Exploring dual effects of dinutuximab beta on cell death and proliferation of insulinoma.

Insulinoma INS-1 cells are pancreatic beta cell tumors. Dinutuximab beta (DB) is a monoclonal antibody used in the treatment of neuroblastoma. The aim of this study is to investigate the effects of DB on pancreatic beta cell tumors at the molecular level. DB (Qarziba®) was available from EUSA Pharma. Streptozotocin (STZ) was used induce to cell cytotoxicity. DB was applied to the cells before or after the STZ application. KCND3, KCNN4, KCNK1, and PTHrP gene expression levels were analyzed by q-RT-PCR, and protein levels were analyzed by Western blotting. Analysis of glucose-stimulated insulin secretion was performed. Ca+2 and CA19-9 levels were determined by the ELISA kit. PERK, CHOP, HSP90, p-c-Jun, p-Atf2, and p-Elk1 protein levels were analyzed by simple WES. Decreased KCND3, KCNK1, and PTHrP protein levels and increased KCND3, KCNN4, KCNK1, and PTHrP gene expression levels were observed with DB applied after STZ application. Cell dysfunction was detected with DB applied before and after STZ application. Ca19-9 and Ca+2 levels were increased with DB applied after STZ application. PERK, CHOP, and p-Elk1 levels decreased, while HSP90 levels increased with DB applied after STZ application. CHOP, p-Akt-2, and p-c-Jun levels increased in the DB group. As a result, INS-1 cells go to cell death via the ERK signaling pathway without ER stress and release insulin with the decrease of K+ channels and an increase in Ca+2 levels with DB applied after STZ application. Moreover, the cells proliferate via JNK signaling with DB application. DB holds promise for the treatment of insulinoma. The study should be supported by in vivo studies.

ESE3-positive PSCs drive pancreatic cancer fibrosis, chemoresistance and poor prognosis via tumour-stromal IL-1ß/NF-κB/ESE3 signalling axis.

BACKGROUND: Desmoplastic stroma, a feature of pancreatic ductal adenocarcinoma (PDAC), contains abundant activated pancreatic stellate cells (PSCs). How PSCs promote PDAC progression remains incompletely understood. METHODS: Effect of epithelium-specific E-twenty six factor 3 (ESE3)-positive PSCs on PDAC fibrosis and chemoresistance was examined by western blot, RT-PCR, immunofluorescence, flow cytometry assay, chromatin immunoprecipitation, luciferase assay, immunohistochemistry and subcutaneous pancreatic cancer mouse model. RESULTS: ESE3 expression increased in PSCs in PDAC tissues compared with those in normal PSCs. Clinical data showed that ESE3 upregulation in PSCs was positively correlated with tumour size, pTNM stage, CA19-9, carcinoembryonic antigen and serum CA242 level. ESE3 overexpression in PSCs was an independent negative prognostic factor for disease-free survival and overall survival amongst patients with PDAC. Mechanistically, the conditional medium from the loss and gain of ESE3-expressing PSCs influenced PDAC chemoresistance and tumour growth. ESE3 directly induced the transcription of α-SMA, collagen-I and IL-1ß by binding to ESE3-binding sites on their promoters to activate PSCs. IL-1ß upregulated ESE3 in PSCs through NF-κB activation, and ESE3 was required for PSC activation by tumour cell-derived IL-1ß. CONCLUSION: Inhibiting the IL-1ß/ESE3 (PSCs)/IL-1ß-positive feedback loop is a promising therapeutic strategy to reduce tumour fibrosis and increase chemotherapeutic efficacy in PDAC.

Protein biomarkers in pancreatic juice and serum for identification of pancreatic cancer.

BACKGROUND AND AIMS: To date, surveillance of high-risk individuals for pancreatic ductal adenocarcinoma (PDAC) has not lived up to expectations, as identification of curable stages through imaging remains challenging. Biomarkers are therefore needed. Pancreatic juice (PJ) may be a promising source, because it is in direct contact with the ductal epithelial lining from which PDAC arises. We aimed to develop a panel of biomarkers from serum and PJ to detect PDAC for future surveillance purposes. METHODS: All patients who underwent PJ collection on secretin stimulation at the Erasmus MC were included. Both PJ and serum were evaluated. Protein levels were determined by the Lowry assay. Potential biomarkers (interleukin-8, interferon-γ, neutrophil gelatinase-associated lipocalin [NGAL], mucin 5, subtype AC [MUC5AC], mucin 2, phospholipase A2 group IB) were selected based on previously reported outcomes and assessed with enzyme-linked immunosorbent assay. Serum carbohydrate antigen 19-9 (CA19-9) values were determined by electrochemiluminescence immunoassay. RESULTS: This study included 59 cases and 126 surveilled control subjects (who underwent PJ collection), of whom 71 had a hereditary predisposition (35 genetic, 36 familial) and 55 had (suspected neoplastic) pancreatic cysts. CA19-9 values were available for 53 cases and 48 control subjects. Serum CA19-9, as well as PJ interleukin-8, NGAL and MUC5AC, were associated with PDAC independent of age, gender, and presence of diabetes mellitus. Serum CA19-9 had a significantly higher area under the curve (AUC; .86; 95% confidence interval [CI], .79-.94) than individual PJ markers (AUC, .62-.70). A combination of PJ markers and serum CA19-9 (panel 2: sensitivity 42% [95% CI, 29-57] and specificity 96% [95% CI, 86-100]) did not improve diagnostic performance compared with CA19-9 alone (sensitivity 70% [95% CI, 56-82] and specificity 85% [95% CI, 72-94]). CONCLUSIONS: High levels of serum CA19-9 and PJ-derived proteins are associated with PDAC. Prospective surveillance studies including individuals at risk of developing PDAC are required to validate these findings.

Application of Proteomics in Pancreatic Ductal Adenocarcinoma Biomarker Investigations: A Review.

Pancreatic ductal adenocarcinoma (PDAC), a highly aggressive malignancy with a poor prognosis is usually detected at the advanced stage of the disease. The only US Food and Drug Administration-approved biomarker that is available for PDAC, CA 19-9, is most useful in monitoring treatment response among PDAC patients rather than for early detection. Moreover, when CA 19-9 is solely used for diagnostic purposes, it has only a recorded sensitivity of 79% and specificity of 82% in symptomatic individuals. Therefore, there is an urgent need to identify reliable biomarkers for diagnosis (specifically for the early diagnosis), ascertain prognosis as well as to monitor treatment response and tumour recurrence of PDAC. In recent years, proteomic technologies are growing exponentially at an accelerated rate for a wide range of applications in cancer research. In this review, we discussed the current status of biomarker research for PDAC using various proteomic technologies. This review will explore the potential perspective for understanding and identifying the unique alterations in protein expressions that could prove beneficial in discovering new robust biomarkers to detect PDAC at an early stage, ascertain prognosis of patients with the disease in addition to monitoring treatment response and tumour recurrence of patients.

GLRX3, a novel cancer stem cell-related secretory biomarker of pancreatic ductal adenocarcinoma.

BACKGROUND: Cancer stem cells (CSCs) are implicated in carcinogenesis, cancer progression, and recurrence. Several biomarkers have been described for pancreatic ductal adenocarcinoma (PDAC) CSCs; however, their function and mechanism remain unclear. METHOD: In this study, secretome analysis was performed in pancreatic CSC-enriched spheres and control adherent cells for biomarker discovery. Glutaredoxin3 (GLRX3), a novel candidate upregulated in spheres, was evaluated for its function and clinical implication. RESULTS: PDAC CSC populations, cell lines, patient tissues, and blood samples demonstrated GLRX3 overexpression. In contrast, GLRX3 silencing decreased the in vitro proliferation, migration, clonogenicity, and sphere formation of cells. GLRX3 knockdown also reduced tumor formation and growth in vivo. GLRX3 was found to regulate Met/PI3K/AKT signaling and stemness-related molecules. ELISA results indicated GLRX3 overexpression in the serum of patients with PDAC compared to that in healthy controls. The sensitivity and specificity of GLRX3 for PDAC diagnosis were 80.0 and 100%, respectively. When GLRX3 and CA19-9 were combined, sensitivity was significantly increased to 98.3% compared to that with GLRX3 or CA19-9 alone. High GLRX3 expression was also associated with poor disease-free survival in patients receiving curative surgery. CONCLUSION: Overall, these results indicate GLRX3 as a novel diagnostic marker and therapeutic target for PDAC targeting CSCs.

Increased carbohydrate antigen 19-9 expression in a thymic neuroendocrine tumor.

Here, we report a case of carbohydrate antigen (CA) 19-9-producing mediastinal neuroendocrine tumor (NET) (atypical carcinoid). A 54-year-old woman with no specific relevant medical history was referred to our hospital because of increased CA19-9 (95.3 U/ml) detected on health screening. Chest computed tomography (CT) revealed an anterior mediastinal mass without localized lymphadenopathy. Thoracic surgery was performed and the histopathological diagnosis was thymic CA19-9-positive NET. The patient developed mediastinal lymph node metastasis at 1 year (CA19-9: 413 U/ml) and multiple bone metastases 4 years (CA19-9: 2303 U/ml) after surgery. Increased CA19-9 levels paralleled the clinical courses of relapse. To our knowledge, this is the first report of CA19-9-producing thymic NET.

Lymphocyte-to-monocyte ratio combined with CA19-9 for predicting postoperative recurrence of colorectal cancer in patients with diabetes.

OBJECTIVE: To investigate the significance of lymphocyte-to-monocyte ratio (LMR) combined with carbohydrate antigen (CA) 19-9 for predicting postoperative recurrence of colorectal cancer (CRC) in patients with type II diabetes. METHODS: We conducted a retrospective analysis of 106 postoperative patients with stage II-III CRC and with type II diabetes. Their clinical indexes such as LMR and CA19-9 were collected, and the patients were followed up for 5 years. RESULTS: The CA19-9 level was 119.7 U/ml at baseline in the relapsed group, while this was 24.81 U/ml in non-relapsed group (p = 0.001). On the contrary, the LMR level was 5.10 and 2.57 for non-relapsed and relapsed group (p < 0.001), respectively. Kaplan-Meier survival curves stratified by CA19-9 and LMR suggested that patients with lower CA19-9 had higher survival probability (p < 0.001), while patients with high LMR level had higher survival probability (p < 0.001). The multivariable Cox proportional hazard regression analysis with CA19-9 and LMR indicated that although the baseline CA19-9 is significantly associated with increasing risk of disease recurrence, the HR (HR = 1.0, 95% CI 1.00-1.01) was small and close to 1, whereas the high baseline LMR (HR = 0.44, 95% CI 0.32-0.61) was associated with decrease in disease recurrence. Model with continuous CA19-9 and LMR was able to better predict (AUC 73.17%) the disease recurrence. CONCLUSION: LMR combined with CA19-9 may become a new index for predicting postoperative recurrence of CRC in patients with diabetes.

Carbohydrate Antigen 19-9 Is an Invasive Malignancy Preoperative Prognostic Factor for Intraductal Papillary Mucinous Neoplasms.

INTRODUCTION: This study aimed to determine the preoperative clinicophysiological and postoperative clinicopathological predictors of malignancy in patients with intraductal papillary mucinous neoplasm (IPMN). METHODS: This was a retrospective observational study. We included 121 patients (73 men and 48 women; mean age: 68.7 years) who had undergone pancreatic resection for IPMN between 2007 and 2018. These patients were grouped into invasive carcinoma (IPMN-INV, N = 21) and low/high-grade IPMN (IPMN-LG/HG, N = 100) groups. Univariate and multivariate analyses of clinicophysiological parameters were carried out. These parameters were also compared between the IPMN-INV/HG (N = 53) and IPMN-LG (N = 68) groups. Survival analyses according to macroscopic type and IPMN subtypes were performed. RESULTS: On univariate analysis, age (p = 0.038), carbohydrate antigen (CA) 19-9 (p < 0.001), IPMN macroscopic type (p = 0.001), IPMN subtype (p < 0.001), pancreatic duct diameter (p < 0.001), and mural nodule (p = 0.042), between IPMN-INV and IPMN-LG/HG were found to be significant prognostic factors of malignancy. CA 19-9 was found to be an independent prognostic factor of IPMN malignancy on multivariate analysis (p = 0.035). The 1-, 3-, and 5-year overall survival (OS) rates of the IPMN-INV and IPMN-LG/HG groups were 94.4/100%, 94.4/100%, and 67.2/100%, respectively. The OS rate in the IPMN-LG/HG group was significantly higher than that in the IPMN-INV group (p < 0.001). On univariate analysis, platelet (p = 0.043), CA 19-9 (p = 0.039), prognostic nutritional index (p = 0.034), platelet/lymphocyte ratio (p = 0.01), IPMN macroscopic type (p < 0.001), IPMN subtype (p < 0.001), pancreatic duct diameter (p = 0.036), and mural nodule (p = 0.032) between IPMN-INV/HG and IPMN-LG were found to be significant prognostic factors of malignancy. On multivariate analysis, CA 19-9 was found to be an independent prognostic factor (p = 0.042) between IPMN-INV/HG and IPMN-LG of malignancy. The 1-, 3-, and 5-year OS rates of the IPMN-INV/HG and IPMN-LG groups were 97.9/100%, 97.9/100%, and 82.6/100%, respectively. The OS rate was significantly higher in the IPMN-LG group than in the IPMN-INV/HG group (p = 0.03). No significant differences in survival were observed in patients with macroscopic tumors (p= 0.544). CONCLUSION: CA 19-9 is an independent invasive malignancy predictor of IPMN.

A Low Total Psoas Muscle Area Index Is a Strong Prognostic Factor in Metastatic Pancreatic Cancer.

OBJECTIVES: The total psoas area index (TPI) is an emerging alternative to the total skeletal muscle area index as a prognostic factor but has never been evaluated in metastatic pancreatic cancer (mPC). METHODS: Areas were manually recorded, as previously described. Sex-specific cutoffs were identified by optimum stratification of TPI using log-rank χ2 statistic associated with mortality to define sarcopenic psoas. Progression-free survival (PFS) and overall survival (OS) were the primary objectives. Two period groups were used as internal validation. RESULTS: During the period study, 79 patients were treated for mPC. The TPI was correlated with PFS (hazards ratio, 0.81; P = 0.02) and OS (hazards ratio, 0.7; P < 0.001). Optimum thresholds defining sarcopenic psoas were less than 5.73 cm2/m2 in men and less than 4.37 cm2/m2 in women. Patients with sarcopenic psoas (62.0%) had shorter median PFS (2.9 months) compared with the others (6.6 months, adjusted P log-rank = 0.01), independently to the intensity of chemotherapy, weight loss, and performance status greater than 1. Similarly, OS was independently shorter in patients with sarcopenic psoas (7.6 months) versus the others (22.2 months, adjusted P < 0.001). These results were confirmed in the 2 period groups. CONCLUSIONS: A low TPI is a stronger independent prognostic factor in mPC.

Sources of variation and establishment of Russian reference intervals for major hormones and tumor markers.

OBJECTIVES: A multicenter study was organized to explore sources of variation (SVs) of reference values (RVs) for 22 major immunochemistry analytes and to determine reference intervals (RIs) for the Russian population. METHODS: According to IFCC Committee on Reference Intervals and Decision Limits (C-RIDL) protocol, 758 healthy volunteers were recruited in St. Petersburg, Moscow, and Yekaterinburg. Serum samples were tested for five tumor markers, 17 hormones and related tests by Beckman Coulter's UniCel DxI 800 immunochemistry analyzer. SVs were explored using multiple regression analysis and ANOVA. Standard deviation ratio (SDR) of 0.4 was used as primary guide for partitioning RIs by gender and age. RESULTS: SDR for between-city difference was <0.4 for all analytes. Secondary exclusion of individuals was done under the following conditions: for female sex-hormones, those with contraceptives (8%); for CA19-9, those supposed to have negative Lewis blood-group (10.5% males and 11.3% females); for insulin, those with BMI≥28 kg/m2 (31%); for the thyroid panel, those with anti-thyroid antibodies (10.3% males; 24.5% females), for CEA those with smoking habit (30% males and 16% females). Gender-specific RIs were required for all analytes except CA19-9, CA15-3, thyroid-related tests, parathyroid hormone, and insulin. Age-specific RIs were required for alpha-fetoprotein, CEA, all sex-hormones for females, FSH and progesterone for both sexes. RIs were generally derived by parametric method after Gaussian transformation using modified Box-Cox formula. Exceptions were growth hormone, estradiol for females in postmenopause, and progesterone for females in premenopause, for which nonparametric method was required due to bimodal distribution and/or insufficient detection limit. CONCLUSION: RIs for major hormones and tumor markers specific for the Russian population were derived based on the up-to-date internationally harmonized protocol by careful consideration of analyte-specific SVs.

Specific Norovirus Interaction with Lewis x and Lewis a on Human Intestinal Inflammatory Mucosa during Refractory Inflammatory Bowel Disease.

Inflammatory bowel disease (IBD), which includes Crohn's disease (CD) and ulcerative colitis (UC), is related to immunological and microbial factors, with the possible implication of enteric viruses. We characterized the interaction between human noroviruses (HuNoVs) and blood group antigens in refractory CD and UC using HuNoV virus-like particles (VLPs) and histological tissues. Immunohistochemistry was conducted on inflammatory tissue samples from the small intestine, colon, and rectum in 15 CD and 9 UC patients. Analysis of the regenerative mucosa of the colon and rectum revealed strong expression of sialylated Lewis a (sLea) and Lewis x (sLex) antigens and HuNoV VLP binding in the absence of ABO antigen expression in both UC and CD. Competition experiments using sialidase, lectins, and monoclonal antibodies demonstrated that HuNoV attachment mostly involved Lea and, to a lesser extent, Lex moieties on regenerative mucosa in both UC and CD. Further studies will be required to understand the implications of specific HuNoV binding to regenerative mucosa in refractory IBD.IMPORTANCE Inflammatory bowel diseases (IBD), including Crohn's disease (CD) and ulcerative colitis (UC), are progressive diseases affecting millions of people each year. Flare-ups during IBD result in severe mucosal alterations of the small intestine (in CD) and in the colon and rectum (in CD and UC). Immunohistochemical analysis of CD and UC samples showed strong expression of known tumoral markers sialyl Lewis a (CA19.9) and sialyl Lewis x (CD15s) antigens on colonic and rectal regenerative mucosa, concurrent with strong human norovirus (HuNov) VLP GII.4 affinity. Sialidase treatment and competition experiments using histo-blood group antigen (HBGA)-specific monoclonal antibodies and lectins clearly demonstrated the implication of the Lewis a moiety and, to a lesser extent, the Lewis x moiety in HuNov recognition in regenerative mucosa of CD and UC tissues. Further studies are required to explore the possible implications of enteric viruses in the impairment of epithelial repair and dysregulation of inflammatory pathways during severe IBD.

Longitudinal analysis of cell-free mutated KRAS and CA 19-9 predicts survival following curative resection of pancreatic cancer.

BACKGROUND: Novel biomarkers and molecular monitoring tools hold potential to improve outcome for patients following resection of pancreatic ductal adenocarcinoma (PDAC). We hypothesized that the combined longitudinal analysis of mutated cell-free plasma KRAS (cfKRASmut) and CA 19-9 during adjuvant treatment and follow-up might more accurately predict disease course than hitherto available parameters. METHODS: Between 07/2015 and 10/2018, we collected 134 plasma samples from 25 patients after R0/R1-resection of PDAC during adjuvant chemotherapy and post-treatment surveillance at our institution. Highly sensitive discriminatory multi-target ddPCR assays were employed to screen plasma samples for cfKRASmut. cfKRASmut and CA 19-9 dynamics were correlated with recurrence-free survival (RFS) and overall survival (OS). Patients were followed-up until 01/2020. RESULTS: Out of 25 enrolled patients, 76% had undergone R0 resection and 48% of resected PDACs were pN0. 17/25 (68%) of patients underwent adjuvant chemotherapy. Median follow-up was 22.0 months, with 19 out of 25 (76%) patients relapsing during study period. Median RFS was 10.0 months, median OS was 22.0 months. Out of clinicopathologic variables, only postoperative CA 19-9 levels and administration of adjuvant chemotherapy correlated with survival endpoints. cfKRASmut. was detected in 12/25 (48%) of patients, and detection of high levels inversely correlated with survival endpoint. Integration of cfKRASmut and CA 19-9 levels outperformed either individual marker. cfKRASmut outperformed CA 19-9 as dynamic marker since increase during adjuvant chemotherapy and follow-up was highly predictive of early relapse and poor OS. CONCLUSIONS: Integrated analysis of cfKRASmut and CA 19-9 levels is a promising approach for molecular monitoring of patients following resection of PDAC. Larger prospective studies are needed to further develop this approach and dissect each marker's specific potential.

Inflammation, Tumoral Markers and Interleukin-17, -10, and -6 Profiles in Pancreatic Adenocarcinoma and Chronic Pancreatitis.

BACKGROUND: Interleukin profiles can be used as biochemical markers regarding the early diagnosis of pancreatic cancer. AIMS: To assess CRP, CA 19-9, CEA levels, and interleukin-6, -10, and -17 profiles in pancreatic ductal adenocarcinoma, chronic pancreatitis was compared with a control group, and the correlation with pancreatic cancer survival. METHODS: A total of 87 patients were prospective divided in pancreatic cancer (n = 53), chronic pancreatitis (n = 22) ,and control group (n = 12). The diagnosis of PDAC was made histologically. The diagnosis of chronic pancreatitis was based on medical history, imaging methods, and endoscopic ultrasound. Systemic concentrations of interleukins were measured using ELISA kits. The patients were followed at 1, 3, and 6 months. RESULTS: CRP, CA 19-9, and CEA were higher in the pancreatic cancer group (p < 0.001). Interleukin-10 was significantly higher in the pancreatic cancer and chronic pancreatitis groups (p < 0.001). Interleukin-17 was statistically higher in the pancreatic cancer group (p < 0.0001). The cut-off of interleukin-17 of 0.273 had a sensitivity of 90.9 and a specificity of 80.9 with a curve under ROC of 0.80 in order to differentiate between pancreatic cancer and chronic pancreatitis. The serum levels of interleukins are not correlated with the stage of the disease. CRP, CA 19-9, CEA, and interleukin-6, -10, and -17 were lower in patients with survival more than 6 months. CONCLUSIONS: We detected high levels of interleukin-6, -10, and -17 in chronic pancreatitis and pancreatic cancer. Serum interleukin-17 levels can discriminate between pancreatic cancer and chronic pancreatitis. The prognostic role of interleukins needs to be established.

Pancreatic body and tail cancer and favorable metastatic lymph node behavior on the left edge of the aorta.

BACKGROUND: Lymph node (LN) metastasis in pancreatic body-tail cancer is a poor prognostic factor and the optimal LN dissection area for distal pancreatectomy (DP) remains unclear. Lymphatic flow from the tumors is thought to depend on the tumor sites. We examined LN metastasis frequency based on tumor site and recurrent patterns post-DP. METHODS: With a retrospective, single institutional study, we examined 100 patients who underwent DP as an upfront surgery for pancreatic cancer over 17 years. Tumor sites were classified as tumor confined to pancreatic body (and neck) (Pb(n)); and pancreatic tail (Pt). We compared metastatic LN and recurrence patterns based on tumor site. The median overall survival (OS) and disease-free survival (DFS) were analyzed. RESULTS: LN metastasis occurred in 59/100 (59.0%), with 23 and 25 tumors located in the Pb(n), and Pt, respectively. Those with the tumor in Pt had metastases to #10, #11d/p, and #18 LN mainly. However, the patients with the Pb(n) tumor had metastases to #8a/p, #11p, and #14p/d LN. There was no metastasis to #10 and #11d LN. The OS and DFS were 34 and 15 months, respectively. No significant difference was found in the OS, DFS, and recurrence patterns based on tumor sites. CONCLUSION: Differences in metastatic LN sites were observed in pancreatic body-tail cancer when tumors were confined to the left or right of the left aortic edge. Although it is necessary to validate this finding with a large-scale study, organ-preserving DP might be a treatment option for selected patients depending on the tumor sites.

Profiling of gastric cancer cell-surface markers to achieve tumour-normal discrimination.

BACKGROUND: Differentiating between malignant and normal cells within tissue samples is vital for molecular profiling of cancer using advances in genomics and transcriptomics. Cell-surface markers of tumour-normal discrimination have additional value in terms of translatability to diagnostic and therapeutic strategies. In gastric cancer (GC), previous studies have identified individual genes or proteins that are upregulated in cancer. However, a systematic analysis of cell-surface markers and development of a composite panel involving multiple candidates to differentiate tumour from normal has not been previously reported. METHODS: Whole transcriptome sequencing (WTS) of GC and matched normal samples from the Singapore Gastric Cancer Consortium (SGCC) was used as a discovery cohort to identify upregulated putative cell-surface proteins. Matched WTS data from the The Cancer Genome Atlas (TCGA) was used as a validation cohort. Promising candidates from this analysis were validated orthogonally using multispectral immunohistochemistry (mIHC) with automated quantitative analysis using the Vectra platform. mIHC was performed on a tissue microarray containing matched normal, marginal and tumour tissues. The receiver-operating characteristic (ROC) curves were analysed to identify markers with the highest diagnostic validity independently and in combination. RESULTS: Analysis of putative membrane protein transcripts from the SGCC discovery cohort WTS data (n=15 matched tumour and normal pairs) identified several differentially and highly expressed candidates in tumour compared with normal tissues. After validation with TCGA data (n=29 matched tumour and normal pairs), the following proteins were selected for mIHC analysis: CEACAM5, CEACAM6, CLDN4, CLDN7, and EpCAM. These were compared with established glycoprotein markers in GC, namely CA19-9 and CA72-4. Individual ROC curves yielded the best performance for CEACAM5 (area under the ROC curve (AUC)=0.80), CEACAM6 (AUC=0.82), EpCAM (AUC=0.83), and CA72-4 (AUC=0.76). Combined multiplexed imaging of these four markers revealed improved specificity and sensitivity for detection of tumour from normal tissue (AUC of 4-plex=0.91). CONCLUSION: CEAMCAM5, CEACAM6, EpCAM, and CA72-4 form a versatile set of markers for robust discrimination of GC from adjacent normal tissue. As cell-surface markers, they are compatible with both IHC and live imaging approaches. These candidates may be exploited to improve automated identification of tumour tissue in GC.

Mucinous cystic neoplasms of the pancreas associated with pregnancy: Two case reports.

RATIONALE: Although rare, pancreatic neoplasms can occur during pregnancy, both in benign and malignant forms. Mucinous cystic neoplasms (MCNs) of the pancreas, a type of these neoplasms, are precursor lesions to invasive pancreatic cancer. The presence of the ovarian-type stroma is a defining feature. PATIENT CONCERNS: The first case was a 38-year-old woman in her 18th week of pregnancy with abdominal pain that worsens a few weeks later. The second case was a 30-year-old woman in her 17th week of pregnancy with abdominal pain in the left hypochondrium. DIAGNOSIS: The patients were under clinical examination and laboratory test including carbohydrate antigen 19-9 (CA 19-9) and carcinoembryonic antigen (CEA). Both patients had magnetic resonance imaging (MRI). The diagnosis of a MCNs of the pancreas was done preoperatively in the 2 cases. INTERVENTIONS: Both patients underwent distal pancreatectomy during pregnancy. One of them was an emergency laparotomy because of a ruptured MCN. OUTCOMES: Both patients were completely recovered from distal pancreatectomy and continued to full term, delivering a healthy baby by Caesarean section. After 6 years of follow-up, the first patient underwent a total gastrectomy, because of a gastric cancer with carcinomatosis. Currently the 2 patients are still alive after 8 years and 5 years of follow-up, respectively. LESSONS: Surgical resection of MCNs during pregnancy should be considered during the second trimester given common distal pancreas location, rapid growth, risk of spontaneous rupture, and malignant potential.

Validation of the clinical utility of 4 guidelines in the initial triage of mucinous cystic lesions of the pancreas based on cross-sectional imaging: Experience with 188 surgically-treated patients.

INTRODUCTION: Over the years, several guidelines have been introduced to guide management of mucinous pancreatic cystic neoplasms (mPCN). In this study, we aimed to evaluate and compare the clinically utility of the Sendai-06, Fukuoka-12, Fukuoka-17 and European-18 guidelines in predicting malignancy of mPCN. METHODS: One hundred and eighty-eight patients with mucinous cystic neoplasms (MCN) or intraductal papillary mucinous neoplasm (IPMN) who underwent surgery were retrospectively reviewed and classified under the 4 guidelines. Malignancy was defined as high grade dysplasia and invasive carcinoma. RESULTS: Raised CA19-9>37U/ml, enhancing mural nodule≥5 mm and main pancreatic duct≥10 mm were significantly associated with malignancy on multivariate analysis. Increasing number of high risk features, absolute indications (European-18), worrisome risk or relative indications (European-18) were significantly associated with an increased likelihood of malignancy. The positive predictive values (PPV) of high risk features for Sendai-06, Fukuoka-12, Fukuoka-17 and absolute indications (European-18) for malignancy were 53%, 76%, 78% and 78% respectively. The negative predictive values (NPV) of the Sendai-06, Fukuoka-12 and Fukuoka-17 were 100%, while that of the European-18 was 92%. Risk of malignancy for patients with ≥4 worrisome features (Fukuoka-17) and ≥3 relative indications (European-18) was 66.7% and 75.0% respectively. CONCLUSIONS: All 4 guidelines studied were useful in the initial triage of mPCN for the risk stratification of malignancy. The Fukuoka-17 had the highest PPV and NPV.

Prognostic Impact of Desmoplastic Reaction Evaluation for Intrahepatic Cholangiocarcinoma.

BACKGROUND/AIM: The purpose of this study was to clarify the relationship between the desmoplastic reaction (DR) and clinicopathological features, and the prognosis using cases of resected intrahepatic cholangiocarcinoma (ICC). PATIENTS AND METHODS: Out of 54 cases that were preoperatively diagnosed with ICC and underwent resection at our department, 47 patients were included in this study. All sections were prepared from resected specimens and were microscopically observed following H&E staining. Stroma were evaluated at the advancing edge of the cancer and stratified into three DR types: mature (DR1), intermediate (DR2), and immature (DR3). RESULTS: DR was correlated to the serum levels of CA19-9, but not to the other tumor factors. In multivariate analysis, only DR and tumor size were determined as independent prognostic factors. CONCLUSION: Evaluation of DR for ICC may be useful for prognostic assessments.