Expression of CD117, CK17, CK20, MUC4, villin and mismatch repair deficiency in pancreatic intraductal papillary mucinous neoplasm.

Among pancreatic intraductal papillary neoplasms, gastric, intestinal, and pancreatobiliary intraductal papillary mucinous neoplasm (IPMN), intraductal oncocytic papillary neoplasm (IOPN), and intraductal tubulopapillary neoplasm (ITPN) have been defined, differing regarding association with invasive carcinoma and prognosis. Immunohistochemistry (IHC) can help in the distinction of these neoplasms, but a proportion is unclassifiable using recommended markers. Hence, additional markers useful for the typing of pancreatic intraductal papillary neoplasms are needed. The reported frequencies of the different types of IPMNs in surgical series vary to some extent, and such data based on Danish patients are currently lacking. Besides, the role of mismatch repair (MMR) deficiency in these neoplasms has not been fully elucidated. We aimed to evaluate the frequency of different types of pancreatic intraductal papillary neoplasms in a Danish cohort. Furthermore, we aimed to examine the utility of CD117, CK17, CK20, MUC4, and villin as markers for their distinction, in addition to the recommended markers MUC1, MUC2, MUC5AC, MUC6 and CDX2, and to evaluate the frequency of MMR deficiency. We typed 40 consecutively resected pancreatic intraductal papillary neoplasms according to the WHO criteria from 2019. IHC for CD117, CDX2, CK17, CK20, MLH1, MSH2, MSH6, MUC1 (H23), MUC1 (Ma695), MUC2, MUC4, MUC5AC, MUC6, PMS2, and villin was performed and evaluated using a five-tiered semiquantitative scale. A subset of the tumours was examined with PCR for microsatellite instability (MSI). Most tumours were intestinal (40 %) and gastric (40 %) IPMNs, followed by pancreatobiliary (17 %) IPMNs and IOPN (3 %). All cases were MMR proficient. We found a higher expression of MUC4, CK20 and villin in intestinal compared to gastric IPMNs (p < 0.01, p < 0.001 and p < 0.001). MUC4 was more strongly expressed in intestinal compared to pancreatobiliary IPMNs, while the opposite was found for CK17 (p < 0.05 and p < 0.05). IOPN showed strong CD117 expression (score 4), while all gastric IPMNs were negative and 50 % and 29 % of intestinal and pancreatobiliary IPMNs only showed weak expression (score 1). Our data suggest that CK20, MUC4 and villin may aid in the identification of intestinal IPMNs, while CK17 and CD117 may aid in the identification of pancreatobiliary IPMNs and IOPN, in some cases. However, additional studies evaluating these markers in pancreatic intraductal papillary neoplasms are needed.

Role of phosphorylated Smad3 signal components in intraductal papillary mucinous neoplasm of pancreas.

BACKGROUND: Malignant intraductal papillary mucinous neoplasm (IPMN) has poor prognosis. The carcinogenesis of IPMN is not clear. The aim of this study was to clarify transitions in phosphorylated Smad3 signaling during IPMN carcinogenesis. METHODS: By using immunohistochemistry, we examined the expression of pSmad3C and pSmad3L from 51 IPMN surgical specimens resected at our institution between 2010 and 2013. We also examined the expression of Ki-67, c-Myc and p-JNK. RESULTS: The median immunostaining index of pSmad3C was 79.2% in low-grade dysplasia, 74.9% in high-grade dysplasia, and 42.0% in invasive carcinoma (P < 0.01), whereas that of pSmad3L was 3.4%, 4.3%, and 42.4%, respectively (P < 0.01). There was a negative relationship between the expression of pSmad3C and c-Myc (P < 0.001, r = -0.615) and a positive relationship between the expression of pSmad3L and c-Myc (P < 0.001, r = 0.696). Negative relationship between the expression of pSmad3C and Ki-67 (P < 0.01, r = -0.610) and positive relationship between the expression of pSmad3L and Ki-67 (P < 0.01, r = 0.731) were confirmed. p-JNK-positive cells were frequently observed among pSmad3L-positive cancer cells. The median of pSmad3L/pSmad3C ratio in the non-recurrence group and the recurrence group were 0.58 (range, 0.05-0.93), 3.83 (range, 0.85-5.96), respectively (P = 0.02). The median immunostaining index of c-Myc in the non-recurrence group and the recurrence group were 2.91 (range, 0-36.9) and 82.1 (range, 46.2-97.1), respectively (P = 0.02). The median immunostaining index of Ki-67 in the non-recurrence group and the recurrence group were 12.9 (range 5.7-30.8) and 90.9 (range 52.9-98.5), respectively (P = 0.02). CONCLUSIONS: pSmad3L was upregulated in malignant IPMN. pSmad3L/pSmad3C ratio may be a useful prognostic factor in IPMN.

Cross Validation of the Monoclonal Antibody Das-1 in Identification of High-Risk Mucinous Pancreatic Cystic Lesions.

BACKGROUND & AIMS: Although pancreatic cystic lesions (PCLs) are frequently and incidentally detected, it is a challenge to determine their risk of malignancy. In immunohistochemical and enzyme-linked immunosorbent assay (ELISA) analyses of tissue and cyst fluid from pancreatic intraductal papillary mucinous neoplasms, the monoclonal antibody Das-1 identifies those at risk for malignancy with high levels of specificity and sensitivity. We aimed to validate the ability of Das-1 to identify high-risk PCLs in comparison to clinical guidelines and clinical features, using samples from a multicenter cohort. METHODS: We obtained cyst fluid samples of 169 PCLs (90 intraductal papillary mucinous neoplasms, 43 mucinous cystic neoplasms, and 36 non-mucinous cysts) from patients undergoing surgery at 4 tertiary referral centers (January 2010 through June 2017). Histology findings from surgical samples, analyzed independently and centrally re-reviewed in a blinded manner, were used as the reference standard. High-risk PCLs were those with invasive carcinomas, high-grade dysplasia, or intestinal-type intraductal papillary mucinous neoplasms with intermediate-grade dysplasia. An ELISA with Das-1 was performed in parallel using banked cyst fluid samples. We evaluated the biomarker's performance, generated area under the curve values, and conducted multivariate logistic regression using clinical and pathology features. RESULTS: The ELISA for Das-1 identified high-risk PCLs with 88% sensitivity, 99% specificity, and 95% accuracy, at a cutoff optical density value of 0.104. In 10-fold cross-validation analysis with 100 replications, Das-1 identified high-risk PCLs with 88% sensitivity and 98% specificity. The Sendai, Fukuoka, and American Gastroenterological Association guideline criteria identified high-risk PCLs with 46%, 52%, and 74% accuracy (P for comparison to Das-1 ELISA <.001). When we controlled for Das-1 in multivariate regression, main pancreatic duct dilation >5 mm (odds ratio, 14.98; 95% confidence interval, 2.63-108; P < .0012), main pancreatic duct dilation ≥1 cm (odds ratio, 47.9; 95% confidence interval, 6.39-490; P < .0001), and jaundice (odds ratio, 6.16; 95% confidence interval, 1.08-36.7; P = .0397) were significantly associated with high-risk PCLs. CONCLUSIONS: We validated the ability of an ELISA with the monoclonal antibody Das-1 to detect PCLs at risk for malignancy with high levels of sensitivity and specificity. This biomarker might be used in conjunction with clinical guidelines to identify patients at risk for malignancy.

Transmembrane mucin MUC13 distinguishes intraductal papillary mucinous neoplasms from non-mucinous cysts and is associated with high-risk lesions.

BACKGROUND: Intraductal papillary mucinous neoplasms (IPMN) are currently managed based on imaging characteristics and cyst fluid sampling. This study was designed to determine if MUC13, a glycoprotein aberrantly overexpressed in pancreatic adenocarcinoma, might aid in distinguishing high-risk lesions (high grade dysplasia/invasive disease) from low-grade lesions. METHODS: MUC13 immunohistochemical staining was performed on surgically resected formalin-fixed tissue specimens from 49 IPMNs and 23 non-mucinous cysts. Membranous MUC13 expression was measured by H-score, which quantifies staining intensity and the percentage of cells involved (range 0-300). RESULTS: MUC13 expression was detected in all IPMNs and was significantly greater than in non-mucinous cysts (median 210 vs 40, p < 0.001). MUC13 expression was similar among main (n = 26), branch (n = 15), and mixed (n = 8) duct lesions (median 210, 200, 225, respectively). The highest expression was observed in tumors with intestinal and pancreatobiliary histologic features (both median 225) and the lowest in gastric type lesions (median 200). MUC13 expression was significantly greater in high-risk lesions (n = 21) compared to those with low-grade dysplasia (n = 28) (median 250 vs 195, p < 0.001). CONCLUSION: MUC13 expression was significantly greater in high-risk IPMNs in this analysis. The preoperative assessment of MUC13 in cyst fluid samples warrants further investigation.

Extracellular matrix proteins and carcinoembryonic antigen-related cell adhesion molecules characterize pancreatic duct fluid exosomes in patients with pancreatic cancer.

BACKGROUND: Exosomes are nanovesicles that have been shown to mediate carcinogenesis in pancreatic ductal adenocarcinoma (PDAC). Given the direct communication of pancreatic duct fluid with the tumor and its relative accessibility, we aimed to determine the feasibility of isolating and characterizing exosomes from pancreatic duct fluid. METHODS: Pancreatic duct fluid was collected from 26 patients with PDAC (n = 13), intraductal papillary mucinous neoplasm (IPMN) (n = 8) and other benign pancreatic diseases (n = 5) at resection. Exosomes were isolated by serial ultracentrifugation, proteins were identified by mass spectrometry, and their expression was evaluated by immunohistochemistry. RESULTS: Exosomes were isolated from all specimens with a mean concentration of 5.9 ± 1 × 108 particles/mL and most frequent size of 138 ± 9 nm. Among the top 35 proteins that were significantly associated with PDAC, multiple carcinoembryonic antigen-related cell adhesion molecules (CEACAMs) and extracellular matrix (ECM) proteins were identified. Interestingly, CEACAM 1/5 expression by immunohistochemistry was seen only on tumor epithelia whereas tenascin C positivity was restricted to stroma, suggesting that both tumor and stromal cells contributed to exosomes. CONCLUSION: This is the first study showing that exosome isolation is feasible from pancreatic duct fluid, and that exosomal proteins may be utilized to diagnose patients with PDAC.