Improved Pancreatic Adenocarcinoma Diagnosis in Jaundiced and Non-Jaundiced Pancreatic Adenocarcinoma Patients through the Combination of Routine Clinical Markers Associated to Pancreatic Adenocarcinoma Pathophysiology

Background

There is still no reliable biomarker for the diagnosis of pancreatic adenocarcinoma. Carbohydrate antigen 19–9 (CA 19–9) is a tumor marker only recommended for pancreatic adenocarcinoma follow-up. One of the clinical problems lies in distinguishing between this cancer and other benign pancreatic diseases such as chronic pancreatitis. In this study we will assess the value of panels of serum molecules related to pancreatic cancer physiopathology to determine whether alone or in combination could help to discriminate between these two pathologies.

Methods

CA 19–9, carcinoembryonic antigen (CEA), C-reactive protein, albumin, insulin growth factor-1 (IGF-1) and IGF binding protein-3 were measured using routine clinical analyzers in a cohort of 47 pancreatic adenocarcinoma, 20 chronic pancreatitis and 15 healthy controls.

Results

The combination of CA 19–9, IGF-1 and albumin resulted in a combined area under the curve (AUC) of 0.959 with 93.6% sensitivity and 95% specificity, much higher than CA 19–9 alone. An algorithm was defined to classify the patients as chronic pancreatitis or pancreatic cancer with the above specificity and sensitivity. In an independent validation group of 20 pancreatic adenocarcinoma and 13 chronic pancreatitis patients, the combination of the four molecules classified correctly all pancreatic adenocarcinoma and 12 out of 13 chronic pancreatitis patients.

Conclusions

Although this panel of markers should be validated in larger cohorts, the high sensitivity and specificity values and the convenience to measure these parameters in clinical laboratories shows great promise for improving pancreatic adenocarcinoma diagnosis.     

Prediagnostic Serum Biomarkers as Early Detection Tools for Pancreatic Cancer in a Large Prospective Cohort Study

BackgroundThe clinical management of pancreatic cancer is severely hampered by the absence of effective screening tools.MethodsSixty-seven biomarkers were evaluated in prediagnostic sera obtained from cases of pancreatic cancer enrolled in the Prostate, Lung, Colorectal, and Ovarian Cancer Screening Trial (PLCO).ResultsThe panel of CA 19-9, OPN, and OPG, identified in a prior retrospective study, was not effective. CA 19-9, CEA, NSE, bHCG, CEACAM1 and PRL were significantly altered in sera obtained from cases greater than 1 year prior to diagnosis. Levels of CA 19-9, CA 125, CEA, PRL, and IL-8 were negatively associated with time to diagnosis. A training/validation study using alternate halves of the PLCO set failed to identify a biomarker panel with significantly improved performance over CA 19-9 alone. When the entire PLCO set was used for training at a specificity (SP) of 95%, a panel of CA 19-9, CEA, and Cyfra 21-1 provided significantly elevated sensitivity (SN) levels of 32.4% and 29.7% in samples collected <1 and >1 year prior to diagnosis, respectively, compared to SN levels of 25.7% and 17.2% for CA 19-9 alone.ConclusionsMost biomarkers identified in previously conducted case/control studies are ineffective in prediagnostic samples, however several biomarkers were identified as significantly altered up to 35 months prior to diagnosis. Two newly derived biomarker combinations offered advantage over CA 19-9 alone in terms of SN, particularly in samples collected >1 year prior to diagnosis. However, the efficacy of biomarker-based tools remains limited at present. Several biomarkers demonstrated significant velocity related to time to diagnosis, an observation which may offer considerable potential for enhancements in early detection.     

Enhanced discrimination of malignant from benign pancreatic disease by measuring the CA 19-9 antigen on specific protein carriers

The CA 19-9 assay detects a carbohydrate antigen on multiple protein carriers, some of which may be preferential carriers of the antigen in cancer. We tested the hypothesis that the measurement of the CA 19-9 antigen on individual proteins could improve performance over the standard CA 19-9 assay. We used antibody arrays to measure the levels of the CA 19-9 antigen on multiple proteins in serum or plasma samples from patients with pancreatic adenocarcinoma or pancreatitis. Sample sets from three different institutions were examined, comprising 531 individual samples. The measurement of the CA 19-9 antigen on any individual protein did not improve upon the performance of the standard CA 19-9 assay (82% sensitivity at 75% specificity for early-stage cancer), owing to diversity among patients in their CA 19-9 protein carriers. However, a subset of cancer patients with no elevation in the standard CA 19-9 assay showed elevations of the CA 19-9 antigen specifically on the proteins MUC5AC or MUC16 in all sample sets. By combining measurements of the standard CA 19-9 assay with detection of CA 19-9 on MUC5AC and MUC16, the sensitivity of cancer detection was improved relative to CA 19-9 alone in each sample set, achieving 67-80% sensitivity at 98% specificity. This finding demonstrates the value of measuring glycans on specific proteins for improving biomarker performance. Diagnostic tests with improved sensitivity for detecting pancreatic cancer could have important applications for improving the treatment and management of patients suffering from this disease.     

A combination of urinary biomarker panel and PancRISK score for earlier detection of pancreatic cancer: A case–control study

BackgroundPancreatic ductal adenocarcinoma (PDAC) is one of the deadliest cancers, with around 9% of patients surviving >5 years. Asymptomatic in its initial stages, PDAC is mostly diagnosed late, when already a locally advanced or metastatic disease, as there are no useful biomarkers for detection in its early stages, when surgery can be curative. We have previously described a promising biomarker panel (LYVE1, REG1A, and TFF1) for earlier detection of PDAC in urine. Here, we aimed to establish the accuracy of an improved panel, including REG1B instead of REG1A, and an algorithm for data interpretation, the PancRISK score, in additional retrospectively collected urine specimens. We also assessed the complementarity of this panel with CA19-9 and explored the daily variation and stability of the biomarkers and their performance in common urinary tract cancers.Methods and findingsClinical specimens were obtained from multiple centres: Barts Pancreas Tissue Bank, University College London, University of Liverpool, Spanish National Cancer Research Center, Cambridge University Hospital, and University of Belgrade. The biomarker panel was assayed on 590 urine specimens: 183 control samples, 208 benign hepatobiliary disease samples (of which 119 were chronic pancreatitis), and 199 PDAC samples (102 stage I–II and 97 stage III–IV); 50.7% were from female individuals. PDAC samples were collected from patients before treatment. The samples were assayed using commercially available ELISAs. Statistical analyses were performed using non-parametric Kruskal–Wallis tests adjusted for multiple comparisons, and multiple logistic regression. Training and validation datasets for controls and PDAC samples were obtained after random division of the whole available dataset in a 1:1 ratio. The substitution of REG1A with REG1B enhanced the performance of the panel to detect resectable PDAC. In a comparison of controls and PDAC stage I–II samples, the areas under the receiver operating characteristic curve (AUCs) increased from 0.900 (95% CI 0.843–0.957) and 0.926 (95% CI 0.843–1.000) in the training (50% of the dataset) and validation sets, respectively, to 0.936 in both the training (95% CI 0.903–0.969) and the validation (95% CI 0.888–0.984) datasets for the new panel including REG1B. This improved panel showed both sensitivity (SN) and specificity (SP) to be >85%. Plasma CA19-9 enhanced the performance of this panel in discriminating PDAC I–II patients from controls, with AUC = 0.992 (95% CI 0.983–1.000), SN = 0.963 (95% CI 0.913–1.000), and SP = 0.967 (95% CI 0.924–1.000). We demonstrate that the biomarkers do not show significant daily variation, and that they are stable for up to 5 days at room temperature. The main limitation of our study is the low number of stage I–IIA PDAC samples (n = 27) and lack of samples from individuals with hereditary predisposition to PDAC, for which specimens collected from control individuals were used as a proxy.ConclusionsWe have successfully validated our urinary biomarker panel, which was improved by substituting REG1A with REG1B. At a pre-selected cutoff of >80% SN and SP for the affiliated PancRISK score, we demonstrate a clinically applicable risk stratification tool with a binary output for risk of developing PDAC (‘elevated’ or ‘normal’). PancRISK provides a step towards precision surveillance for PDAC patients, which we will test in a prospective clinical study, UroPanc.

Silvana Debernardi and colleagues establish a clinical risk score and a biomarker panel for early detection of pancreatic cancer.     

Diverse monoclonal antibodies against the CA 19-9 antigen show variation in binding specificity with consequences for clinical interpretation

The CA 19-9 antigen is currently the best individual marker for the detection of pancreatic cancer. In order to optimize the CA 19-9 assay and to develop approaches to further improve cancer detection, it is important to understand the specificity differences between CA 19-9 antibodies and the consequential affect on biomarker performance. Antibody arrays enabled multiplexed comparisons between five different CA 19-9 antibodies used in the analysis of plasma samples from pancreatic cancer patients and controls. Major differences were observed between antibodies in their detection of particular patient samples. Glycan array analysis revealed that certain antibodies were highly specific for the canonical CA 19-9 epitope, sialyl-Lewis A, while others bound sialyl-Lewis A in addition to a related structure called sialyl-Lewis C and modification with Nue5Gc. In a much larger patient cohort, we confirmed the binding of sialyl-Lewis C glycan by one of the antibodies and showed that the broader specificity led to the detection of an increased number of cancer patients without increasing detection of pancreatitis patient samples. This work demonstrates that variation between antibody specificity for cancer-associated glycans can have significant implications for biomarker performance and highlights the value of characterizing and detecting the range of glycan structures that are elevated in cancer.     

Laminin,gamma 2 (LAMC2): A Promising New Putative Pancreatic Cancer Biomarker Identified by Proteomic Analysis of Pancreatic Adenocarcinoma Tissues

In pancreatic cancer, the incidence and mortality curves coincide. One major reason for this high mortality rate in pancreatic ductal adenocarcinoma (PDAC) patients is the dearth of effective diagnostic, prognostic, and disease-monitoring biomarkers. Unfortunately, existing tumor markers, as well as current imaging modalities, are not sufficiently sensitive and/or specific for early-stage diagnosis. There is, therefore, an urgent need for improved serum markers of the disease. Herein, we performed Orbitrap® mass spectrometry proteomic analysis of four PDAC tissues and their adjacent benign tissues and identified a total of 2190 nonredundant proteins. Sixteen promising candidates were selected for further scrutiny using a systematic scoring algorithm. Our preliminary serum verification of the top four candidates (DSP, LAMC2, GP73, and DSG2) in 20 patients diagnosed with pancreatic cancer and 20 with benign pancreatic cysts, showed a significant (p < 0.05) elevation of LAMC2 in pancreatic cancer serum. Extensive validation of LAMC2 in healthy, benign, and PDAC sera from geographically diverse cohorts (n = 425) (Japan, Europe, and USA) demonstrated a significant increase in levels in early-stage PDAC compared with benign diseases. The sensitivity of LAMC2 was comparable to CA19.9 in all data sets, with an AUC value greater than 0.85 in discriminating healthy patients from early-stage PDAC patients. LAMC2 exhibited diagnostic complementarity with CA19.9 by showing significant (p < 0.001 in two out of three cohorts) elevation in PDAC patients with clinically low CA19.9 levels.Pancreatic ductal adenocarcinoma (PDAC)¹ is one of the most devastating cancers and the fourth leading cause of cancer-related deaths in North America (1). Ninety-five percent of patients will not survive beyond five years; this high mortality rate is primarily attributed to the lack of effective diagnostic techniques and treatment regimens. The hallmark features of pancreatic cancer (PC) are late presentation and aggressive metastatic progression (2, 3). The National Cancer Institute statistics estimate that approximately $1.9 billion is being spent in the United States alone each year on PC diagnosis and treatment. PDAC is classified into resectable (∼10–20%), locally advanced unresectable (∼30–40%), and metastatic (∼50%) (3). PDAC diagnosed at resectable stage can possibly be cured with complete surgical removal. This could improve the survival rates and considerably lower treatment costs. It is projected that 20–40% of patients with resectable PDAC survive more than five years after complete surgical removal, highlighting the importance of early-stage diagnosis. Unfortunately, carbohydrate antigen 19–9 (CA19.9), the current standard serum tumor marker for PDAC, has certain limitations as an early detection biomarker (its sensitivity for small tumors {<3 cm} is ∼50% and it is significantly elevated in many benign conditions (e.g. biliary obstruction, hepatic cirrhosis, chronic pancreatitis)) (4, 5). In light of the scarcity of other, more reliable markers, CA19.9 is currently used in the clinic as a prognostic and surveillance marker. Undoubtedly, the need for a more reliable consistent biomarker (or biomarker panel) for early PDAC diagnosis remains unmet. In pursuit of novel PDAC biomarker candidates, we have previously delineated the proteomes of malignant pancreatic ascitic fluids, pools of pancreatic juice, and pancreatic cancer cell lines (BxPC3, CAPAN, CFPAC1, MIA-Paca2, PANC1, and SU.86.86). We identified a panel of five potential candidate biomarkers, which, in combination, slightly outperformed CA19.9 in a pilot verification study (40 individuals; 20 healthy, and 20 PDAC) (6).From a different perspective, in the current study, we deployed a comparative quantitative tissue proteomic methodology to compare the proteome of malignant pancreatic tissues with that of their adjacent normal counterparts. A total of 2190 nonredundant proteins were identified, which were further scrutinized using a systematic scoring algorithm based on their quantified cancer-versus-normal ratios, on their identification in malignant pancreatic ascites fluid, on their cancer-specific nature, and on their tissue-expression profiles. Our analysis resulted in sixteen promising candidate biomarkers, which fulfilled our criteria and selected for further validation studies. In a multistep validation approach, the selected candidates were first verified in serum samples obtained from 20 patients with benign pancreatic diseases and 20 patients with pancreatic cancer, using commercially available ELISA kits. The best candidate (LAMC2) was further tested in three geographically diverse cohorts from Germany, Japan, and the US composed of 435 serum samples from healthy, benign, and early and late stage cancer patients. Our approach brought to light a previously unknown, promising PDAC candidate biomarker, LAMC2.     

Protein alterations associated with pancreatic cancer and chronic pancreatitis found in human plasma using global quantitative proteomics profiling

Pancreatic cancer is a lethal disease that is difficult to diagnose at early stages when curable treatments are effective. Biomarkers that can improve current pancreatic cancer detection would have great value in improving patient management and survival rate. A large scale quantitative proteomics study was performed to search for the plasma protein alterations associated with pancreatic cancer. The enormous complexity of the plasma proteome and the vast dynamic range of protein concentration therein present major challenges for quantitative global profiling of plasma. To address these challenges, multidimensional fractionation at both protein and peptide levels was applied to enhance the depth of proteomics analysis. Employing stringent criteria, more than 1300 proteins total were identified in plasma across 8-orders of magnitude in protein concentration. Differential proteins associated with pancreatic cancer were identified, and their relationship with the proteome of pancreatic tissue and pancreatic juice from our previous studies was discussed. A subgroup of differentially expressed proteins was selected for biomarker testing using an independent cohort of plasma and serum samples from well-diagnosed patients with pancreatic cancer, chronic pancreatitis, and nonpancreatic disease controls. Using ELISA methodology, the performance of each of these protein candidates was benchmarked against CA19-9, the current gold standard for a pancreatic cancer blood test. A composite marker of TIMP1 and ICAM1 demonstrate significantly better performance than CA19-9 in distinguishing pancreatic cancer from the nonpancreatic disease controls and chronic pancreatitis controls. In addition, protein AZGP1 was identified as a biomarker candidate for chronic pancreatitis. The discovery and technical challenges associated with plasma-based quantitative proteomics are discussed and may benefit the development of plasma proteomics technology in general. The protein candidates identified in this study provide a biomarker candidate pool for future investigations.     

Can IL-2R alpha be a valuable marker along with CA 19-9 in the diagnosis of chronic pancreatitis and pancreatic cancer?

BACKGROUND: Pancreatic cancer is characterized initially by non-specific abdominal symptoms followed by rapid tumor progression. Although chronic pancreatitis is a benign disorder, it can be one of the causative factors of pancreatic cancer. The level of the tumor marker carbohydrate antigen 19-9 (CA 19-9) in pancreatic cancer does not correlate with the stage of the neoplasm. Soluble interleukin 2 receptor (sIL-2R) is a cytokine that shows increased levels during some inflammatory processes and malignant disorders. AIM: Our aim in this study was to investigate whether sIL-2Ralpha levels can be used in association with CA 19-9 in the early diagnosis of pancreatic cancer and chronic pancreatitis. PATIENTS: Serum samples were obtained from the blood of 21 pancreatic cancer patients without distant metastasis who were deemed inoperable, 16 chronic pancreatitis patients and 20 normal volunteers. RESULTS: We did not find any significant differences in CA 19-9 levels between normal controls and patients with chronic pancreatitis. There was a significant difference in the levels between the control group and the pancreatic cancer group (p = 0.003) and between patients with chronic pancreatitis and those with pancreatic cancer (p = 0.004). Although there was no significant difference in sIL-2Ralpha levels between the control group and the patient groups, we found a slight correlation between sIL-2Ralpha and CA 19-9 levels in the pancreatic cancer group (p = 0.003, r = 0.623) and a more marked correlation in the chronic pancreatitis group (p < 0.01, r = 0.751). CONCLUSION: According to our results, sIL-2Ralpha alone is not a good candidate marker in the diagnosis of pancreatic cancer; it can, however, be used in association with CA 19-9 for this purpose.     

血清miR-92a在胰腺癌诊断及预后评估中的临床意义研究

目的:探讨血清miR-92a在胰腺癌诊断和预后分析中的价值,为胰腺癌早诊断以及预后评估提供潜在的分子标志物。方法:回顾性分析我院及重庆医科大学附属第一、第二医院2014年8月~2016年12月收治的30例胰腺癌未转移患者、30例胰腺癌转移患者和30例慢性胰腺炎患者的临床资料,另选择同期在我院进行健康体检的30例健康人作为健康组。收集血清,应用定量PCR法检测各组血清miR-92a的表达水平,利用化学发光法检测各组血清中的糖蛋白抗原19-9(CA-19-9)含量。以ROC分析比较血清miR-92a与CA 19-9在胰腺癌诊断中的特异度、敏感性。结果:胰腺癌未转移组患者和胰腺癌转移组患者血清中miR-92a水平显著高于健康组和慢性胰腺炎组(P0.05),胰腺癌转移组患者血清中miR-92a水平显著高于胰腺癌未转移组患者(P0.05)。胰腺癌未转移组患者和胰腺癌转移组患者血清中CA19-9水平显著高于健康组和慢性胰腺炎组(P0.05)。miR-92a诊断胰腺癌的敏感度高于CA19-9和miR-92a+CA 19-9,而miR-92a+CA 19-9诊断胰腺癌的特异度显著高于miR-92a和CA19-9(P0.05),且有较高的胰腺癌转移预测应用价值。结论:血清miR-92a联合CA 19-9检测能够诊断胰腺癌,具有良好的敏感度和特异度,miR-92a还具有较好的胰腺癌转移预测价值,可作为胰腺癌早期无创筛查方法加以应用。     

IPO-38 is identified as a novel serum biomarker of gastric cancer based on clinical proteomics technology

Gastric cancer is one of the most common malignancies in China. So far, there are few reliable serum biomarkers for diagnosis. The available biomarkers of CEA, CA19-9 and CA72-4 are not sufficiently sensitive and specific for gastric cancer. In this study, a high density antibody microarray was used for identifying new biomarkers from serum samples of gastric cancer. Serum samples from colorectal cancer, pancreatic cancer, hepatocellular cancer, and breast cancer were also screened for comparative study. As result, some candidate biomarkers were identified. IPO-38, an up-regulated serum protein in gastric cancer was selected for subsequent validation including serum IPO-38 expression by ELISA and IPO-38 protein expression by immunohistochemistry. The immunoprecipitation by IPO-38 for gastric cancer cell line and MALDI-TOF/TOF mass spectrometer suggested that pull-down of IPO-38 belongs to H2B histone, which was supported by co-localization study of laser scanning confocal microscope. A follow-up study showed that the survival rate of IPO-38 negative group was better than that in IPO-38 positive group. The study first clarified the property of IPO-38 proliferating marker, and proposed that IPO-38 protein is a promising biomarker both for diagnosis and for predicting prognosis of gastric cancer.     

Roles of CA19-9 in pancreatic cancer: Biomarker,predictor and promoter

Carbohydrate antigen 19-9 (CA19-9) is the best validated biomarker and an indicator of aberrant glycosylation in pancreatic cancer. CA19-9 functions as a biomarker, predictor, and promoter in pancreatic cancer. As a biomarker, the sensitivity is approximately 80%, and the major challenges involve false positives in conditions of inflammation and nonpancreatic cancers and false negatives in Lewis-negative Individuals. Lewis antigen status should be determined when using CA19-9 as a biomarker. CA19-9 has screening potential when combined with symptoms and/or risk factors. As a predictor, CA19-9 could be used to assess stage, prognosis, resectability, recurrence, and therapeutic efficacy. Normal baseline levels of CA19-9 are associated with long-term survival. As a promoter, CA19-9 could be used to evaluate the biology of pancreatic cancer. CA19-9 can accelerate pancreatic cancer progression by glycosylating proteins, binding to E-selectin, strengthening angiogenesis, and mediating the immunological response. CA19-9 is an attractive therapeutic target for cancer, and strategies include therapeutic antibodies and vaccines, CA19-9-guided nanoparticles, and inhibition of CA19-9 biosynthesis.     

Diagnostic and Prognostic Impact of Circulating YKL-40, IL-6, and CA 19.9 in Patients with Pancreatic Cancer

Purpose

We tested the hypothesis that high plasma YKL-40 and IL-6 associate with pancreatic cancer and short overall survival.

Patients and Methods

In all, 559 patients with pancreatic cancer from prospective biomarker studies from Denmark (n = 448) and Germany (n = 111) were studied. Plasma YKL-40 and IL-6 were determined by ELISAs and serum CA 19.9 by chemiluminescent immunometric assay.

Results

Odds ratios (ORs) for prediction of pancreatic cancer were significant for all biomarkers, with CA 19.9 having the highest AUC (CA 19.9: OR = 2.28, 95% CI 1.97 to 2.68, p<0.0001, AUC = 0.94; YKL-40: OR = 4.50, 3.99 to 5.08, p<0.0001, AUC = 0.87; IL-6: OR = 3.68, 3.08 to 4.44, p<0.0001, AUC = 0.87). Multivariate Cox analysis (YKL-40, IL-6, CA 19.9, age, stage, gender) in patients operated on showed that high preoperative IL-6 and CA 19.9 (dichotomized according to normal values) were independently associated with short overall survival (CA 19.9: HR = 2.51, 1.22–5.15, p = 0.013; IL-6: HR = 2.03, 1.11 to 3.70, p = 0.021). Multivariate Cox analysis of non-operable patients (Stage IIB-IV) showed that high pre-treatment levels of each biomarker were independently associated with short overall survival (YKL-40: HR = 1.30, 1.03 to 1.64, p = 0.029; IL-6: HR = 1.71, 1.33 to 2.20, p<0.0001; CA 19.9: HR = 1.54, 1.06 to 2.24, p = 0.022). Patients with preoperative elevation of both IL-6 and CA 19.9 had shorter overall survival (p<0.005) compared to patients with normal levels of both biomarkers (45% vs. 92% alive after 12 months).

Conclusions

Plasma YKL-40 and IL-6 had less diagnostic impact than CA 19.9. Combination of pretreatment YKL-40, IL-6, and CA 19.9 may have clinical value to identify pancreatic cancer patients with the poorest prognosis.     

Serum Protein Profile at Remission Can Accurately Assess Therapeutic Outcomes and Survival for Serous Ovarian Cancer

Background

Biomarkers play critical roles in early detection, diagnosis and monitoring of therapeutic outcome and recurrence of cancer. Previous biomarker research on ovarian cancer (OC) has mostly focused on the discovery and validation of diagnostic biomarkers. The primary purpose of this study is to identify serum biomarkers for prognosis and therapeutic outcomes of ovarian cancer.

Experimental Design

Forty serum proteins were analyzed in 70 serum samples from healthy controls (HC) and 101 serum samples from serous OC patients at three different disease phases: post diagnosis (PD), remission (RM) and recurrence (RC). The utility of serum proteins as OC biomarkers was evaluated using a variety of statistical methods including survival analysis.

Results

Ten serum proteins (PDGF-AB/BB, PDGF-AA, CRP, sFas, CA125, SAA, sTNFRII, sIL-6R, IGFBP6 and MDC) have individually good area-under-the-curve (AUC) values (AUC = 0.69–0.86) and more than 10 three-marker combinations have excellent AUC values (0.91–0.93) in distinguishing active cancer samples (PD & RC) from HC. The mean serum protein levels for RM samples are usually intermediate between HC and OC patients with active cancer (PD & RC). Most importantly, five proteins (sICAM1, RANTES, sgp130, sTNFR-II and sVCAM1) measured at remission can classify, individually and in combination, serous OC patients into two subsets with significantly different overall survival (best HR = 17, p<10⁻³).

Conclusion

We identified five serum proteins which, when measured at remission, can accurately predict the overall survival of serous OC patients, suggesting that they may be useful for monitoring the therapeutic outcomes for ovarian cancer.     

Quantitative proteomics analysis reveals that proteins differentially expressed in chronic pancreatitis are also frequently involved in pancreatic cancer

The effective treatment of pancreatic cancer relies on the diagnosis of the disease at an early stage, a difficult challenge. One major obstacle in the development of diagnostic biomarkers of early pancreatic cancer has been the dual expression of potential biomarkers in both chronic pancreatitis and cancer. To better understand the limitations of potential protein biomarkers, we used ICAT technology and tandem mass spectrometry-based proteomics to systematically study protein expression in chronic pancreatitis. Among the 116 differentially expressed proteins identified in chronic pancreatitis, most biological processes were responses to wounding and inflammation, a finding consistent with the underlining inflammation and tissue repair associated with chronic pancreatitis. Furthermore 40% of the differentially expressed proteins identified in chronic pancreatitis have been implicated previously in pancreatic cancer, suggesting some commonality in protein expression between these two diseases. Biological network analysis further identified c-MYC as a common prominent regulatory protein in pancreatic cancer and chronic pancreatitis. Lastly five proteins were selected for validation by Western blot and immunohistochemistry. Annexin A2 and insulin-like growth factor-binding protein 2 were overexpressed in cancer but not in chronic pancreatitis, making them promising biomarker candidates for pancreatic cancer. In addition, our study validated that cathepsin D, integrin beta1, and plasminogen were overexpressed in both pancreatic cancer and chronic pancreatitis. The positive involvement of these proteins in chronic pancreatitis and pancreatic cancer will potentially lower the specificity of these proteins as biomarker candidates for pancreatic cancer. Altogether our study provides some insights into the molecular events in chronic pancreatitis that may lead to diverse strategies for diagnosis and treatment of these diseases.     

Identification of blood-protein carriers of the CA 19-9 antigen and characterization of prevalence in pancreatic diseases

The current best serum marker for pancreatic cancer, CA 19-9, detects a carbohydrate antigen on multiple protein carriers. Better knowledge of the protein carriers of the CA 19-9 antigen in various disease states may lead to improved diagnostic tests. To identify proteins that carry the CA 19-9 antigen, we immunoprecipitated the CA 19-9 antigen from pooled sera and identified the associated proteins using MS. Among the high-confidence identifications, we confirmed the presence of the CA 19-9 antigen on Apolipoprotein B-100 by antibody arrays and Western blot and on kininogen, ARVCF, and Apolipoprotein E by antibody arrays. We characterized the frequency and levels of the CA 19-9 antigen on the four proteins across various patient groups (pancreatic cancer, pancreatitis, and healthy controls) using antibody arrays. Nearly, 10-25% of the subjects showed elevations of the antigen on each protein, but the elevations were not associated with disease state or total CA 19-9 levels. These results contribute to our knowledge of the carrier proteins of an important functional glycan and the rate at which the glycan is displayed. This work also demonstrates a strategy for using the complementary methods of MS and antibody microarrays to identify protein carriers of glycans and assess the diagnostic value of measuring glycans on individual proteins.     

Seminal Plasma as a Source of Prostate Cancer Peptide Biomarker Candidates for Detection of Indolent and Advanced Disease

Background

Extensive prostate specific antigen screening for prostate cancer generates a high number of unnecessary biopsies and over-treatment due to insufficient differentiation between indolent and aggressive tumours. We hypothesized that seminal plasma is a robust source of novel prostate cancer (PCa) biomarkers with the potential to improve primary diagnosis of and to distinguish advanced from indolent disease.

Methodology/Principal Findings

In an open-label case/control study 125 patients (70 PCa, 21 benign prostate hyperplasia, 25 chronic prostatitis, 9 healthy controls) were enrolled in 3 centres. Biomarker panels a) for PCa diagnosis (comparison of PCa patients versus benign controls) and b) for advanced disease (comparison of patients with post surgery Gleason score <7 versus Gleason score >7) were sought. Independent cohorts were used for proteomic biomarker discovery and testing the performance of the identified biomarker profiles. Seminal plasma was profiled using capillary electrophoresis mass spectrometry. Pre-analytical stability and analytical precision of the proteome analysis were determined. Support vector machine learning was used for classification. Stepwise application of two biomarker signatures with 21 and 5 biomarkers provided 83% sensitivity and 67% specificity for PCa detection in a test set of samples. A panel of 11 biomarkers for advanced disease discriminated between patients with Gleason score 7 and organ-confined (<pT3a) or advanced (≥pT3a) disease with 80% sensitivity and 82% specificity in a preliminary validation setting. Seminal profiles showed excellent pre-analytical stability. Eight biomarkers were identified as fragments of N-acetyllactosaminide beta-1,3-N-acetylglucosaminyltransferase, prostatic acid phosphatase, stabilin-2, GTPase IMAP family member 6, semenogelin-1 and -2. Restricted sample size was the major limitation of the study.

Conclusions/Significance

Seminal plasma represents a robust source of potential peptide makers for primary PCa diagnosis. Our findings warrant further prospective validation to confirm the diagnostic potential of identified seminal biomarker candidates.     

Serum CA19–9, cathepsin D,and matrix metalloproteinase‐7 as a diagnostic panel for pancreatic ductal adenocarcinoma

Pancreatic ductal adenocarcinoma (PDAC) accounts for 95% of pancreatic cancers. CA19‐9 is not widely used for screening PDAC due to its low sensitivity. Here, we studied the clinical usefulness of cathepsin D, matrix metalloproteinases (MMPs), and tissue inhibitors of MMPs (TIMPs) for screening patients with PDAC. A total of 248 patients with PDAC and 216 control subjects were recruited (109 PDAC patients and 70 controls in the training set and 139 PDAC patients and 146 controls in the validation set). We measured serum levels of cathepsin D, TIMPs (?1, ?3, and ?4), and MMPs (?1, ?7, ?8, and ?9) using Fluorokine MAP multiplex kits. The concentrations of cathepsin D and MMP‐7 were significantly higher in PDAC subjects than control subjects. In the training set, the diagnostic sensitivity and AUC of the panel of CA19‐9, cathepsin D, and MMP‐7 for PDAC were increased to 88% and 0.900, compared to 74% and 0.835 of CA19‐9 single marker at 80% specificity. The sensitivity using cut‐off value of biomarker panel was significantly increased in the validation set as well as training set. Our findings indicate that a serum biomarker panel consisting of CA19‐9, cathepsin D, and MMP‐7 may provide the most effective screening test currently feasible for PDAC.     

Serum apolipoproteins C-I and C-III are reduced in stomach cancer patients: results from MALDI-based peptidome and immuno-based clinical assays

Finding new peptide biomarkers for stomach cancer in human sera that can be implemented into a clinically practicable prediction method for monitoring of stomach cancer. We studied the serum peptidome from two different biorepositories. We first employed a C8-reverse phase liquid chromatography approach for sample purification, followed by mass-spectrometry analysis. These were applied onto serum samples from cancer-free controls and stomach cancer patients at various clinical stages. We then created a bioinformatics analysis pipeline and identified peptide signature discriminating stomach adenocarcinoma patients from cancer-free controls. Matrix Assisted Laser Desorption/Ionization-Time of Flight (MALDI-TOF) results from 103 samples revealed 9 signature peptides; with prediction accuracy of 89% in the training set and 88% in the validation set. Three of the discriminating peptides discovered were fragments of Apolipoproteins C-I and C-III (apoC-I and C-III); we further quantified their serum levels, as well as CA19-9 and CRP, employing quantitative commercial-clinical assays in 142 samples. ApoC-I and apoC-III quantitative results correlated with the MS results. We then employed apoB-100-normalized apoC-I and apoC-III, CA19-9 and CRP levels to generate rules set for stomach cancer prediction. For training, we used sera from one repository, and for validation, we used sera from the second repository. Prediction accuracies of 88.4% and 74.4% were obtained in the training and validation sets, respectively. Serum levels of apoC-I and apoC-III combined with other clinical parameters can serve as a basis for the formulation of a diagnostic score for stomach cancer patients.     

Autoantibodies against tumor-associated antigens in sputum as biomarkers for lung cancer

Tumor antigens (TAs) can initiate host immune responses and produce TA-associated autoantibody (TAAbs), potential cancer biomarkers. Sputum is directly generated from the upper and lower airways, and thus can be used as a surrogate sample for the diagnosis of lung cancer based on molecular analysis. To develop sputum TAAb biomarkers for the early detection of lung cancer, the leading cause of cancer death, we probed a protein microarray containing more than 9,000 antigens with sputum supernatants of a discovery set of 30 lung cancer patients and 30 cancer-free smokers. Twenty-eight TAs with higher reactivity in sputum of lung cancer cases vs. controls were identified. The diagnostic significance of TAAbs against the TAs was determined by enzyme-linked immunosorbent assays (ELISAs) in sputum of the discovery set and additional 166 lung cancer patients and 213 cancer-free smokers (validation set). Three sputum TAAbs against DDX6, ENO1, and 14–3-3ζ were developed as a biomarker panel with 81% sensitivity and 83% specificity for diagnosis of lung cancer, regardless of stages, locations, and histological types of lung tumors. This study provides the first evidence that sputum TAAbs could be used as biomarkers for the early detection of lung cancer.     

HOTAIRM1 as a potential biomarker for diagnosis of colorectal cancer functions the role in the tumour suppressor

Recent studies have revealed many different long noncoding RNAs (lncRNA), however, the investigation for their function and clinical value as tumour biomarkers has scarcely begun. Here, we found that expression of HOTAIRM1 was reduced in colorectal cancer (CRC) tissues compared with matched normal tissues, and plasma HOTAIRM1 levels in CRC patients were less than in controls. The cut‐off point was chosen as 0.003 with a sensitivity of 64.00% and a specificity of 76.50% in the validation set. The performance of HOTAIRM1 was highly comparable to carcinoembryonic antigen (CEA), and better than CA19‐9 and CA125. The combined assay of HOTAIRM1 and CEA raised the sensitivity and specificity to 84.00%. HOTAIRM1 knockdown resulted in obvious changes in expression of the cell proliferation related to genes and promoted cell proliferation. HOTAIRM1 plays a role of tumour suppressor in CRC; Down‐regulation of HOTAIRM1 can serve as a biomarker for CRC, and combined HOTAIRM1 and CEA assay might provide a promising diagnosis for CRC.