Evaluation of Serum Glycoprotein Biomarker Candidates for Detection of Esophageal Adenocarcinoma and Surveillance of Barrett's Esophagus

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Highlights

• •C9, GSN, PON1, and PON3 validated as serum biomarker candidates of EAC.
• •Multimarker panel with AUROC of 0.93 to aid current endoscopy surveillance of BE.
• •Induction of tissue C9 in BE, dysplastic BE and EAC.
• •Alteration of complement pathway glycoproteins during BE-EAC pathogenesis.

     

Serum Fucosylated Haptoglobin as a Novel Diagnostic Biomarker for Predicting Hepatocyte Ballooning and Nonalcoholic Steatohepatitis

Nonalcoholic fatty liver disease (NAFLD) is a growing medical problem around the world. NAFLD patients with nonalcoholic steatohepatitis (NASH) can develop cirrhosis and hepatocellular carcinoma. The ability to distinguish NASH from simple steatosis would be of great clinical significance. Ballooning hepatocytes are characteristic of typical pathological NASH; here, the polarized secretion of proteins is disrupted due to destruction of the cytoskeleton. We previously reported that fucosylated glycoproteins are secreted into bile, but not into sera in normal liver. Therefore, we hypothesized that the fucosylation-based sorting machinery would be disrupted in ballooning hepatocytes, and serum fucosylated glycoproteins would increase in NASH patients. To confirm our hypothesis, we evaluated serum fucosylated haptoglobin (Fuc-Hpt) levels in biopsy-proven NAFLD patients (n = 126) using a lectin-antibody ELISA kit. Fuc-Hpt levels were significantly increased in NASH patients compared with non-NASH (NAFLD patients without NASH) patients. Interestingly, Fuc-Hpt levels showed a significant stepwise increase with increasing hepatocyte ballooning scores. Multiple logistic regression analysis showed that Fuc-Hpt levels were independent and significant determinants of the presence of ballooning hepatocytes. Moreover, Fuc-Hpt levels were useful in monitoring liver fibrosis staging. Next, to investigate the significance of serum Fuc-Hpt in a larger population, we measured Fuc-Hpt levels in ultrasound-diagnosed NAFLD subjects (n = 870) who received a medical health checkup. To evaluate NAFLD disease severity, we used the FIB-4 index (based on age, serum AST and ALT levels, and platelet counts). Fuc-Hpt levels increased stepwise with increasing FIB-4 index.

Conclusion

Measurement of serum Fuc-Hpt levels can distinguish NASH from non-NASH patients, and predict the presence of ballooning hepatocytes in NAFLD patients with sufficient accuracy. These results support the potential usefulness of measuring Fuc-Hpt levels in clinical practice.     

Follistatin Is a Novel Biomarker for Lung Adenocarcinoma in Humans

Background

Follistatin (FST), a single chain glycoprotein, is originally isolated from follicular fluid of ovary. Previous studies have revealed that serum FST served as a biomarker for pregnancy and ovarian mucinous tumor. However, whether FST can serve as a biomarker for diagnosis in lung adenocarcinoma of humans remains unclear.

Methods and Results

The study population consisted of 80 patients with lung adenocarcinoma, 40 patients with ovarian adenocarcinoma and 80 healthy subjects. Serum FST levels in patients and healthy subjects were measured using ELISA. The results showed that the positive ratio of serum FST levels was 51.3% (41/80), which was comparable to the sensitivity of FST in 40 patients with ovarian adenocarcinoma (60%, 24/40) using the 95th confidence interval for the healthy subject group as the cut-off value. FST expressions in lung adenocarcinoma were examined by immunohistochemical staining, we found that lung adenocarcinoma could produce FST and there was positive correlation between the level of FST expression and the differential degree of lung adenocarcinoma. Furthermore, the results showed that primary cultured lung adenocarcinoma cells could secrete FST, while cells derived from non-tumor lung tissues almost did not produce FST. In addition, the results of CCK8 assay and flow cytometry showed that using anti-FST monoclonal antibody to neutralize endogenous FST significantly augmented activin A-induced lung adenocarcinoma cells apoptosis.

Conclusions

These data indicate that lung adenocarcinoma cells can secret FST into serum, which may be beneficial to the survival of adenocarcinoma cells by neutralizing activin A action. Thus, FST can serve as a promising biomarker for diagnosis of lung adenocarcinoma and a useful biotherapy target for lung adenocarcinoma.     

Identification of Novel Biomarker Candidates for the Immunohistochemical Diagnosis of Cholangiocellular Carcinoma

The aim of this study was the identification of novel biomarker candidates for the diagnosis of cholangiocellular carcinoma (CCC) and its immunohistochemical differentiation from benign liver and bile duct cells. CCC is a primary cancer that arises from the epithelial cells of bile ducts and is characterized by high mortality rates due to its late clinical presentation and limited treatment options. Tumorous tissue and adjacent non-tumorous liver tissue from eight CCC patients were analyzed by means of two-dimensional differential in-gel electrophoresis and mass-spectrometry-based label-free proteomics. After data analysis and statistical evaluation of the proteins found to be differentially regulated between the two experimental groups (fold change ≥ 1.5; p value ≤ 0.05), 14 candidate proteins were chosen for determination of the cell-type-specific expression profile via immunohistochemistry in a cohort of 14 patients. This confirmed the significant up-regulation of serpin H1, 14-3-3 protein sigma, and stress-induced phosphoprotein 1 in tumorous cholangiocytes relative to normal hepatocytes and non-tumorous cholangiocytes, whereas some proteins were detectable specifically in hepatocytes. Because stress-induced phosphoprotein 1 exhibited both sensitivity and specificity of 100%, an immunohistochemical verification examining tissue sections of 60 CCC patients was performed. This resulted in a specificity of 98% and a sensitivity of 64%. We therefore conclude that this protein should be considered as a potential diagnostic biomarker for CCC in an immunohistochemical application, possibly in combination with other candidates from this study in the form of a biomarker panel. This could improve the differential diagnosis of CCC and benign bile duct diseases, as well as metastatic malignancies in the liver.Cholangiocellular carcinoma (CCC)¹ is a malignant neoplasm that arises from the cholangiocytes, the epithelial cells lining the bile ducts. The tumors, consisting of a significant amount of fibrous stroma, are classified as intrahepatic, extrahepatic, or hilar according to their anatomic location. Most common are the Klatskin tumors, originating from the confluence of the right and left hepatic ducts (1). Compared with other types of cancer, CCC is a relatively rare disease, accounting for about 3% of all gastrointestinal malignancies (2). However, its incidence is increasing, and as a result of poor patient outcomes it has overtaken hepatocellular carcinoma as the main cause of death from a primary hepatobiliary tumor (3). Reasons for the high mortality rate (5-year survival rate of about 5%) (4) are the difficult diagnosis and limited treatment options. At present, extensive surgical resection of the extrahepatic bile ducts and parts of the liver or liver transplantation remain the only potentially curative treatment options, although most patients are considered inoperable at the time of diagnosis (5).In general, the diagnosis of CCC is made based on histomorphological evaluation of core biopsies or cytological specimens. However, distinction between CCC and benign diseases such as reactive bile ductules or bile duct adenomas can be challenging when based on conventional histology alone. Additionally, it may be difficult to distinguish CCC from metastatic adenocarcinoma in the liver, especially when it originates from the pancreas like pancreatic ductal adenocarcinoma. Therefore, specific immunohistochemical tissue markers for CCC would be highly beneficial for further validation of the diagnosis. In routine immunohistochemical diagnosis of CCC, so far, the detection of p53 (a product of a tumor suppressor gene) has proven useful, although its application is limited because of low sensitivity (6). The cytokeratins Ck7, Ck8, Ck18, and Ck19 have been reported to have sensitivities of between 80% and 97% for CCC cells, but at low specificities and a similar expression as in non-tumorous cholangiocytes (7). In addition, the tumor marker carcinoembryonic antigen, which is a commonly applied serum marker, has been used for immunohistochemical staining of CCC tissue. Although this was reported to be positive in 100% of the tested CCC sections, it also was immunoreactive in 60% of hepatocellular carcinomas (8). Recently, it has been shown that the polycomb group protein EZH2 may be useful for differential diagnosis of cholangiolocellular carcinoma (a subtype of CCC), bile duct adenomas, and ductular reaction. This, however, applies only to this certain type of CCC (9). Establishing reliable immunohistochemical tumor markers specific for CCC therefore remains a challenge.Several proteomic studies using different sample types and various techniques have been performed in order to identify CCC-specific proteins. The analysis of CCC cell lines, for example, has led to the identification of potential diagnostic and prognostic biomarker candidates (10–12). In addition, cell lines have been used to discover proteins predictive of the response to chemotherapy (13). Because results from cell culture experiments do not always reflect the actual conditions in the tumor, the use of patient samples can be advantageous. The most appropriate source of tumor-specific signals is tumor tissue, which in the past has been analyzed via two-dimensional electrophoresis (14) and mass-spectrometry-based proteomic approaches such as histology-directed MALDI-TOF-MS (15), Surface-enhanced laser desorption/ionization (SELDI) TOF-MS (16), or LC-MS/MS (17). So far, however, none of the potential biomarkers have been successfully implemented into clinical routine.Recently, we demonstrated that the application of two complementary techniques, two-dimensional differential in-gel electrophoresis (2D-DIGE) and mass-spectrometry-based label-free LC-MS/MS, is an auspicious tactic for the discovery of novel biomarker candidates in hepatocellular carcinoma tissue (18). Here, we applied this well-established workflow as the initial step for the discovery of tissue markers that improve the differential diagnosis of intrahepatic CCC from benign bile duct diseases. In these experiments, CCC tumor tissue was compared with non-tumorous liver tissue (n = 8). Because this does not allow discrimination among different cell types such as hepatocytes, cholangiocytes, and tumor cells, an immunohistochemical determination of the cell-type-specific expression was subsequently performed for the most promising biomarker candidates. Stress-induced phosphoprotein 1, the protein showing the greatest specificity and sensitivity for CCC tumor cells, was verified as a suitable biomarker candidate for CCC in a larger patient cohort (n = 60).     

基于支持向量机的大肠黏液腺癌术前血清蛋白质标志物检测及分析

应用表面加强激光解吸电离-飞行时间质谱(SELDI-TOF-MS)技术和CM10蛋白质芯片从大肠黏液腺癌和非黏液腺癌患者中成功地筛选出了大肠黏液腺癌患者血清特异性相关蛋白.应用美国CipherGen公司CM10蛋白质芯片和PBSⅡ型蛋白质芯片阅读仪检测53例大肠癌患者(黏液腺癌12例,非黏液腺癌41例)患者血清蛋白质指纹图谱.采用ZUCI-Protein Chip Data Analyze System分析软件包进行分析,离散小波去噪音,结合支持向量机筛选肿瘤标志物,建立大肠黏液腺癌的术前诊断模型.12例大肠黏液腺癌患者与41例大肠非黏液腺癌患者的血清蛋白质有12个蛋白质峰强度有显著差异.其中质荷比为24 297和23 434 m/z处的蛋白质峰强度统计学P值分别为0.0067和0.0092,差异有极显著统计学意义.支持向量机筛选出24 297、3 322、3 822和4 353 m/z蛋白质峰作为生物标志物进行检测和预测准确率,其中12例大肠黏液腺癌患者中有10例患者被正确识别,41例大肠癌非黏液腺癌患者中有39例被正确识别,准确率为92.45%(49/53).该方法可以较好地应用于区别大肠黏液腺癌和非黏液腺癌,进行术前病理鉴别,指导进行大肠黏液腺癌的手术和综合治疗.     

Serum N-Glycans: A New Diagnostic Biomarker for Light Chain Multiple Myeloma

The aim of this study was to evaluate the diagnostic and differential diagnostic power of serum N-glycans for light chain multiple myeloma (LCMM). A total of 167 cases of subjects, including 42 LCMM, 42 IgG myeloma, 41 IgA myeloma, and 42 healthy controls were recruited in this study. DNA sequencer-assisted fluorophore-assisted capillary electrophoresis (DSA-FACE) was applied to determine the quantitive abundance of serum N-glycans. The core fucosylated, bisecting and sialylated modifications were analyzed in serum of LCMM patients (n=20) and healthy controls (n=20) randomly selected from the same cohort by lectin blot. Moreover, serum sialic acid (SA) level was measured by enzymatic method. We found two N-glycan structures (NG1A2F, Peak3; NA2FB, Peak7) showed the optimum diagnostic efficacy with area under the ROC curve (AUC) 0.939 and 0.940 between LCMM and healthy control. The sensitivity and specificity of Peak3 were 88.1% and 92.9%, while Peak7 were 92.9% and 97.6%, respectively. The abundance of Peak3 could differentiate LCMM from IgG myeloma with AUC 0.899, sensitivity 100% and specificity 76.2%, and Peak7 could be used to differentiate LCMM from IgA myeloma with AUC 0.922, sensitivity 92.9% and specificity 82.9%. Serum SA level was significantly higher in patients with LCMM than that in healthy controls. Moreover, the decreased core fucosylation, bisecting and increased sialylation characters of serum glycoproteins were observed in patients with LCMM. We concluded that serum N-glycan could provide a simple, reliable and non-invasive biomarker for LCMM diagnosis and abnormal glycosylation might imply a new potential therapeutic target in LCMM.     

Peptidomic Analysis of Maternal Serum to Identify Biomarker Candidates for Prenatal Diagnosis of Tetralogy of Fallot

Tetralogy of Fallot (TOF) is the most common form of cyanotic congenital heart disease. To identify endogenous peptides possibly involved in the progression of TOF, we performed comparative peptidomic profiling of maternal serum between normal fetuses and fetuses suffering from TOF. A total of 278 differentially expressed peptides, including 94 over‐expressed peptides and 184 under‐expressed peptides, originating from 227 protein precursors were identified by liquid chromatography/mass spectrometry (LC/MS) in maternal serum of fetuses with TOF compared to normal controls. Further, ingenuity pathway analysis (IPA) was used to identify putative roles for these peptides in cardiovascular development. Two peptides were derived from functional domains of proteins involved in heart development and associated with TOF; these may represent candidate bioactive peptides involved in TOF. These peptides may be related to the pathologic changes in the heart associated with TOF, and may be useful as novel biomarkers for prenatal diagnosis of TOF. J. Cell. Biochem. 119: 468–477, 2018. © 2017 Wiley Periodicals, Inc.     

Computational Biomarker Pipeline from Discovery to Clinical Implementation: Plasma Proteomic Biomarkers for Cardiac Transplantation

Recent technical advances in the field of quantitative proteomics have stimulated a large number of biomarker discovery studies of various diseases, providing avenues for new treatments and diagnostics. However, inherent challenges have limited the successful translation of candidate biomarkers into clinical use, thus highlighting the need for a robust analytical methodology to transition from biomarker discovery to clinical implementation. We have developed an end-to-end computational proteomic pipeline for biomarkers studies. At the discovery stage, the pipeline emphasizes different aspects of experimental design, appropriate statistical methodologies, and quality assessment of results. At the validation stage, the pipeline focuses on the migration of the results to a platform appropriate for external validation, and the development of a classifier score based on corroborated protein biomarkers. At the last stage towards clinical implementation, the main aims are to develop and validate an assay suitable for clinical deployment, and to calibrate the biomarker classifier using the developed assay. The proposed pipeline was applied to a biomarker study in cardiac transplantation aimed at developing a minimally invasive clinical test to monitor acute rejection. Starting with an untargeted screening of the human plasma proteome, five candidate biomarker proteins were identified. Rejection-regulated proteins reflect cellular and humoral immune responses, acute phase inflammatory pathways, and lipid metabolism biological processes. A multiplex multiple reaction monitoring mass-spectrometry (MRM-MS) assay was developed for the five candidate biomarkers and validated by enzyme-linked immune-sorbent (ELISA) and immunonephelometric assays (INA). A classifier score based on corroborated proteins demonstrated that the developed MRM-MS assay provides an appropriate methodology for an external validation, which is still in progress. Plasma proteomic biomarkers of acute cardiac rejection may offer a relevant post-transplant monitoring tool to effectively guide clinical care. The proposed computational pipeline is highly applicable to a wide range of biomarker proteomic studies.     

A Flexible Integration and Visualisation System for Biomarker Discovery

Biological data have accumulated at an unprecedented pace as a result of improvements in molecular technologies. However, the translation of data into information, and subsequently into knowledge, requires the intricate interplay of data access, visualisation and interpretation. Biological data are complex and are organised either hierarchically or non-hierarchically. For non-hierarchically organised data, it is difficult to view relationships among biological facts. In addition, it is difficult to make changes in underlying data storage without affecting the visualisation interface. Here, we demonstrate a platform where non-hierarchically organised data can be visualised through the application of a customised hierarchy incorporating medical subject headings (MeSH) classifications. This platform gives users flexibility in updating and manipulation. It can also facilitate fresh scientific insight by highlighting biological impacts across different hierarchical branches. An example of the integration of biomarker information from the curated Proteome database using MeSH and the StarTree visualisation tool is presented.     

Potential Diagnostic Value of Serum p53 Antibody for Detecting Esophageal Cancer: A Meta-Analysis

Background

Mutant p53 protein overexpression has been reported to induce serum antibodies against p53. Various studies assessing the diagnostic value of serum p53 antibody in patients with esophageal cancer remain controversial. This study aims to comprehensively and quantitatively summarize the potential diagnostic value of serum p53 antibody in esophageal cancer.

Methods

We systematically searched PubMed and Embase until 31st May 2012, without language restriction. Studies were assessed for quality using QUADAS (quality assessment of studies of diagnostic accuracy). Positive likelihood ratio (PLR) and negative likelihood ratio (NLR) were pooled separately and compared with overall accuracy measures diagnostic odds ratio (DOR) and symmetric summary receiver operating characteristic (sROC). The PLR and NLR and their 95% confidence interval (CI) were calculated using a fixed effects model according to the Mantel-Haensed method and random effects model based on the work of Der Simonian and laird, respectively.

Results

Fifteen studies (cases = 1079, controls = 2260) met the inclusion criteria for the meta-analysis. Approximately 53.33% (8/15) of the included studies were of high quality (QUADAS score≥8), which were retrospective case-control studies. The summary estimates for quantitative analysis of serum p53 antibody in the diagnosis of esophageal cancer were PLR 6.95 (95% CI: 4.77–9.51), NLR 0.75 (95%CI: 0.72–0.78) and DOR 9.65 (95%CI: 7.04–13.22). However, we found significant heterogeneity between NLRs.

Conclusions

The current evidence suggests serum p53 antibody has a potential diagnostic value for esophageal cancer. However, its discrimination power is not perfect because of low sensitivity.

Impact

These results suggest that s-p53-antibody may be useful for monitoring residual tumor cells and for aiding in the selection of candidates for less invasive treatment procedures because of the high specificity of s-p53-antibody. Further studies may need to identify patterns of multiple biomarkers to further increase the power of EC detection.     

Shotgun Proteomics Identifies Serum Fibronectin as a Candidate Diagnostic Biomarker for Inclusion in Future Multiplex Tests for Ectopic Pregnancy

Ectopic pregnancy (EP) is difficult to diagnose early and accurately. Women often present at emergency departments in early pregnancy with a ‘pregnancy of unknown location’ (PUL), and diagnosis and exclusion of EP is challenging due to a lack of reliable biomarkers. The objective of this study was to identify novel diagnostic biomarkers for EP. Shotgun proteomics, incorporating combinatorial-ligand library pre-fractionation, was used to interrogate pooled sera (n = 40) from women undergoing surgery for EP, termination of viable intrauterine pregnancy and management of non-viable intrauterine pregnancy. Western blot was used to validate results in individual sera. ELISAs were developed to interrogate sera from women with PUL (n = 120). Sera were collected at time of first symptomatic presentation and categorized according to pregnancy outcome. The main outcome measures were differences between groups and area under the receiver operating curve (ROC). Proteomics identified six biomarker candidates. Western blot detected significant differences in levels of two of these candidates. ELISA of sera from second cohort revealed that these differences were only significant for one of these candidates, fibronectin. ROC analysis of ability of fibronectin to discriminate EP from other pregnancy outcomes suggested that fibronectin has diagnostic potential (ROC 0.6439; 95% CI 0.5090 to 0.7788; P>0.05), becoming significant when ‘ambiguous’ medically managed PUL excluded from analysis (ROC 0.6538; 95% CI 0.5158 to 0.7918; P<0.05). Fibronectin may make a useful adjunct to future multiplex EP diagnostic tests.     

Towards Plant Species Identification in Complex Samples: A Bioinformatics Pipeline for the Identification of Novel Nuclear Barcode Candidates

Monitoring of the food chain to fight fraud and protect consumer health relies on the availability of methods to correctly identify the species present in samples, for which DNA barcoding is a promising candidate. The nuclear genome is a rich potential source of barcode targets, but has been relatively unexploited until now. Here, we show the development and use of a bioinformatics pipeline that processes available genome sequences to automatically screen large numbers of input candidates, identifies novel nuclear barcode targets and designs associated primer pairs, according to a specific set of requirements. We applied this pipeline to identify novel barcodes for plant species, a kingdom for which the currently available solutions are known to be insufficient. We tested one of the identified primer pairs and show its capability to correctly identify the plant species in simple and complex samples, validating the output of our approach.     

Discovery of Novel Serum Biomarkers for Prenatal Down Syndrome Screening by Integrative Data Mining

Background

To facilitate the experimental search for novel maternal serum biomarkers in prenatal Down Syndrome screening, we aimed to create a set of candidate biomarkers using a data mining approach.

Methodology/Principal Findings

Because current screening markers are derived from either fetal liver or placental trophoblasts, we reasoned that new biomarkers can primarily be found to be derived from these two tissues. By applying a three-stage filtering strategy on publicly available data from different sources, we identified 49 potential blood-detectable protein biomarkers. Our set contains three biomarkers that are currently widely used in either first- or second-trimester screening (AFP, PAPP-A and fβ-hCG), as well as ten other proteins that are or have been examined as prenatal serum markers. This supports the effectiveness of our strategy and indicates the set contains other markers potentially applicable for screening.

Conclusions/Significance

We anticipate the set will help support further experimental studies for the identification of new Down Syndrome screening markers in maternal blood.     

Targeting miR-9 in Glioma Stem Cell-Derived Extracellular Vesicles: A Novel Diagnostic and Therapeutic Biomarker

BackgroundGlioblastoma (GBM) is a lethal brain tumor with no effective strategies in early diagnosis and treatment. This study was aimed to assess the miRNA expression profiles in EVs from CSF and tissue of glioblastoma patients to identify significantly upregulated miRNAs and investigate the underlying neoplastic mechanisms.MethodsEVs were measured by TEM and NTA assays. Differentially regulated miRNAs were measured using RNA sequencing in GBM CSF EVs and in GBM tissues compared with controls. RT-qPCR was employed to analyze miRNA and gene expression. Luciferase report assay was used to investigate gene target of miR-9. The proliferation ability was detected by EdU and CCK-8 experiment while cell migration was measured by transwell and wound healing assay.ResultsThe expression level of miR-9 was significantly higher in GBM CSF EVs and tissues than controls (p = 0.038). The area under curve for CSF EV miR-9 was 0.800 (95% CI: 0.583–1.000, p = 0.033). The expression of miR-9 was significantly higher in Glioma stem cells (GSCs) and GSC-derived EVs than in glioblastoma cells. GSC-derives EVs could promote GBM growth and migration Moreover, inhibition of miR-9 in GSCs showed the reverse anti-tumor effects through secreted EVs. MiR-9 could bind to the 3’UTR region of DACT3 and suppress its expression. The miR-9/DACT3 axis might attribute to GBM malignant phenotype.ConclusionMiR-9 in CSF EVs may act as a novel diagnostic biomarker for GBM and targeting miR-9 by GSC-derived EVs may be a specific and efficient strategy for GBM biotherapy.     

IGFBP3 Methylation Is a Novel Diagnostic and Predictive Biomarker in Colorectal Cancer

Background and Aim

Aberrant hypermethylation of cancer-related genes has emerged as a promising strategy for the development of diagnostic, prognostic and predictive biomarkers in human cancer, including colorectal cancer (CRC). The aim of this study was to perform a systematic and comprehensive analysis of a panel of CRC-specific genes as potential diagnostic, prognostic and predictive biomarkers in a large, population-based CRC cohort.

Patients and Methods

Methylation status of the SEPT9, TWIST1, IGFBP3, GAS7, ALX4 and miR137 genes was studied by quantitative bisulfite pyrosequencing in a population-based cohort of 425 CRC patients.

Results

Methylation levels of all genes analyzed were significantly higher in tumor tissues compared to normal mucosa (p<0.0001); however, cancer-associated hypermethylation was most frequently observed for miR137 (86.7%) and IGFBP3 (83%) in CRC patients. Methylation analysis using the combination of these two genes demonstrated greatest accuracy for the identification of colonic tumors (sensitivity 95.5%; specificity 90.5%). Low levels of IGFBP3 promoter methylation emerged as an independent risk factor for predicting poor disease free survival in stage II and III CRC patients (HR = 0.49, 95% CI: 0.28–0.85, p = 0.01). Our results also suggest that stage II & III CRC patients with high levels of IGFBP3 methylation do not benefit from adjuvant 5FU-based chemotherapy.

Conclusion

By analyzing a large, population-based CRC cohort, we demonstrate the potential clinical significance of miR137 and IGFBP3 hypermethylation as promising diagnostic biomarkers in CRC. Our data also revealed that IGFBP3 hypermethylation may serve as an independent prognostic and predictive biomarker in stage II and III CRC patients.