Angao Xu

Clinical significance of HSP27 expression in colorectal cancer

The heat shock protein 27-kDa (HSP27) has been found overexpressed in several types of human cancer and is associated with treatment resistance and poor prognosis. Recent proteomic studies demonstrate that HSP27 is significantly overexpressed in colorectal cancer (CRC). However, the relationship between HSP27 expression and patient prognosis remains nascent. In the present study, we aimed to investigate the expression of HSP27 and its correlation with clinicopathological parameters in CRC patients. Dysregulated expression of HSP27 was observed in neoplastic lesions, and appears to be involved in disease progression. Immunohistochemical analysis showed that detectable HSP27 expression was found in 145/182 (79.7%) CRC cases. There was a significant correlation between HSP27 expression and TNM stage (P=0.003). Patients with low HSP27 expression had better survival than those with high HSP27 expression. Additionally, multivariate analysis indicated that HSP27 expression is an independent prognostic marker for CRC. These results suggest that elevated expression of HSP27 protein is a frequent event during the progression of CRC. HSP27 might be used as a valuable prognostic marker for patients with CRC.

Diagnostic value of stool DNA testing for multiple markers of colorectal cancer and advanced adenoma: a meta-analysis.

BACKGROUND AND OBJECTIVES The diagnostic value of stool DNA (sDNA) testing for colorectal neoplasms remains controversial. To compensate for the lack of large-scale unbiased population studies, a meta-analysis was performed to evaluate the diagnostic value of sDNA testing for multiple markers of colorectal cancer (CRC) and advanced adenoma. METHODS The PubMed, Science Direct, Biosis Review, Cochrane Library and Embase databases were systematically searched in January 2012 without time restriction. Meta-analysis was performed using a random-effects model using sensitivity, specificity, diagnostic OR (DOR), summary ROC curves, area under the curve (AUC), and 95% CIs as effect measures. Heterogeneity was measured using the χ(2) test and Q statistic; subgroup analysis was also conducted. RESULTS A total of 20 studies comprising 5876 individuals were eligible. There was no heterogeneity for CRC, but adenoma and advanced adenoma harboured considerable heterogeneity influenced by risk classification and various detection markers. Stratification analysis according to risk classification showed that multiple markers had a high DOR for the high-risk subgroups of both CRC (sensitivity 0.759 [95% CI 0.711 to 0.804]; specificity 0.883 [95% CI 0.846 to 0.913]; AUC 0.906) and advanced adenoma (sensitivity 0.683 [95% CI 0.584 to 0.771]; specificity 0.918 [95% CI 0.866 to 0.954]; AUC 0.946) but not for the average-risk subgroups of either. In the methylation subgroup, sDNA testing had significantly higher DOR for CRC (sensitivity 0.753 [95% CI 0.685 to 0.812]; specificity 0.913 [95% CI 0.860 to 0.950]; AUC 0.918) and advanced adenoma (sensitivity 0.623 [95% CI 0.527 to 0.712]; specificity 0.926 [95% CI 0.882 to 0.958]; AUC 0.910) compared with the mutation subgroup. There was no significant heterogeneity among studies for subgroup analysis. CONCLUSION sDNA testing for multiple markers had strong diagnostic significance for CRC and advanced adenoma in high-risk subjects. Methylation makers had more diagnostic value than mutation markers.

Lyn is involved in CD24-induced ERK1/2 activation in colorectal cancer

CD24 expression is associated with human colorectal cancer (CRC). Our previous data indicated that CD24 promoted the proliferation and invasion of colorectal cancer cells through the activation of ERK1/2. Since Src family kinases are frequently deregulated in CRC and closely related to the MAPK signaling pathway, we investigated the impact of Lyn, an important member of SFKs, on CD24-induced ERK1/2 activation in CRC. The interaction of CD24 and Lyn was identified by co-immunoprecipitation (Co-IP) and ectopic expression of CD24-induced Lyn activation. Inhibition of Lyn activation by phosphatase PP2 in SW480CD24cells abrogated CD24-induced invasion. The results of the Co-IP and immunofluorescence assay revealed that overexpression of CD24 enhanced the interaction of Lyn and ERK1/2 and induced the nuclear translocation of Lyn. However, inhibition of Lyn activity attenuated CD24-induced ERK1/2 activation, and depletion of CD24 disrupted Lyn-ERK1/2 interaction. Immunohistochemistry analysis for 202 cases of CRC showed that the expression of both CD24 and Lyn was positively correlated with tumor grade, stage, lymph node and distant metastasis. Patients with lower expression of CD24 or Lyn had a higher survival rate. The Cox multivariate analysis showed that CD24 expression, but not Lyn expression, was an independent prognostic factor of CRC. Our results suggest that Lyn is involved in CD24-induced ERK1/2 activation in CRC. The expression of CD24 is associated with activation of Lyn and ERK1/2, which might be a novel mechanism related to CD24-mediated regulation of CRC development.

Is High Resolution Melting Analysis (HRMA) Accurate for Detection of Human Disease-Associated Mutations? A Meta Analysis

Background High Resolution Melting Analysis (HRMA) is becoming the preferred method for mutation detection. However, its accuracy in the individual clinical diagnostic setting is variable. To assess the diagnostic accuracy of HRMA for human mutations in comparison to DNA sequencing in different routine clinical settings, we have conducted a meta-analysis of published reports. Methodology/Principal Findings Out of 195 publications obtained from the initial search criteria, thirty-four studies assessing the accuracy of HRMA were included in the meta-analysis. We found that HRMA was a highly sensitive test for detecting disease-associated mutations in humans. Overall, the summary sensitivity was 97.5% (95% confidence interval (CI): 96.8–98.5; I2 = 27.0%). Subgroup analysis showed even higher sensitivity for non-HR-1 instruments (sensitivity 98.7% (95%CI: 97.7–99.3; I2 = 0.0%)) and an eligible sample size subgroup (sensitivity 99.3% (95%CI: 98.1–99.8; I2 = 0.0%)). HRMA specificity showed considerable heterogeneity between studies. Sensitivity of the techniques was influenced by sample size and instrument type but by not sample source or dye type. Conclusions/Significance These findings show that HRMA is a highly sensitive, simple and low-cost test to detect human disease-associated mutations, especially for samples with mutations of low incidence. The burden on DNA sequencing could be significantly reduced by the implementation of HRMA, but it should be recognized that its sensitivity varies according to the number of samples with/without mutations, and positive results require DNA sequencing for confirmation.

Validation of methylation-sensitive high-resolution melting (MS-HRM) for the detection of stool DNA methylation in colorectal neoplasms

BACKGROUND Methylation-sensitive high-resolution melting (MS-HRM) is a new technique for assaying DNA methylation, but its feasibility for assaying stool in patients with colorectal cancer (CRC) is unknown. METHODS First, the MS-HRM and methylation-specific PCR (MSP) detection limits were tested. Second, the methylation statuses of SFRP2 and VIM were analyzed in stool samples by MS-HRM, and in matching tumor and normal colon tissues via bisulfite sequencing PCR (BSP). Third, a case-control study evaluated the diagnostic sensitivity and specificity of MS-HRM relative to results obtained with MSP and the fecal immunochemical test (FIT). Finally, the linearity and reproducibility of MS-HRM were assessed. RESULTS The detection limits of MS-HRM and MSP were 1% and 5%, respectively. The diagnostic sensitivities of MS-HRM (87.3%, 55/63) in stool and BSP in matching tumor tissue (92.1%, 58/63) were highly consistent (κ=0.744). The MS-HRM assay detected 92.5% (37/40) methylation in CRCs, 94.4% (34/36) in advanced adenomas, and 8.8% (5/57) in normal controls. The results of MS-HRM analysis were stable and reliable and showed fairly good linearity for both SFRP2 (P<0.001, R(2)=0.957) and VIM (P<0.001, R(2)=0.954). CONCLUSIONS MS-HRM shows potential for CRC screening.

High-Resolution Melting Assay (HRMA) is a Simple and Sensitive Stool-Based DNA Test for the Detection of Mutations in Colorectal Neoplasms

BACKGROUND Stool-based DNA testing for colorectal cancer is becoming a favored alternative to existing DNA screening tests. However, current methods of analysis often become more complicated and costly with increased sensitivity. The high-resolution melting assay (HRMA) is a simple and rapid mutation scanning method with low cost and superb accuracy. In this study, we verified the accuracy of HRMA for screening KRAS/TP53 mutations in stool-isolated DNA from patients with colorectal cancer. MATERIALS AND METHODS Comparing to direct DNA sequencing, the accuracy of HRMA was verified by detecting KRAS/TP53 mutations in 2 independent stages. In study stage I, both tissue and stool samples from colorectal neoplasm patients were analyzed. In study stage II, stool samples from patients with colorectal neoplasms, and normal controls in clinical screening settings were examined. RESULTS In study stage I, the HRMA identified 14 of 17 target mutations (82.4%) in stools from cancer patients, and 4 of 5 (80.0%) target mutations in stools from advanced adenoma patients. The mutation detection rate in fecal samples (45.0%; 18/40) and referred tissue samples (55.0%; 22/40) was highly consistent (κ = 0.79). The HRMA detected 1% mutant DNA in a background of wild type DNA. In study stage II, the HRMA assay detected 58.8% (20/34) mutations in tumor samples, 41.5% (17/41) in advanced adenomas samples, and 3.33% (2/60) in age-matched normal control samples. The results from HRMA and DNA sequencing revealed 100% sensitivity and specificity in both tissue and stool samples. CONCLUSION HRMA is a simple, reliable, and sensitive method for detecting DNA mutations in the stool samples from patients with colorectal neoplasms.

Diagnostic Performance of DNA Hypermethylation Markers in Peripheral Blood for the Detection of Colorectal Cancer: A Meta-Analysis and Systematic Review

DNA hypermethylation in blood is becoming an attractive candidate marker for colorectal cancer (CRC) detection. To assess the diagnostic accuracy of blood hypermethylation markers for CRC in different clinical settings, we conducted a meta-analysis of published reports. Of 485 publications obtained in the initial literature search, 39 studies were included in the meta-analysis. Hypermethylation markers in peripheral blood showed a high degree of accuracy for the detection of CRC. The summary sensitivity was 0.62 [95% confidence interval (CI), 0.56–0.67] and specificity was 0.91 (95% CI, 0.89–0.93). Subgroup analysis showed significantly greater sensitivity for the methylated Septin 9 gene (SEPT9) subgroup (0.75; 95% CI, 0.67–0.81) than for the non-methylated SEPT9 subgroup (0.58; 95% CI, 0.52–0.64). Sensitivity and specificity were not affected significantly by target gene number, CRC staging, study region, or methylation analysis method. These findings show that hypermethylation markers in blood are highly sensitive and specific for CRC detection, with methylated SEPT9 being particularly robust. The diagnostic performance of hypermethylation markers, which have varied across different studies, can be improved by marker optimization. Future research should examine variation in diagnostic accuracy according to non-neoplastic factors.

Correction: Lyn is involved in CD24-induced ERK1/2 activation in colorectal cancer

Correction To further confirm that the inactivation of Lyn inhibited CD24-induced ERK1/2 activation, we submitted the results of another CD24siRNA (CD24siRNA-2) as the Additional file 1: Figure S2. After publication of the article [1] the authors noted that the wrong figure for Additional file 1: Figure S2 was submitted by mistake. The correct image was conducted in SW480 using CD24siRNA-2 (new Additional file 1: attached). The authors apologise for any inconvenience caused.

945 CD24 Regulated the Properties of Self-Renewal, Drug-Resistant, and Tumorigenicity of Colorectal Cancer Stem Cell

isms of any sensitizing or direct action. Methods: Afferent mechanoand chemosensitivity was assessed in mouse colo-rectum In Vitro. High-threshold nociceptive afferents were characterised by responses to mechanical stimuli (3). Cytokine receptor abundance was assessed using PCR of Dorsal Root Ganglia (DRG) (L6-S1) and specifically in laser-captured colonic DRG neurons. Healthy and PIVH (TNBS 130μg/mL with 28 day recovery) mice were compared (3). Results: In healthy mice TNF-α (100ng/mL) induced potent mechanical hypersensitivity (P<0.0001) that was blocked by ruthenium red and selective TRPA1 antagonism (HC-030031), but not by selective antagonism of TRPV1 (capsazepine). TNF-α had no direct chemosensory effects. A switch in effect occurred in PIVH whereby TNF-α inhibited mechanical hypersensitivity. This inhibition was blocked by iberiotoxin, a selective potassium channel (BKCa) antagonist. TNF1 or 2 receptor mRNA abundance in DRG was unaltered. In healthy mice IL-1β (20ng/mL) induced modest mechanical hypersensitivity but caused a direct chemosensory effect in 4/7 afferent recordings, that was inhibited by tetrodotoxin (TTX). Similar responses occurred in PIVH, and IL-1R1 mRNA abundance was unaltered. IL-2 (100ng/mL) had no effect in healthy mice (n=7), but induced a profound TTX-sensitive chemosensory effect in PIVH (7/10 afferents), that was greater than observed for IL-1β, and a significant (P<0.05) increase in IL-2 receptor mRNA. Data are shown for pelvic serosal afferents, but were broadly similar in splanchnic serosal afferents. Discussion: IL-1β and IL2 induce direct chemosensory effects in colonic afferents via sodium channels without alteringmechanosensory function. In contrast, TNF-α exclusively induced mechanical hypersensitivity, predominately via TRPA1. In PIVH, IL-1β effects are unchanged, IL-2 effects are increased, and TNF-α effects are in the opposite direction whereby TNF-α now causes inhibition via BKCa channels. Therefore cytokines interact with visceral afferents via different mechanisms, which undergo markedly different changes in PIVH. 1: Leibregts Gastro 2007. 2: Hughes Gut 2009. 3: Hughes Gut 2009.