Humoral Predictors of Malignancy in IPMN: A Review of the Literature.

Pancreatic cystic lesions are increasingly detected in cross-sectional imaging. Intraductal papillary mucinous neoplasm (IPMN) is a mucin-producing subtype of the pancreatic cyst lesions arising from the pancreatic duct system. IPMN is a potential precursor of pancreatic cancer. The transformation of IPMN in pancreatic cancer is progressive and requires the occurrence of low-grade dysplasia, high-grade dysplasia, and ultimately invasive cancer. Jaundice, enhancing mural nodule >5 mm, main pancreatic duct diameter >10 mm, and positive cytology for high-grade dysplasia are considered high-risk stigmata of malignancy. While increased levels of carbohydrate antigen 19-9 (CA 19-9) (>37 U/mL), main pancreatic duct diameter 5-9.9 mm, cyst diameter >40 mm, enhancing mural nodules <5 mm, IPMN-induced acute pancreatitis, new onset of diabetes, cyst grow-rate >5 mm/year are considered worrisome features of malignancy. However, cross-sectional imaging is often inadequate in the prediction of high-grade dysplasia and invasive cancer. Several studies evaluated the role of humoral and intra-cystic biomarkers in the prediction of malignancy in IPMN. Carcinoembryonic antigen (CEA), CA 19-9, intra-cystic CEA, intra-cystic glucose, and cystic fluid cytology are widely used in clinical practice to distinguish between mucinous and non-mucinous cysts and to predict the presence of invasive cancer. Other biomarkers such as cystic fluid DNA sequencing, microRNA (mi-RNA), circulating microvesicles, and liquid biopsy are the new options for the mini-invasive diagnosis of degenerated IPMN. The aim of this study is to review the literature to assess the role of humoral and intracystic biomarkers in the prediction of advanced IPMN with high-grade dysplasia or invasive carcinoma.

Bioinformatic Analysis and Integration of Transcriptome and Proteome Results Identify Key Coding and Noncoding Genes Predicting Malignancy in Intraductal Papillary Mucinous Neoplasms of the Pancreas.

BACKGROUND: Intraductal papillary mucinous neoplasms (IPMNs) are precursor lesions of pancreatic ductal adenocarcinoma (PDAC). IPMNs are generally associated with high risk of developing malignancy and therefore need to be diagnosed and assessed accurately, once detected. Existing diagnostic methods are inadequate, and identification of efficient biomarker capable of detecting high-risk IPMNs is necessitated. Moreover, the mechanism of development of malignancy in IPMNs is also elusive. METHODS: Gene expression meta-analysis conducted using 12 low-risk IPMN and 23 high-risk IPMN tissue samples. We have also listed all the altered miRNAs and long noncoding RNAs (lncRNAs), identified their target genes, and performed pathway analysis. We further enlisted cyst fluid proteins detected to be altered in high-risk or malignant IPMNs and compared them with fraction of differentially expressed genes secreted into cyst fluid. RESULTS: Our meta-analysis identified 270 upregulated and 161 downregulated genes characteristically altered in high-risk IPMNs. We further identified 61 miRNAs and 14 lncRNAs and their target genes and key pathways contributing towards understanding of the gene regulation during the progression of the disease. Most importantly, we have detected 12 genes altered significantly both in cystic lesions and cyst fluid. CONCLUSION: Our study reports, for the first time, a meta-analysis identifying key changes in gene expression between low-risk and high-risk IPMNs and also explains the regulatory aspect through construction of a miRNA-lncRNA-mRNA interaction network. The 12-gene-signature could function as potential biomarker in cyst fluid for detection of IPMN with a high risk of developing malignancy.

Diabetes in Humans Activates Pancreatic Stellate Cells via RAGE in Pancreatic Ductal Adenocarcinoma.

Pancreatic stellate cells (PSCs) mainly consist of cancer-associating fibroblasts in pancreatic ductal adenocarcinoma (PDAC). The receptor for advanced glycation end products (RAGE) is implicated in the pathophysiology of diabetic complications. Here, we studied the implication of RAGE in PSC activation in PDAC. The activation of cultured mouse PSCs was evaluated by qPCR. The induction of epithelial mesenchymal transition (EMT) in PDAC cell lines was assessed under stimulation with culture supernatant from activated PSCs. A total of 155 surgically resected PDAC subjects (83 nondiabetic, 18 with ≦3-years and 54 with >3-years history of diabetes) were clinicopathologically evaluated. A high-fat diet increased the expression of activated markers in cultured PSCs, which was abrogated by RAGE deletion. Culture supernatant from activated PSCs facilitated EMT of PDAC cells with elevation of TGF-ß and IL-6, but not from RAGE-deleted PSCs. Diabetic subjects complicated with metabolic syndrome, divided by cluster analysis, showed higher PSC activation and RAGE expression. In such groups, PDAC cells exhibited an EMT nature. The complication of metabolic syndrome with diabetes significantly worsened disease-free survival of PDAC subjects. Thus, RAGE in PSCs can be viewed as a new promoter and a future therapeutic target of PDAC in diabetic subjects with metabolic syndrome.

Simple mucinous cyst: another potential cancer precursor in the pancreas? Case report with molecular characterization and systematic review of the literature.

Cystic lesions of the pancreas may range from benign to precursors of pancreatic cancer. Simple mucinous cyst (SMC) is larger than 1 cm, has a gastric-type flat mucinous lining, and minimal atypia without ovarian-type stroma. We report a new case of pancreatic SMC, coupling a systematic review of the English literature mainly focused on their clinic-pathological features. We reviewed 103 cases of SMC in adults (73 women), averaging 57 (range, 26-70) years. The SMCs were located in the body-tail region of the pancreas in 60 (58%) cases, presenting as single cystic lesions in 94% of cases; 43% of patients were asymptomatic. A preoperative fine-needle aspiration of the cyst fluid detected amylase and carcinoembryonic antigen positivity in 71% and 76% of cases, respectively. Patients underwent surgery mostly for suspected malignancy; in 83% of cases, a standard pancreatic resection was performed. Mean SMC size was 4.9 (range, 1.5-12.0) cm. Mucins MUC5AC and MUC6 resulted positive in 77% and 81% of cases performed, respectively, whereas MUC2 was negative in all but one patient. The SMC from our institution was characterized by a KRAS somatic mutation. The diagnosis of SMC should be considered when a solitary pancreatic cyst larger than 1 cm is detected in asymptomatic patients. To establish a correct diagnosis, an extensive histologic/immunohistochemical analysis is essential. The presence of a KRAS mutation highlights that SMC may represent another potential pancreatic cancer precursor.

Intestinal phenotype is maintained by Atoh1 in the cancer region of intraductal papillary mucinous neoplasm.

Intraductal papillary mucinous neoplasm (IPMN) is a precancerous lesion of pancreatic cancer. Although there are 4 types of IPMN, among which intestinal-type IPMN is likely to progress into invasive cancer known as colloid carcinoma, no information regarding the involvement of the intestinal phenotype in the carcinogenesis of IPMN exists. The present study was conducted to explore how the intestinal differentiation system is maintained during the tumor progression of intestinal-type IPMN using surgical resection specimens. Results showed that Atoh1, a critical transcriptional factor for intestinal differentiation toward the secretory lineages of intestinal epithelial cells, was expressed in an invasive-grade IPMN. To determine the function of Atoh1 in pancreatic cancer, we generated a pancreatic ductal adenocarcinoma (PDAC) cell line overexpressing Atoh1. In a xenograft model, we successfully induced an IPMN phenotype in PDAC cells via Atoh1 induction. Finally, for the first time, we discovered that GPA33 is expressed in intestinal-type IPMN, thereby suggesting a novel target for cancer therapy. In conclusion, the intestinal differentiation system might be maintained during tumor progression of intestinal-type IPMN. Further analysis of the function of Atoh1 in IPMN might be useful for understanding the molecular mechanism underlying the malignant potential during the tumor progression of IPMN.

Possibility of detecting intraductal papillary mucinous neoplasms using metabolite biomarkers for pancreatic cancer.

Aim: The aim of this study was to identify whether metabolite biomarker candidates for pancreatic cancer (PC) could aid detection of intraductal papillary mucinous neoplasms (IPMN), recognized as high-risk factors for PC. Materials & methods: The 12 metabolite biomarker candidates, which were found to be useful to detect PC in our previous study, were evaluated for plasma samples from patients with PC (n = 44) or IPMN (n = 24) or healthy volunteers (n = 46). Results: Regarding the performance of individual biomarkers of PC and PC high-risk IPMN, lysine exhibited the best performance (sensitivity: 67.8%; specificity: 86.9%). The multiple logistic regression analysis-based detection model displayed high sensitivity and specificity values of 92.5 and 90.6%, respectively. Conclusion: Metabolite biomarker candidates for PC are useful for detecting high-risk IPMN, which can progress to PC.

CD117 Is a Specific Marker of Intraductal Papillary Mucinous Neoplasms (IPMN) of the Pancreas, Oncocytic Subtype.

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.

Differential diagnosis of pancreatic cysts: A prospective study on the role of intra-cystic glucose concentration.

BACKGROUND: The accuracy and costs of current diagnostic methods in the differential diagnosis of pancreatic cystic lesions still has ample room for improvement. AIMS: The aim of the study was to confirm the diagnostic yield of intracystic glucose in the diagnosis of pancreatic cyst subtypes. METHODS: We prospectively recruited all patients who underwent Endoscopic Ultrasound with Fine Needle Aspiration of pancreatic cyst at our Institution. RESULTS: Fifty-six patients were included in the study. We found that intracystic glucose concentration < 50 mg/dL was significantly more sensitive than a concentration of Carcinoembryonic Antigen > 192 ng/mL (93.6% vs 54.8%; p = 0.003) for the diagnosis of mucinous cysts. In terms of specificity, the two markers were not different (96% vs 100%; p = 1). Regarding the diagnosis of non-mucinous cysts, intracystic glucose concentration ≥ 50 mg/mL showed higher sensitivity than Carcinoembryonic Antigen level < 5 ng/mL (96% vs 72%) although a statistical significance could not be reached (p = 0.07). The two markers were not statistically different in terms of specificity (93.6% vs 87.1%; p = 0.62). CONCLUSION: Given its diagnostic performance and ease of measurement, intracystic glucose may replace Carcinoembryonic Antigen in the differential diagnosis of mucinous versus non-mucinous pancreatic cysts.

Molecular characterization of organoids derived from pancreatic intraductal papillary mucinous neoplasms.

Intraductal papillary mucinous neoplasms (IPMNs) are commonly identified non-invasive cyst-forming pancreatic neoplasms with the potential to progress into invasive pancreatic adenocarcinoma. There are few in vitro models with which to study the biology of IPMNs and their progression to invasive carcinoma. Therefore, we generated a living biobank of organoids from seven normal pancreatic ducts and ten IPMNs. We characterized eight IPMN organoid samples using whole genome sequencing and characterized five IPMN organoids and seven normal pancreatic duct organoids using transcriptome sequencing. We identified an average of 11,344 somatic mutations in the genomes of organoids derived from IPMNs, with one sample harboring 61,537 somatic mutations enriched for T→C transitions and T→A transversions. Recurrent coding somatic mutations were identified in 15 genes, including KRAS, GNAS, RNF43, PHF3, and RBM10. The most frequently mutated genes were KRAS, GNAS, and RNF43, with somatic mutations identified in six (75%), four (50%), and three (37.5%) IPMN organoid samples, respectively. On average, we identified 36 structural variants in IPMN derived organoids, and none had an unstable phenotype (> 200 structural variants). Transcriptome sequencing identified 28 genes differentially expressed between normal pancreatic duct organoid and IPMN organoid samples. The most significantly upregulated and downregulated genes were CLDN18 and FOXA1. Immunohistochemical analysis of FOXA1 expression in 112 IPMNs, 113 mucinous cystic neoplasms, and 145 pancreatic ductal adenocarcinomas demonstrated statistically significant loss of expression in low-grade IPMNs (p < 0.0016), mucinous cystic neoplasms (p < 0.0001), and pancreatic ductal adenocarcinoma of any histologic grade (p < 0.0001) compared to normal pancreatic ducts. These data indicate that FOXA1 loss of expression occurs early in pancreatic tumorigenesis. Our study highlights the utility of organoid culture to study the genetics and biology of normal pancreatic duct and IPMNs. © 2020 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Upregulation of follistatin and low apoptotic activity in intraductal oncocytic papillary neoplasm of the pancreatobiliary system.

Intraductal oncocytic papillary neoplasm (IOPN) is a rare intraductal tumor of the pancreatobiliary system. Currently, little is known about its distinct characteristics, unlike intraductal papillary mucinous neoplasms (IPMN) and intraductal papillary neoplasms of the bile duct (IPNB). The present study compared 22 IOPNs (18 pancreatic and 4 biliary) with those of 61 IPMNs/8 IPNBs. IOPNs were classified into pure and combined types, depending on the coexistence of IPMN/IPNB. Multiple gene expression analysis (nCounter system) was performed, and hierarchical clustering analysis separated IOPNs(n = 4) and IPMNs(n = 3)/ IPNBs(n = 3), and pathway score analysis supported the result. Volcano plot identified follistatin (FST) as the most upregulated mRNA in IOPN in comparison to the gastric subtype (log2 fold change of 5.34) and the intestinal subtype (that of 5.81) of IPMN/IPNB. The expression of FST in IOPN was also high in quantitative polymerase chain reaction and immunohistochemical analysis. We also found lower apoptotic activity in IOPN, particularly in pure type, compared to high-grade or invasive IPMN/IPNB using immunohistochemistry for cleaved caspase 3. But, combined type IOPN was more similar to IPMN/IPNB than pure IOPN. In conclusion, we proved that IOPN, particularly pure IOPN, is distinct from IPMN/IPNB in FST mRNA overexpression and exhibits lower apoptotic activity.

RNA cargos in extracellular vesicles derived from blood serum in pancreas associated conditions.

Exosomes are extracellular vesicles which are released from healthy and tumor cells into blood circulation. Unique biomolecular cargos such as RNA and protein are loaded in these vesicles. These molecules may have biological functions such as signaling, cell communications and have the potential to be analyzed as biomarkers. In this initial study, we describe the analysis of exosomes in the serum of healthy subjects, intraductal papillary mucosal neoplasms and pancreatic ductal adenocarcinoma including the characterization of their RNA cargos by next generation sequencing (EXO-NGS). Results indicate the presence of a wide variety of RNAs including mRNA, miRNA, lincRNA, tRNA and piRNA in these vesicles. Based on the differential mRNA expression observed upon EXO-NGS analysis, we independently evaluated two protein coding genes, matrix metalloproteinase-8 (MMP-8) and transcription factor T-Box 3 (TBX3) by qRT-PCR for selective expression in the serum samples. Results indicate a variable expression pattern of these genes across serum samples between different study groups. Further, qRT-PCR analysis with the same serum exosomes processed for EXO-NGS, we observed two long non-coding RNAs, malat-1 and CRNDE to be variably expressed. Overall, our observations emphasize the potential value of different exosome components in distinguishing between healthy, premalignant and malignant conditions related to the pancreas.

Excellent Accuracy of Glucose Level in Cystic Fluid for Diagnosis of Pancreatic Mucinous Cysts.

BACKGROUND: CEA in pancreatic cystic fluid (PCF) is standard for mucinous cysts diagnosis. Glucose is an alternative, but its accuracy remains poorly described. AIMS: To evaluate PCF glucose using a glucometer and compare its accuracy with CEA for mucinous cysts diagnosis. MATERIALS AND METHODS: In frozen PCF obtained by EUS-FNA, glucose was evaluated using a glucometer. CEA and cytology were available as standard of care. The accuracy of glucose and CEA was calculated using receiver operator (ROC) curves. Definitive diagnoses were surgical or clinicopathological. RESULTS: We evaluated 82 patients with a mean age of 61.3 ± 14.8 years (25-91), predominantly (59%) females. Diagnoses included 17 serous cystadenomas, five pseudocysts, 20 intraductal papillary mucinous neoplasms, three mucinous cystic neoplasms, five adenocarcinomas, four neuroendocrine tumors, two other types, 26 non-defined. The median glucose levels (interquartile range) were 19 mg/dL (19-19) in mucinous and 105 mg/dL (96-127) in non-mucinous cysts (p < 0.0001). The median CEA level was 741 ng/mL (165-28,567) in mucinous and 9 ng/mL (5-19) in non-mucinous cysts (p < 0.0001). For mucinous cyst diagnosis, a CEA > 192 ng/mL had a sensitivity of 72% (95% CI 51-88) and a specificity of 96% (95% CI 82-100), and ROC analysis showed an area under the curve (AUC) of 0.842 (95% CI 0.726-0.959), while glucose < 50 mg/dL had a sensitivity of 89% (95% CI 72-98), a specificity of 86% (95% CI 67-96), and an AUC of 0.86 (95% CI 0.748-0.973). Pseudocysts presented low glucose, identically to mucinous cysts, with CEA allowing differential diagnosis. CONCLUSION: Glucose measured by a glucometer is accurate for mucinous cyst diagnosis, with significantly higher levels in non-mucinous cysts, except pseudocysts.

The TLR7/8 agonist R848 remodels tumor and host responses to promote survival in pancreatic cancer.

A priority in cancer research is to innovate therapies that are not only effective against tumor progression but also address comorbidities such as cachexia that limit quality and quantity of life. We demonstrate that TLR7/8 agonist R848 induces anti-tumor responses and attenuates cachexia in murine models of pancreatic ductal adenocarcinoma (PDAC). In vivo, tumors from two of three cell lines were R848-sensitive, resulting in smaller tumor mass, increased immune complexity, increased CD8+ T-cell infiltration and activity, and decreased Treg frequency. R848-treated mice demonstrated improvements in behavioral and molecular cachexia manifestations, resulting in a near-doubling of survival duration. Knockout mouse studies revealed that stromal, not neoplastic, TLR7 is requisite for R848-mediated responses. In patient samples, we found Tlr7 is ubiquitously expressed in stroma across all stages of pancreatic neoplasia, but epithelial Tlr7 expression is relatively uncommon. These studies indicate immune-enhancing approaches including R848 may be useful in PDAC and cancer-associated cachexia.

Oncogenic KRAS Reduces Expression of FGF21 in Acinar Cells to Promote Pancreatic Tumorigenesis in Mice on a High-Fat Diet.

BACKGROUND & AIMS: Obesity is a risk factor for pancreatic cancer. In mice, a high-fat diet (HFD) and expression of oncogenic KRAS lead to development of invasive pancreatic ductal adenocarcinoma (PDAC) by unknown mechanisms. We investigated how oncogenic KRAS regulates the expression of fibroblast growth factor 21, FGF21, a metabolic regulator that prevents obesity, and the effects of recombinant human FGF21 (rhFGF21) on pancreatic tumorigenesis. METHODS: We performed immunohistochemical analyses of FGF21 levels in human pancreatic tissue arrays, comprising 59 PDAC specimens and 45 nontumor tissues. We also studied mice with tamoxifen-inducible expression of oncogenic KRAS in acinar cells (KrasG12D/+ mice) and fElasCreERT mice (controls). KrasG12D/+ mice were placed on an HFD or regular chow diet (control) and given injections of rhFGF21 or vehicle; pancreata were collected and analyzed by histology, immunoblots, quantitative polymerase chain reaction, and immunohistochemistry. We measured markers of inflammation in the pancreas, liver, and adipose tissue. Activity of RAS was measured based on the amount of bound guanosine triphosphate. RESULTS: Pancreatic tissues of mice expressed high levels of FGF21 compared with liver tissues. FGF21 and its receptor proteins were expressed by acinar cells. Acinar cells that expressed KrasG12D/+ had significantly lower expression of Fgf21 messenger RNA compared with acinar cells from control mice, partly due to down-regulation of PPARG expression-a transcription factor that activates Fgf21 transcription. Pancreata from KrasG12D/+ mice on a control diet and given injections of rhFGF21 had reduced pancreatic inflammation, infiltration by immune cells, and acinar-to-ductal metaplasia compared with mice given injections of vehicle. HFD-fed KrasG12D/+ mice given injections of vehicle accumulated abdominal fat, developed extensive inflammation, pancreatic cysts, and high-grade pancreatic intraepithelial neoplasias (PanINs); half the mice developed PDAC with liver metastases. HFD-fed KrasG12D/+ mice given injections of rhFGF21 had reduced accumulation of abdominal fat and pancreatic triglycerides, fewer pancreatic cysts, reduced systemic and pancreatic markers of inflammation, fewer PanINs, and longer survival-only approximately 12% of the mice developed PDACs, and none of the mice had metastases. Pancreata from HFD-fed KrasG12D/+ mice given injections of rhFGF21 had lower levels of active RAS than from mice given vehicle. CONCLUSIONS: Normal acinar cells from mice and humans express high levels of FGF21. In mice, acinar expression of oncogenic KRAS significantly reduces FGF21 expression. When these mice are placed on an HFD, they develop extensive inflammation, pancreatic cysts, PanINs, and PDACs, which are reduced by injection of FGF21. FGF21 also reduces the guanosine triphosphate binding capacity of RAS. FGF21 might be used in the prevention or treatment of pancreatic cancer.

A Consensus Study of the Grading and Typing of Intraductal Papillary Mucinous Neoplasms of the Pancreas.

OBJECTIVE: The grading and typing of intraductal papillary mucinous neoplasms (IPMNs) of the pancreas are challenging for pathologists. We aimed to clarify the points of consistency and disagreement in assessing the grades and types of IPMNs. METHODS: Digital slide images of 20 IPMNs were independently assessed by 10 Japanese pathologists, who then held a consensus meeting to discuss the points of disagreement and develop a consensus and recommendations. RESULTS: The average agreement rates for grade and type were 83.5% (range, 100%-40%) and 82.5% (range, 100%-50%) and the Fleiss' κ values were 0.567 and 0.636, respectively. CONCLUSIONS: The disagreement points and recommendations were as follows: destructed ductal walls with desquamated neoplastic epithelia or mucin lakes partially lined with neoplastic cells could be invasion; intraductal stromal invasion could be dismissed unless vascular or lymphatic invasion existed; elastica staining may help visualize ducts in colloidal nodules; high-grade can be distinguished from low/intermediate grade by marked nuclear disarrangements and complex architecture in the intestinal papillae; oncocytic papillae are characterized by eosinophilic cells with round disoriented nuclei; high-grade gastric papillae can be distinguished from pancreatobiliary papillae by relatively low but complex architecture; and the most dysplastic papillae should be used to assess type in mixed papillae types.

Intratumoral Heterogeneity of SMAD4 Immunohistochemical Expression and Its Role in Prediction of Recurrence Pattern in Patients with Resectable Pancreatic Cancer.

BACKGROUND: The aim of our study was to evaluate consistency of SMAD4 expression in different tumor areas and its correlation with recurrence pattern in patients after resection for pancreatic cancer (PC). METHODS: Records of patients who underwent resection for nonmetastatic PC between 2001 and 2015 were analyzed. Formalin-fixed, paraffin-embedded tissue sections from different areas of primary tumor and lymph node metastases were analyzed immunohistochemically (IHC) for SMAD4 expression using TMA technology. RESULTS: SMAD4 expression was assessed in 356 tissue sections obtained from 91 patients. SMAD4 expression was positive in all assessed tumor slides only in 7 of 26 patients (26.9%). There were 54 recurrences (9 locoregional, 41 distant, and 4 both local and distant) with median follow-up of 21.7 months. There was no correlation between SMAD4 expression and locoregional recurrence pattern (p = 0.30). SMAD4 status influenced neither distant recurrence-free survival (p = 0.99) nor overall survival (p = 0.13). CONCLUSIONS: Different areas inside primary tumor and lymph node metastases express SMAD4 heterogeneously. SMAD4 IHC expression is not a biomarker of the recurrence pattern after surgical resection for PC.

Next generation sequencing of the cellular and liquid fraction of pancreatic cyst fluid supports discrimination of IPMN from pseudocysts and reveals cases with multiple mutated driver clones: First findings from the prospective ZYSTEUS biomarker study.

Approximately half of all pancreatic cysts are neoplastic, mainly comprising intraductal papillary mucinous neoplasms (IPMN), which can progress to invasive carcinoma. Current Fukuoka guidelines have limited sensitivity and specificity in predicting progression of asymptomatic pancreatic cysts. We present first results of the prospective ZYSTEUS biomarker study investigating (i) whether detection of driver mutations in IPMN by liquid biopsy is technically feasible, (ii) which compartment of IPMN is most suitable for analysis, and (iii) implications for clinical diagnostics. Twenty-two patients with clinical inclusion criteria were enrolled in ZYSTEUS. Fifteen cases underwent endoscopic ultrasound (EUS)-guided fine-needle aspiration and cytological diagnostics. Cellular and liquid fraction of the cysts of each case were separated and subjected to deep targeted next generation sequencing (NGS). Clinical parameters, imaging findings (EUS and MRI), and follow-up data were collected continuously. All IPMN cases (n = 12) showed at least one mutation in either KRAS (n = 11) or GNAS (n = 4). Three cases showed both KRAS and GNAS mutations. Six cases harbored multiple KRAS/GNAS mutations. In the three cases with pseudocysts, no KRAS or GNAS mutations were detected. DNA yields were higher and showed higher mutation diversity in the cellular fraction. In conclusion, mutation detection in pancreatic cyst fluid is technically feasible with more robust results in the cellular than in the liquid fraction. Current results suggest that, together with imaging, targeted sequencing supports discrimination of IPMN from pseudocysts. The prospective design of ZYSTEUS will provide insight into diagnostic value of NGS in preoperative risk stratification. Our data provide evidence for an oligoclonal nature of IPMN.

Diagnostic and Prognostic Impact of Neutrophil-to-Lymphocyte Ratio for Intraductal Papillary Mucinous Neoplasms of the Pancreas With High-Grade Dysplasia and Associated Invasive Carcinoma.

OBJECTIVES: The aim of this study was to evaluate the diagnostic and prognostic impact of systemic inflammatory markers for IPMN with high-grade dysplasia (HGD)/invasive carcinoma. METHODS: Neutrophil-to-lymphocyte ratio (NLR), derived NLR, platelet-to-lymphocyte ratio, and C-reactive protein-to-albumin ratio were compared across the different histological grades of 205 IPMN cases. We also tested the diagnostic performance for IPMN with HGD/invasive carcinoma. RESULTS: The median (interquartile range) preoperative NLR was higher in IPMN with HGD/invasive carcinoma (2.03 [1.48-2.93]) than IPMN with low-grade dysplasia (1.74 [1.42-2.24], P = 0.0137). The C-reactive protein-to-albumin ratio and derived NLR values were also significant higher in cases with HGD/invasive carcinoma. A combination assay of NLR, carcinoembryonic antigen, and carbohydrate antigen 19-9 revealed a 58.8% sensitivity and 76.8% specificity. Among the cases with worrisome features, the high NLR values increased the positive predictive value (68.8%) compared with low values (31.8%). In IPMN cases with the associated invasive carcinoma, high NLR values showed association with the deeper vertical invasion and shorter survival periods. CONCLUSIONS: Preoperative NLR, combined with tumor markers and image findings, can be a useful predictive marker for the presence of HGD/invasive carcinoma in IPMNs. Preoperative NLR also predicts the long-term outcomes in IPMN cases with invasive carcinoma.

From somatic mutation to early detection: insights from molecular characterization of pancreatic cancer precursor lesions.

Pancreatic cancer arises from noninvasive precursor lesions, including pancreatic intraepithelial neoplasia (PanIN), intraductal papillary mucinous neoplasm (IPMN), and mucinous cystic neoplasm (MCN), which are curable if detected early enough. Recently, these types of precursor lesions have been extensively characterized at the molecular level, defining the timing of critical genetic alterations in tumorigenesis pathways. The results of these studies deepen our understanding of tumorigenesis in the pancreas, providing novel insights into tumor initiation and progression. Perhaps more importantly, they also provide a rational foundation for early detection approaches that could allow clinical intervention prior to malignant transformation. In this review, we summarize the results of comprehensive molecular characterization of PanINs, IPMNs, and MCNs and discuss the implications for cancer biology as well as early detection. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Pancreatic DCLK1+ cells originate distinctly from PDX1+ progenitors and contribute to the initiation of intraductal papillary mucinous neoplasm in mice.

PanINs and IPMNs are the two most common precursor lesions that can progress to invasive pancreatic ductal adenocarcinoma (PDA). DCLK1 has been identified as a biomarker of progenitor cells in PDA progressed from PanINs. To explore the potential role of DCLK1-expressing cells in the genesis of IPMNs, we compared the incidence of DCLK1-positive cells in pancreatic tissue samples from genetically-engineered mouse models (GEMMs) for IPMNs, PanINs, and acinar to ductal metaplasia by immunohistochemistry and immunofluorescence. Mouse lineage tracing experiments in the IPMN GEMM showed that DCLK1+ cells originated from a cell lineage distinct from PDX1+ progenitors. The DCLK1+ cells shared the features of tuft cells but were devoid of IPMN tumor biomarkers. The DCLK1+ cells were detected in the earliest proliferative acinar clusters prior to the formation of metaplastic ductal cells, and were enriched in the "IPMN niches". In summary, DCLK1 labels a unique pancreatic cellular lineage in the IPMN GEMM. The clustering of DCLK1+ cells is an early event in Kras-induced pancreatic tumorigenesis and may contribute to IPMN initiation.