druggable alterations and one lacking actionable opportunities. CONCLUSIONS: This study provides new insights into the molecular landscape and the actionable alterations of ECC. Our findings represent a step toward improved ECC molecular taxonomy and therapeutic strategies for precision oncology.
Subject(s)
Bile Duct Neoplasms , Cholangiocarcinoma , Bile Duct Neoplasms/pathology , Bile Ducts, Intrahepatic/metabolism , Bile Ducts, Intrahepatic/pathology , Cholangiocarcinoma/pathology , Genomics , Humans , Mutation , Precision Medicine , Transcriptome , Tumor Microenvironment/geneticsABSTRACT
The GNA15 gene is ectopically expressed in human pancreatic ductal adenocarcinoma cancer cells. The encoded Gα15 protein can promiscuously redirect GPCR signaling toward pathways with oncogenic potential. We sought to describe the distribution of GNA15 in adenocarcinoma from human pancreatic specimens and to analyze the mechanism driving abnormal expression and the consequences on signaling and clinical follow-up. We detected GNA15 expression in pre-neoplastic pancreatic lesions and throughout progression. The analysis of biological data sets, primary and xenografted human tumor samples, and clinical follow-up shows that elevated expression is associated with poor prognosis for GNA15, but not any other GNA gene. Demethylation of the 5' GNA15 promoter region was associated with ectopic expression of Gα15 in pancreatic neoplastic cells, but not in adjacent dysplastic or non-transformed tissue. Down-modulation of Gα15 by shRNA or CRISPR/Cas9 affected oncogenic signaling, and reduced adenocarcimoma cell motility and invasiveness. We conclude that de novo expression of wild-type GNA15 characterizes transformed pancreatic cells. The methylation pattern of GNA15 changes in preneoplastic lesions coincident with the release a transcriptional blockade that allows ectopic expression to persist throughout PDAC progression. Elevated GNA15 mRNA correlates with poor prognosis. In addition, ectopic Gα15 signaling provides an unprecedented mechanism in the early steps of pancreas carcinogenesis distinct from classical G protein oncogenic mutations described previously in GNAS and GNAQ/GNA11.
Subject(s)
Carcinoma, Pancreatic Ductal/genetics , GTP-Binding Protein alpha Subunits, Gq-G11/genetics , Gene Expression Regulation, Neoplastic/genetics , Pancreatic Neoplasms/genetics , CRISPR-Cas Systems , Carcinoma, Pancreatic Ductal/pathology , Cell Line, Tumor , Cell Movement/genetics , GTP-Binding Proteins/metabolism , Gene Expression/genetics , Humans , Methylation , Neoplasm Invasiveness/genetics , Pancreatic Neoplasms/pathology , Prognosis , Promoter Regions, Genetic/genetics , RNA, Messenger , RNA, Small Interfering , Signal TransductionABSTRACT
Hepatoid tumors (HT) are rare neoplasms morphologically resembling hepatocellular carcinoma, which arise in several organs other than the liver. A comprehensive molecular profile of this group of neoplasms is still lacking. Genomic characterization of 19 HTs from different organs (three colon HTs, four esophagogastric HTs, four biliary HTs, six genitourinary HTs, two lung HTs) was performed using a multigene next-generation sequencing panel. NGS unraveled a composite molecular profile of HT. Their genetic alterations were clearly clustered by tumor site: (i) colorectal HT displayed microsatellite instability, high tumor mutational burden, mutations in ARID1A/B genes and NCOA4-RET gene fusion (2/3 cases); (ii) gastric HT had TP53 mutations (2/4); (iii) biliary HT displayed loss of CDKN2A (3/4) and loss of chromosome 18 (2/4); (iv) genital HT showed gain of chromosome 12 (3/6); (v) lung HT had STK11 somatic mutations (2/2). The only commonly mutated gene occurring in HT of different sites was TP53 (8/19 cases: colon 2, esophagogastric 2, biliary 2, genital 1, lungs 1). This study shows that most genetic alterations of HT were clustered by site, indicating that context matters. The novel potential targets for HT precision oncology are also clustered based on the anatomic origin. This study shed light on the biology of these rare cancers and may have important consequences for treatment decisions and clinical trial selection for HT patients.