Pancreatic Cyst Fluid Research Articles

Abstract PO-009: Multi-omic profiling of patient pancreatic cyst fluid for the identification of a novel biomarker panel of patient cancer risk

Abstract Background: Pancreatic cancer was responsible for almost 500,000 deaths globally in 2020 according to GLOBOCAN 2020. Pancreatic cystic lesions (PCLs) are fluid-filled protrusions either on or inside the pancreas. PCLs can either be benign or pre-malignant, however, current guidelines based on clinical features are limited in their ability to accurately stratify patients based on cancer risk. Multi-omic profiling of the pancreatic cyst fluid (PCF) could aid in the identification of a novel biomarker panel of patient cancer risk. Methods: PCF was collected from 40 patients by EUS-FNA. Patients were stratified using the 2018 European evidence-based guidelines into low-risk (n=15), high risk (n=15) and no-risk/pseudocyst (n=10). PCF was sonicated and subsequently processed using a single-pot solid-phase-enhanced sample preparation (SP3) protocol with Sera-Mag SpeedBead carboxylate-modified beads prior to LC-MS. Samples were run on a Thermo Scientific Q Exactive mass spectrometer coupled to a Dionex Ultimate 3000 (RSLCnano) chromatography system. MS-generated proteomic data were analysed in Perseus (v1.6.13). HTG microRNA whole transcriptome sequencing was run on whole PCF. Transcriptomic data were analysed using HTG EdgeSeq Reveal (v3.1). Results: MS-analysis revealed 1,266 proteins present across all PCF samples. Proteins were filtered based on potential contaminants and valid values. Only proteins expressed in a minimum of six PCF samples were included in the analysis. After data clean-up, 465 proteins were examined for differential expression. A total of eight proteins were upregulated in high-risk PCF compared to low-risk (p<0.05, FDR=0.05, s0=0.1). Among them, seven have been shown to be upregulated in pancreatic cancer. Conversely, one protein which is reported to be downregulated in pancreatic cancer, was significantly upregulated in the high-risk patient cohort. A total of 2,096 miRNAs were identified across all PCF samples. MiRNAs were filtered based on fold-change > ±2 between the groups, with 202 miRNAs meeting this criteria. Forty-six miRNAs were significantly upregulated in high-risk PCF compared to low-risk PCF (adj-p<0.05, FDR=0.05, s0=0.1). Five of these miRNAs are known to be upregulated in pancreatic ductal adenocarcinoma (PDAC) tissues. Furthermore, three of the miRNAs are upregulated in the circulation of PDAC patients. Importantly, seven miRNAs identified as being upregulated in high-risk PCF have been shown to be downregulated in PDAC tissues. Differentially expressed proteins and miRNAs are currently being utilised to create an integrated, multi-omic predictive algorithm for patient risk. Conclusion: Multi-omic profiling of pancreatic cyst fluid provides an abundance of potential biomarkers that could be utilised for the stratification of patients into high-and low-risk groups for malignancy. Integration of multi-omic data has the potential to provide more robust biomarker panels of patient risk. Validation of biomarkers in independent patient cohorts will be key to the development of novel clinical biomarkers. Citation Format: Laura E. Kane, Gregory S. Mellotte, Simone Marcone, Barbara M. Ryan, Stephen G. Maher. Multi-omic profiling of patient pancreatic cyst fluid for the identification of a novel biomarker panel of patient cancer risk [abstract]. In: Proceedings of the AACR Virtual Special Conference on Pancreatic Cancer; 2021 Sep 29-30. Philadelphia (PA): AACR; Cancer Res 2021;81(22 Suppl):Abstract nr PO-009.

Co-analysis of pancreatic cyst fluid carcinoembryonic antigen and glucose with novel cut-off levels better distinguishes between mucinous and non-mucinous neoplastic pancreatic cystic lesions.

Pancreatic cyst fluid analysis plays an important role in distinguishing between mucinous and non-mucinous cyst lesions. We aimed to compare the diagnostic performances of cyst fluid carcinoembryonic antigen (CEA), CA 19-9, and glucose in differentiating mucinous from non-mucinous neoplastic pancreatic cystic lesions (PCLs) and determine the best cut-off levels. Patients' data were evaluated retrospectively. 102 patients' PCLs were grouped as non-neoplastic (n = 25), non-mucinous neoplastic (n = 20), mucinous neoplastic (n = 47) and pancreatic adenocarcinomas with cystic degeneration (n = 10); and CEA, CA 19-9, and glucose levels were compared. Receiver-operating characteristic analysis was performed, and the ideal cut-off values were determined. Cyst fluid CEA and CA 19-9, levels were significantly higher (P < 0.001, P < 0.001, respectively) and glucose levels were significantly lower (P = 0.001) in mucinous than in non-mucinous neoplastic PCLs. Area under curve with 95% confidence interval of CEA, glucose and CEA and glucose test combination was 0.939 (95% CI = 0.885-0.993, P = 0.001), 0.809 (95% CI = 0.695-0.924, P < 0.001) and 0.937 (95% CI = 0.879-0.995), respectively. CEA cut-offs to rule-in and rule-out mucinous neoplastic were 135.1ng/mL (sensitivity = 62%, specificity = 94.7%) and 6.12ng/mL (sensitivity = 94.1%, specificity = 80.4%), respectively. Glucose cut-off of 2.8mmol/L was chosen both to rule-in and rule-out mucinous neoplastic PCLs (sensitivity = 78%, specificity = 80%). Co-analysis of CEA and glucose to distinguish mucinous from non-mucinous neoplastic PCLs had sensitivity = 87.8%, specificity = 93.3%, and diagnostic accuracy = 89.3%. We concluded that co-analysis of cyst fluid CEA (cut-off = 135.1ng/mL) and glucose (cut-off = 2.8mmol/L) at novel cut-offs had the best testing performance to rule-in mucinous neoplastic PCLs. To rule-out mucinous PCLs co-analysis of CEA (cut-off = 6.12ng/mL) and glucose (cut-off = 2.8mmol/L) added value to prediction.

Clinical Management of Pancreatic Premalignant Lesions

Clinical Management of Pancreatic Premalignant Lesions

Evaluation of pancreatic cyst prostaglandin E2 as a Marker for Differentiation between Mucinous and Non Mucinous Cysts and Prediction of Dysplasia in Mucinous Pancreatic Cysts

Abstract Background There are different types of pancreatic cysts.They can be classified by different ways. Each of them has different plan of management. that is why their differentiation became mandatory before setting their treatment plans. Aim of the study To Evaluate PGE2 as a marker of differentiation of mucin containing pancreatic cysts and prediction of dysplasia in these cyst. A Secondary aim is differentiation of mucin from non mucin containing pancreatic cystic lesions Patients and Methods 40 patients were recruited from the Department of Internal Medicine and the Outpatient Clinic of Cairo University hospital in the period between December 2018 and August 2019. A case control study design was adopted where the patients were grouped as follow : Group 1 - composed of 20 patients with pancreatic mucinous cystic lesions defined by EUS guided samples, Histopathology and Radiological findings. Group 2: - composed of 20 patients with pancreatic non mucinous cystic lesions defined by EUS guided samples, Histopathology and Radiological findings. Results In our study, there was a significant differences in PGE2 level between true and inflammatory pancreatic cysts (p = 0.001) .However, there is insignificant differences between mucinous, non mucinous (p = 0.406). There was insignificant differences between different grades of IPMN dysplasia (p = 0.615) Conclusion Prostaglandin E2 is pancreatic cystic fluid marker which can be used for differentiation between true pancreatic cysts from inflammatory pancreatic cysts which have different plans of management. Prostaglandin E2 is not good marker to be used for differentiation between different grades of IPMN dysplasia.

Cyst fluid metabolites distinguish malignant from benign pancreatic cysts.

OBJECTIVESCurrent standard of care imaging, cytology, or cystic fluid analysis cannot reliably differentiate malignant from benign pancreatic cystic neoplasms. This study sought to determine if the metabolic profile of cystic fluid could distinguish benign and malignant lesions, as well as mucinous and non-mucinous lesions.MethodsMetabolic profiling by untargeted mass spectrometry and quantitative nuclear magnetic resonance was performed in 24 pancreatic cyst fluid from surgically resected samples with pathological diagnoses and clinicopathological correlation.Results(Iso)-butyrylcarnitine distinguished malignant from benign pancreatic cysts, with a diagnostic accuracy of 89%. (Iso)-butyrylcarnitine was 28-fold more abundant in malignant cyst fluid compared with benign cyst fluid (P=.048). Furthermore, 5-oxoproline (P=.01) differentiated mucinous from non-mucinous cysts with a diagnostic accuracy of 90%, better than glucose (82% accuracy), a previously described metabolite that distinguishes mucinous from non-mucinous cysts. Combined analysis of glucose and 5-oxoproline did not improve the diagnostic accuracy. In comparison, standard of care cyst fluid carcinoembryonic antigen (CEA) and cytology had a diagnostic accuracy of 40% and 60% respectively for mucinous cysts. (Iso)-butyrylcarnitine and 5-oxoproline correlated with cyst fluid CEA levels (P<.0001 and P<.05 respectively). For diagnosing malignant pancreatic cysts, the diagnostic accuracies of cyst size > 3 cm, ≥ 1 high-risk features, cyst fluid CEA, and cytology are 38%, 75%, 80%, and 75%, respectively.Conclusions(Iso)-butyrylcarnitine has potential clinical application for accurately distinguishing malignant from benign pancreatic cysts, and 5-oxoproline for distinguishing mucinous from non-mucinous cysts.

Biomarker Risk Score Algorithm and Preoperative Stratification of Patients with Pancreatic Cystic Lesions

Pancreatic cysts are incidentally detected in up to 13% of patients undergoing radiographic imaging. Of the most frequently encountered types, mucin-producing (mucinous) pancreatic cystic lesions may develop into pancreatic cancer, while nonmucinous ones have little or no malignant potential. Accurate preoperative diagnosis is critical for optimal management, but has been difficult to achieve, resulting in unnecessary major surgery. Here, we aim to develop an algorithm based on biomarker risk scores to improve risk stratification. Patients undergoing surgery and/or surveillance for a pancreatic cystic lesion, with diagnostic imaging and banked pancreatic cyst fluid, were enrolled in the study after informed consent (n= 163 surgical, 67 surveillance). Cyst fluid biomarkers with high specificity for distinguishing nonmucinous from mucinous pancreatic cysts (vascular endothelial growth factor [VEGF], glucose, carcinoembryonic antigen [CEA], amylase, cytology, and DNA mutation) were selected. Biomarker risk scores were used to design an algorithm to predict preoperative diagnosis. Performance was tested using surgical (retrospective) and surveillance (prospective) cohorts. In the surgical cohort, the biomarker algorithm outperformed the preoperative clinical diagnosis in correctly predicting the final pathologic diagnosis (91% vs 73%; p < 0.000001). Specifically, nonmucinous serous cystic neoplasms (SCN) and mucinous cystic neoplasms (MCN) were correctly classified more frequently by the algorithm than clinical diagnosis (96% vs 30%; p < 0.000008 and 92% vs 69%; p= 0.04, respectively). In the surveillance cohort, the algorithm predicted a preoperative diagnosis with high confidence based on a high biomarker score and/or consistency with imaging from ≥1 follow-up visits. A biomarker risk score-based algorithm was able to correctly classify pancreatic cysts preoperatively. Importantly, this tool may improve initial and dynamic risk stratification, reducing overdiagnosis and underdiagnosis.

Early detection of pancreatic cancer using DNA-based molecular approaches.

Due to its poor prognosis and the late stage at which it is typically diagnosed, early detection of pancreatic cancer is a pressing clinical problem. Advances in genomic analysis of human pancreatic tissue and other biospecimens such as pancreatic cyst fluid, pancreatic juice and blood have opened the possibility of DNA-based molecular approaches for early detection of pancreatic cancer. In this Review, we discuss and focus on the pathological and molecular features of precancerous lesions of the pancreas, including pancreatic intraepithelial neoplasia, intraductal papillary mucinous neoplasm and mucinous cystic neoplasm, which are target lesions of early detection approaches. We also discuss the most prevalent genetic alterations in these precancerous lesions, including somatic mutations in the oncogenes KRAS and GNAS as well as tumour suppressor genes CDKN2A, TP53 and SMAD4. We highlight the latest discoveries related to genetic heterogeneity and multifocal neoplasia in precancerous lesions. In addition, we review specific approaches, challenges and clinically available assays for early detection of pancreatic cancer using DNA-based molecular techniques. Although detection and risk stratification of precancerous pancreatic neoplasms are difficult problems, progress in this field highlights the promise of molecular approaches for improving survival of patients with this disease.

Micro-RNA Analysis of Pancreatic Cyst Fluid for Diagnosing Malignant Transformation of Intraductal Papillary Mucinous Neoplasm by Comparing Intraductal Papillary Mucinous Adenoma and Carcinoma.

Although intraductal papillary mucinous neoplasm (IPMN) is thought to be a precursor lesion of pancreatic cancer, diagnosing malignant transformation of IPMN using non-invasive diagnostic methods is difficult and complicated. Micro-RNAs (miRNAs) are currently recognized as biomarkers and molecular targets of various diseases, including malignancy. In this study, we investigated a potential diagnostic approach using miRNA in pancreatic cyst fluid as a marker for evaluating malignant alternation of IPMN. Cystic fluid was sampled mainly during surgical resection. The collected samples were evaluated by performing comprehensive analysis of miRNA using a highly sensitive DNA chip. miRNA expression was compared between IPM adenoma (IPMA) and IPM carcinoma (IPMC) to evaluate the related biomarkers for malignant transformation of IPMN. miRNA analysis revealed that six miRNAs (miR-711, miR-3679-5p, miR-6126, miR-6780b-5p, miR-6798-5p, and miR-6879-5p) in IPMC were significantly enriched compared to those in IPMA. The difference was validated using quantitative real-time PCR. Cyst fluid miRNA analysis might be useful for diagnosing malignant alteration of IPMN. Further evaluations of diagnostic capability as well as functional analysis using the identified miRNAs are required with larger cohorts to confirm its efficacy.

Molecular epidemiology of vancomycin-resistant Enterococcus faecium clinical isolates in a tertiary care hospital in southern Thailand: a retrospective study

ObjectiveVancomycin-resistant enterococci are nosocomial pathogens that are responsible for commonly causing healthcare-associated infections, and they exhibit increased resistance to many antimicrobials, particularly to vancomycin. The epidemiological data available on vancomycin-resistant enterococci (VRE) in Thailand are inadequate.MethodsUsing enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR), this study investigated genes that encode antimicrobial resistance and genetic relatedness to further understand VRE prevalence. Ninety VRE isolates were collected between 2011 and 2019 from a tertiary care hospital in southern Thailand. Antimicrobial susceptibility was determined using the disk diffusion method and E-test methods. Multiplex PCR was performed to detect the van gene and virulence genes.ResultsThe study showed a high prevalence of diverse multidrug-resistant VRE strains. The prevalence of VRE infection was the highest in 2014 (28 isolates, 39.4%). VRE were mostly found in the urogenital tract (26 isolates, 28.9%), followed by the digestive tract (20%), body fluid, i.e., pancreatic cyst fluid, peritoneal dialysis fluid, Jackson–Pratt (JP) drain (20%), and blood specimens (10%). Patients in medical and surgical wards had 71.1% multi-drug-resistant and 28.9% extensively drug-resistant (XDR) VRE strains, respectively. The most prevalent antibiotic resistance was to ampicillin (74.4%). Susceptibility to gentamicin and meropenem were similar (7% and 10%, respectively). Four isolates (4.4%) were resistant to colistin. Only vanA was detected among the strains. The virulence gene test showed that the detection rates of enterococcal surface protein (esp) and hyaluronidase (hyl) genes were 91.1% and 5.6%, respectively. According to ERIC-PCR analysis, 51 of 90 strains had clonality, with a similarity rate of 95%.ConclusionsWe conclude that there is a need to implement infection control practices and active surveillance. Molecular techniques can effectively detect antibiotic-resistant genes, which would allow monitoring to control VRE infection in hospitals.

Intracystic Glucose Levels Appear Useful for Diagnosis of Pancreatic Cystic Lesions: A Systematic Review and Meta-Analysis.

Carcinoembryonic antigen (CEA) in the pancreatic cystic fluid is the most important biomarker for differentiating mucinous from non-mucinous pancreatic cystic lesions (PCLs). However, recent studies have shown that glucose levels in pancreatic cystic fluid can discriminate mucinous from non-mucinous cysts. To perform a meta-analysis to determine the utility of intracystic fluid glucose of pancreatic mucinous cysts compared with intracystic CEA. We conducted a systematic review of the literature in the PubMed, OVID Medline, and Cochrane databases. This meta-analysis considers studies published up to October 2020. Six studies comprising 506 patients were selected; 61.2% of the population was female. Of the 480 PCLs, 287 (59.7%) were mucinous. Pooled sensitivity and specificity of cystic fluid glucose levels for mucinous PCLs were 91% and 85%, respectively. The positive likelihood ratio (PLR) and negative likelihood ratio (NLR) were 6.33 and 0.11, respectively. Pooled diagnostic odds ratio (DOR) was 60.94. The pooled area under the summary receiver operating characteristic (SROC) curve was 0.959. Pooled sensitivity and specificity of pancreatic cystic fluid CEA levels were 61% and 93%. The PLR and NLR were 8.51 and 0.40, respectively. Pooled DOR was 23.52, and the pooled area under the SROC curve was 0.861. Glucose has become a useful method and appears to be better than CEA for differentiating between mucinous PCLs and non-mucinous PCLs. We suggest that the analysis of glucose in PCLs be routinely performed for the differential diagnosis of these lesions.

Comparative analysis of glucose and carcinoembryonic antigen in the diagnosis of pancreatic mucinous cysts: a systematic review and meta-analysis

Pancreatic cystic fluid (PCF) analysis is useful to distinguish between different cyst types and to guide management. The aim of our study was to compare the diagnostic accuracy of glucose level with carcinoembryonic antigen (CEA) in PCF for mucinous cyst diagnosis. We identified studies with PCF obtained by EUS before surgery, with cysts classified as mucinous and nonmucinous according to surgical specimens. A random-effects model was used for quantitative meta-analysis. Pooled sensitivities, specificities, and summary receiver operating characteristic (ROC) curve analysis were conducted. For CEA, we included 31 studies with 5268 patients, of which 2083 were referred for surgery. For glucose, we included 4 studies with 345 patients, of which 275 were referred for surgery. Glucose performed better than CEA for mucinous cysts diagnosis (premalignant and malignant) with sensitivities of .90 (95% confidence interval [CI], .85-.94) and .67 (95% CI, .65-.70), specificities of .82 (95% CI, .72-.89) and .80 (95% CI, 0.76-0.83), and areas under the ROC curve of .96 and .79, respectively. Glucose had a higher sensitivity (90%), with uncommon false-negative results, making it an excellent biomarker to exclude a mucinous cyst. Sensitivity analysis demonstrated that the findings of the current meta-analysis are robust. Glucose level in PCF is more accurate than CEA for preoperative diagnosis of mucinous cysts. It may become a useful first-line test, particularly in small cysts with a limited volume of PCF. Larger studies are awaited to confirm glucose as the single test for mucinous cyst diagnosis.

Molecular analysis of EUS-acquired pancreatic cyst fluid for KRAS and GNAS mutations for diagnosis of intraductal papillary mucinous neoplasia and mucinous cystic lesions: a systematic review and meta-analysis

Although molecular analysis of pancreatic cyst fluid may aid pancreatic cyst classification, clinical practice remains highly variable. Therefore, we performed a systematic review and meta-analysis to evaluate the diagnostic performance of KRAS and GNAS mutations in EUS-acquired pancreatic cyst fluid for diagnosis of intraductal papillary mucinous neoplasms (IPMNs) and mucinous cystic lesions (MCLs). Individualized searches were developed in accordance with Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) and Meta-Analysis of Observational Studies in Epidemiology (MOOSE) guidelines and meta-analysis analyzed according to the Cochrane Diagnostic Test Accuracy working group methodology. A bivariate model was used to compute the pooled sensitivity and specificity and to plot the summary receiver operating characteristics curve with summary point and corresponding 95% confidence interval (95% CI). Six studies (785 lesions) were included. For IPMNs and MCLs, KRAS + GNAS (combination) had significantly higher diagnostic accuracy than KRAS alone and GNAS alone (all P< .001). The pooled sensitivity, specificity, and diagnostic accuracy of KRAS + GNAS mutations for diagnosis of IPMNs were 94% (95% CI, 72-99; I2= 86.74%), 91% (95% CI, 72-98; I2= 89.83), and 97% (95% CI, 95-98), respectively, with each significantly higher compared with carcinoembryonic antigen (CEA) alone (all P< .001). For diagnosis of MCLs, KRAS + GNAS had a similar sensitivity and specificity compared with CEA alone; however, diagnostic accuracy was significantly improved (97% [95% CI, 95-98] vs 89% [95% CI, 86-91]; P< .001). Molecular analysis for KRAS + GNAS mutations in EUS-acquired pancreatic cyst fluid has high sensitivity and specificity with significantly improved diagnostic accuracy for diagnosis of IPMNs and MCLs when compared with CEA alone.

Intracystic Glucose Levels in Differentiating Mucinous From Nonmucinous Pancreatic Cysts: A Systematic Review and Meta-analysis.

Mucinous pancreatic cysts are well reported to transform into pancreatic adenocarcinoma, whereas nonmucinous cysts are mostly benign with low risk for malignant transformation. Nonsurgical methods of differentiating mucinous and nonmucinous pancreatic cysts are challenging and entail a multi investigational approach. Low intracystic glucose levels have been evaluated in multiple studies for its accuracy in differentiating mucinous from nonmucinous cysts of the pancreas. Multiple databases were searched and studies that reported on the utility of intracystic glucose levels in diagnosing mucinous pancreatic cysts were analyzed. Meta-analysis was conducted using the random-effects model, heterogeneity was assessed by I2%, and pooled diagnostic test accuracy values were calculated. Seven studies were included in the analysis from an initial total of 375 citations. The pooled sensitivity of low glucose in differentiating mucinous pancreatic cyst was 90.5% [95% confidence interval (CI): 88.1-92.5; I2=0%] and the pooled specificity was 88% (95% CI: 80.8-92.7; I2=79%). The sensitivity at a glucose cut-off of 50 was 90.1% (95% CI: 87.2-92.5; I2=0%) and the specificity was 85.3% (95% CI: 76.8-91.1; I2=76%). The sensitivity of glucose levels in pancreatic cyst fluid taken by endoscopic ultrasound guided fine-needle aspiration was 90.8% (95% CI: 87.9-93.1; I2=0%) and the specificity was 90.5% (95% CI: 81.7-95.3; I2=83%). The sensitivity of point-of-care glucometers was 89.5% (95% CI: 87.9-93.1; I2=0%) and specificity was 83.9% (95% CI: 68.5-92.6; I2=43%). Low glucose level at a cut-off of 50 mg/dL on fluid samples collected by endoscopic ultrasound guided fine-needle aspiration and analyzed by point-of-care glucometer achieves excellent diagnostic accuracy in differentiating mucinous pancreatic cysts.

KRAS/GNAS-testing by highly sensitive deep targeted next generation sequencing improves the endoscopic ultrasound-guided workup of suspected mucinous neoplasms of the pancreas.

Pancreatic cysts or dilated pancreatic ducts are often found by cross-sectional imaging, but only mucinous lesions can become malignant. Therefore, distinction between mucinous and non-mucinous lesions is crucial for adequate patient management. We performed a prospective study including targeted next generation sequencing (NGS) of cell-free DNA in the diagnostic endoscopic ultrasound (EUS)-guided workup. Pancreatic cyst(s) or main duct fluid obtained by EUS-guided FNA was analysed by carcinoembryonic antigen (CEA), cytology and deep targeted NGS of 14 known gastrointestinal cancer genes (AKT1, BRAF, CTNNB1, EGFR, ERBB2, FBXW7, GNAS, KRAS, MAP2K1, NRAS, PIK3CA, SMAD4, TP53, APC) with a limit of detection down to variant allele frequency of 0.01%. Results were correlated to histopathology and clinical follow-up. One hundred and thirteen patients with pancreatic cyst(s) and/or a dilated pancreatic main duct (≥5 mm) were screened. Sixty-six patients had to be excluded, mainly due to inoperability or small cyst size (≤10 mm). Forty-seven patients were enrolled for further analysis. A final diagnosis was available in 27 cases including 8 negative controls. In 43/47 (91.5%) of patients a KRAS- and/or GNAS-mutation was diagnosed by NGS. 27.0% of the KRAS-mutated and 10.0% of the GNAS-mutated lesions harbored multiple mutations. KRAS/GNAS-testing by NGS, cytology, and CEA had a sensitivity and specificity of 94.7/100%, 38.1/100%, and 42.1/75.0%, respectively. KRAS/GNAS-testing was significantly superior to CEA (P = .0209) and cytology (P = .0016). In conclusion, KRAS/GNAS-testing by deep targeted NGS is a suitable method to distinguish mucinous from non-mucinous pancreatic lesions, suggesting its usage as a single diagnostic test. Results must be confirmed in a larger cohort.

Practical Applications of Molecular Testing in the Cytologic Diagnosis of Pancreatic Cysts

Mucinous pancreatic cysts are precursor lesions of ductal adenocarcinoma. Discoveries of the molecular alterations detectable in pancreatic cyst fluid (PCF) that help to define a mucinous cyst and its risk for malignancy have led to more routine molecular testing in the preoperative evaluation of these cysts. The differential diagnosis of pancreatic cysts is broad and ranges from non-neoplastic to premalignant to malignant cysts. Not all pancreatic cysts—including mucinous cysts—require surgical intervention, and it is the preoperative evaluation with imaging and PCF analysis that determines patient management. PCF analysis includes biochemical and molecular analysis, both of which are ancillary studies that add significant value to the final cytological diagnosis. While testing PCF for carcinoembryonic antigen (CEA) is a very specific test for a mucinous etiology, many mucinous cysts do not have an elevated CEA. In these cases, detection of a KRAS and/or GNAS mutation is highly specific for a mucinous etiology, with GNAS mutations supporting an intraductal papillary mucinous neoplasm. Late mutations in the progression to malignancy such as those found in TP53, p16/CDKN2A, and/or SMAD4 support a high-risk lesion. This review highlights PCF triage and analysis of pancreatic cysts for optimal cytological diagnosis.

A novel microfluidic-based assay to determine high-risk pancreatic cysts for surgery utilizing pancreatic cyst fluid: an international multicenter study

Presenter: Ajay Maker MD | University of California San Francisco Background: Pancreatic cysts are increasingly diagnosed in the general population. We previously reported that the majority of cysts resected in the country are low risk lesions on final pathology, despite multiple established clinical guidelines. Thus, selecting which patients harbor a cyst at high-risk of occult malignancy or malignant degeneration for operative resection remains a clinical dilemma. The aim of the current study is to develop a high-throughout molecular diagnostic platform to identify pancreatic cysts at high risk of malignancy or high-grade dysplasia. Methods: Pancreatic cyst fluid from an international multi-center patient cohort were analyzed on a newly constructed microfluidic TaqMan low density array that was customized with our gene target panel of five miRNAs and seven mRNAs. Signatures were validated utilizing a machine learning random forest model for the optimal accuracy to determine high-risk cysts (high-grade dysplasia or invasive cancer) from low risk cysts for operative intervention. Results: Of 358 pancreatic cyst fluids analyzed, 157 cysts were intraductal papillary mucinous neoplasms (IPMNs), 62 were mucinous cystic neoplasms (MCNs), and the remainder were of various non-mucinous histologies. Of the 219 mucinous cysts, 60 (27%) were found to harbor high-grade dysplasia (HGD) or invasive cancer on final pathological examination. The models were established and then internally validated revealing a maximal accuracy of 87% to determine benign from potentially malignant cysts with an out-of-bag error of 8-12%. Further, the assay performed with an accuracy of 81% to identify high-risk from low-risk mucinous lesions (IPMN+MCN) for operative intervention. Conclusion: A high-throughput, cost-effective microfluidics-based molecular diagnostic assay has been developed that identifies high-risk cystic lesions of the pancreas for surgical evaluation. In this large, international, multi-center study, robust validation sets were performed. Performance of the biosignatures are currently being enhanced with clinical and radiographic features.

Molecular analysis of EUS-acquired pancreatic cyst fluid for KRAS and GNAS mutations for diagnosis of intraductal papillary mucinous neoplasia and mucinous cystic lesions：a systematic review and meta-analysis 1）.

Molecular analysis of EUS-acquired pancreatic cyst fluid for KRAS and GNAS mutations for diagnosis of intraductal papillary mucinous neoplasia and mucinous cystic lesions：a systematic review and meta-analysis^1）.

Pancreatic cyst nodules in clinical practice: ethereal or real?

Presenter: Oswaldo Aguirre MD | Medstar Georgetown University Hospital/Georgetown University School of Medicine Background: Risk stratification in patients with pancreatic cysts continues to rely heavily on structural characteristics, as imaged both by endoscopic ultrasound and axial imaging with CT and MRI. The most recent international guidelines of the IAP use the presence of a cyst wall nodule, the nodule’s size in mm, and the presence of flow within the nodule to help risk stratify patients under investigation for pancreatic cysts. At present, the guidelines do not differentiate between nodules or solid components as seen by EUS and those imaged by CT/MRI. Further, the rate of discordance between modalities is not well known. The aim of this study was to examine the structural characteristics as seen by axial imaging in patients with endoscopically identified nodules and to determine the rate of concordance. Methods: All patients who had endoscopic ultrasound and pancreatic cyst fluid molecular testing at a single institution over a 7 year period (2012-2019) were studied. Cyst fluid molecular alterations, including DNA quantity and quality, KRAS/GNAS mutations, and tumor suppressor gene mutations were included, in addition to routine endoscopic variables. The presence of a nodule and its associated features were recorded on the endoscopic report. Clinical variables, including imaging characteristics, clinical features, and pathologic/cytologic data, were collected retrospectively using primary source data. Continuous and categorical variables were summarized and compared using standard statistical testing. Results: A total of 791 endoscopic ultrasound procedures with associated cyst fluid samples with molecular testing were available from the selected time period. Of those, 93 patients were found to have a pancreatic cyst nodule by high resolution endoscopic ultrasound. The median age of that subset was 68 (SD 14.6), and median body mass index was 26 (SD 6.4). The median cyst size was 20 mm (SD 19 mm) in maximum diameter. 60% of patients with EUS identified nodules were categorized by cyst fluid molecular testing as low risk, and 2% were predicted to have “aggressive” phenotypes. Only 42% of cysts with nodules by EUS had CEA levels >192. Importantly, only 12.8% of patients were noted to have nodules visualized by axial imaging, and just 30% of those demonstrated enhancement. Despite that rate of discordance, 75% of patients had at least one “worrisome” structural imaging feature (e.g. cyst size, solid component, cyst nodules, main duct diameter, growth rate). Conclusion: : The presence of a pancreatic nodule has a prominent place in risk stratification tools for pancreatic cysts, and both high quality endoscopic ultrasound imaging (with cyst fluid analysis) and high resolution axial imaging are central to this determination. However, in practice, there remains a significant rate of discordance between these modalities in the determination of cyst wall nodules, likely due to the inherent differences in resolution and limitations of each. Future study into other variables which modulate the prognostic importance of pancreatic cystic nodules as imaged by both EUS and CT/MRI is needed.

Management of pancreatic cysts and guidelines: what the gastroenterologist needs to know.

The prevalence of pancreatic cysts has increased significantly over the last decade, partly secondary to increased quality and frequency of cross-sectional imaging. While the majority never progress to cancer, a small number will and need to be followed. The management of pancreatic cysts can be both confusing and intimidating due to the multiple guidelines with varying recommendations. Despite the differences in the specifics of the guidelines, they all agree on several high-risk features that should get the attention of any clinician when assessing a pancreatic cyst: presence of a mural nodule or solid component, dilation of the main pancreatic duct (or presence of main duct intraductal papillary mucinous neoplasm), pancreatic cyst size ⩾3–4 cm, or positive cytology on pancreatic cyst fluid aspiration. Other important criteria to consider include rapid cyst growth (⩾5 mm/year), elevated serum carbohydrate antigen 19-9 levels, new-onset diabetes mellitus, or acute pancreatitis thought to be related to the cystic lesion.

Isolation of pancreatic microbiota from cystic precursors of pancreatic cancer with intracellular growth and DNA damaging properties

ABSTRACT Emerging research suggests gut microbiome may play a role in pancreatic cancer initiation and progression, but cultivation of the cancer microbiome remains challenging. This pilot study aims to investigate the possibility to cultivate pancreatic microbiome from pancreatic cystic lesions associated with invasive cancer. Intra-operatively acquired pancreatic cyst fluid samples showed culture-positivity mainly in the intraductal papillary mucinous neoplasm (IPMN) group of lesions. MALDI-TOF MS profiling analysis shows Gammaproteobacteria and Bacilli dominate among individual bacteria isolates. Among cultivated bacteria, Gammaproteobacteria, particularly Klebsiella pneumoniae, but also Granulicatella adiacens and Enterococcus faecalis, demonstrate consistent pathogenic properties in pancreatic cell lines tested in ex vivo co-culture models. Pathogenic properties include intracellular survival capability, cell death induction, or causing DNA double-strand breaks in the surviving cells resembling genotoxic effects. This study provides new insights into the role of the pancreatic microbiota in the intriguing link between pancreatic cystic lesions and cancer.