Haochang Hu

The role of TFPI2 hypermethylation in the detection of gastric and colorectal cancer

Gastrointestinal cancer is a prevalent disease with high morbidity and mortality. Tissue factor pathway inhibitor 2 (TFPI2) gene could protect the extracellular matrix of cancer cells from degradation and tumor invasion. The goal of our study was to estimate the diagnostic value of TFPI2 hypermethylation in gastric cancer (GC) and colorectal cancer (CRC). TFPI2 methylation was measured by quantitative methylation-specific polymerase chain reaction (qMSP) method in 114 GC and 80 CRC tissues and their paired non-tumor tissues. Our results showed that TFPI2 methylation was significantly higher in tumor tissues (GC: 29.940% vs. 12.785%, P < 0.001; CRC: 26.930% vs. 5.420%, P < 0.001). The methylation level of TFPI2 in colorectal tumor tissues was significantly higher than that in colorectal normal tissues (26.930% versus 0.002%, P < 0.00001). In GC, TFPI2 hypermethylation yielded an area under the curve (AUC) of 0.762 (95% CI: 0.696–0.828) with a sensitivity of 68% and a specificity of 83%. In CRC, TFPI2 hypermethylation yielded an AUC of 0.759 (95% CI: 0.685–0.834) with a sensitivity of 61% and a specificity of 84%. Similarly, TCGA data also supported TFPI2 hypermethylation was a promising diagnostic marker for GC and CRC. Moreover, the dual-luciferase reporter assay showed TFPI2 fragment could upregulate gene expression (fold change = 5, P = 0.005). Data mining further indicated that TFPI2 expression in CRC cell lines was significantly increased after 5’-AZA-deoxycytidine treatment (fold change > 1.37). In conclusion, TFPI2 hypermethylation might be a promising diagnostic biomarker for GC and CRC.

CCL2 promoter hypomethylation is associated with gout risk in Chinese Han male population

OBJECTIVESnC-C motif chemokine ligand 2 (CCL2) encodes a chemokine which amplifies and maintains inflammation through chemokine-cytokine networks. Gout is a type of painful inflammation arthritis in response to the deposition of monosodium urate (MSU) crystals in joints. The purpose of this study was to figure out the role of CCL2 promoter methylation in gout.nnnMETHODSnA quantitative methylation-specific polymerase chain reaction (qMSP) was used to measure CCL2 promoter methylation in 93 gout patients and 101 healthy controls, respectively. All participants were Chinese Han males. The percent of methylated reference (PMR) was used to evaluate methylation level of each sample.nnnRESULTSnIn the current study, we have found that CCL2 promoter methylation was significantly lower in gout than in healthy controls (24.95% versus 42.68%, P<0.001). CCL2 promoter hypermethylation was considered as a risk factor of gout [OR (95% CI)=3.59E-04 (5.67E-06, 0.023), P=1.79E-04]. Receiver operating characteristic (ROC) curve analysis showed there was a significant diagnostic value of CCL2 hypomethylation for gout (AUC=0.763, 95% CI=0.695-0.830, sensitivity=0.644, specificity=0.849).nnnCONCLUSIONSnCCL2 promoter hypomethylation in peripheral blood could be a potential biomarker for the diagnosis of gout in Chinese Han males.

Diagnostic value of WIF1 methylation for colorectal cancer: a meta-analysis

As a common antagonist of Wnt/β-catenin signaling, Wnt inhibitory factor 1 (WIF1) plays an important role in the tumor progression. The aim of our meta-analysis was to summarize the diagnostic value of WIF1 methylation in colorectal cancer (CRC). Eligible studies were retrieved by a systemic search among PubMed, Embase, CNKI, and Wanfang literature databases. The diagnostic value of WIF1 methylation for CRC was assessed by the summary receiver operating characteristics (SROC) test. Our meta-analysis of 12 studies between 1420 CRC samples and 946 control samples showed that WIF1 hypermethylation was significantly associated with CRC (P < 0.001, OR = 30.10, 95% CI = 19.48-46.50). WIF1 hypermethylation, as a diagnostic biomarker for CRC, has a pooled sensitivity of 0.40 (95% CI: 0.37-0.42), a pooled specificity of 0.95 (95% CI: 0.93-0.96), a pooled positive-likelihood ratio (PLR) of 8.65 (95% CI, 4.47-16.73), and a pooled negative-likelihood ratio (NLR) of 0.41 (95% CI, 0.30-0.55), a diagnostic odds ratio (DOR) of 26.86 (95% CI: 15.73-45.89), and an area under the curve (AUC) of 0.9115. In conclusion, our study established that WIF1 hypermethylation might be a promising diagnostic biomarker for CRC.

Elevated UMOD methylation level in peripheral blood is associated with gout risk

Uromodulin (UMOD) encodes an uromodulin glycoprotein, and its mutation results in uromodulin glycoprotein dysfunction and the occurrence of gout. The aim of our study was to assess whether UMOD methylation could predict the risk of gout. A total of 89 sporadic gout cases and 103 age and gender-matched healthy controls were recruited in this study. UMOD methylation level was determined by quantitative methylation-specific PCR (qMSP) in peripheral blood, and the percentage of methylated reference (PMR) was described to represent the methylation level. Our results showed that UMOD methylation was significantly higher in gout cases than controls (median: 1.45 versus 0.75, Pu2009<u20090.001). The area under curve (AUC) of UMOD methylation in gout was 0.764 (Pu2009=u20092.90E-10) with a sensitivity of 65.2% and a specificity of 88.3%. UMOD methylation level was shown to be significantly correlated with the serum level of uric acid (UA) (ru2009=u2009−0.208, Pu2009=u20090.035). Besides, the luciferase reporter assay showed that UMOD CpG island region was able to upregulate gene expression (fold changeu2009=u20092, Pu2009=u20090.004). In conclusion, UMOD methylation assessment might be used to predict the occurrence of gout.

FOXF2 promoter methylation is associated with prognosis in esophageal squamous cell carcinoma

Esophageal squamous cell carcinoma is a commonly malignant tumor of digestive tract with poor prognosis. Previous studies suggested that forkhead box F2 (FOXF2) could be a candidate gene for assessing and predicting the prognosis of human cancers. However, the relationship between FOXF2 promoter methylation and the prognosis of esophageal squamous cell carcinoma remained unclear. Formalin-fixed, paraffin-embedded esophageal squamous cell carcinoma tissues of 135 esophageal squamous cell carcinoma patients were detected for FOXF2 promoter methylation status by methylation-specific polymerase chain reaction approach. DNA methylation results were evaluated with regard to clinicopathological features and overall survival. Our study confirmed that FOXF2 promoter hypermethylation could independently predict a poorer overall survival of esophageal squamous cell carcinoma patients (pu2009=u20090.002), which was consistent with the data mining results of the data from 82 esophageal squamous cell carcinoma patients in The Cancer Genome Atlas datasets (pu2009=u20090.036). In addition, no correlation was found between FOXF2 promoter methylation and other clinic pathological parameters (age, gender, differentiation, lymph node metastasis, stage, cutting edge, vascular invasion, smoking behavior, and drinking history). In conclusion, FOXF2 methylation might be a useful prognostic biomarker for esophageal squamous cell carcinoma patients.

TNFRSF10C methylation is a new epigenetic biomarker for colorectal cancer

Background Abnormal methylation of TNFRSF10C was found to be associated with different types of cancers, excluding colorectal cancer (CRC). In this paper, the performance of TNFRSF10C methylation in CRC was studied in two stages. Method The discovery stage was involved with 38 pairs of CRC tumor and paired adjacent non-tumor tissues, and 69 pairs of CRC tumor and paired adjacent non-tumor tissues were used for the validation stage. Quantitative methylation specific PCR (qMSP) method and percentage of methylated reference (PMR) were used to test and represent the methylation level of TNFRSF10C, respectively. A dual-luciferase reporter gene experiment was conducted to evaluate the promoter activity of TNFRSF10C fragment. Results A significant association of TNFRSF10C promoter hypermethylation with CRC was found and validated (discovery stage: 24.67 ± 7.52 vs. 3.36 ± 0.89; P = 0.003; validation stage: 31.21 ± 12.48 vs. 4.52 ± 1.47; P = 0.0005). Subsequent analyses of TCGA data among 46 pairs of CRC samples further confirmed our findings (cg23965061: P = 4E − 6; cg14015044: P = 1E − 7). Dual-luciferase reporter gene assay revealed that TNFRSF10C fragment was able to significantly promote gene expression (Fold change = 2.375, P = 0.013). Our data confirmed that TNFRSF10C promoter hypermethylation can predict shorter overall survival of CRC patients (P = 0.032). Additionally, bioinformatics analyses indicated that TNFRSF10C hypermethylation was significantly associated with lower TNFRSF10C expression. Conclusion Our work suggested that TNFRSF10C hypermethylation was significantly associated with the risk of CRC.

Diagnostic value of RASSF1A hypermethylation in colorectal cancer: a meta-analysis

PURPOSEnRas association domain family 1 isoform A (RASSF1A), a member of Ras association domain family, plays an important role in tumorigenesis. The goal of our meta-analysis was to assess the diagnostic value of RASSF1A hypermethylation in colorectal cancer (CRC).nnnMETHODSnPubMed, Embase, CNKI and Wanfang databases were used to conduct literature selection. The association between RASSF1A methylation and CRC risk was evaluated by odds ratios (ORs) and 95% confidence intervals (CIs). Summary receiver operating characteristics (SROC) test was used to estimate the diagnostic value of RASSF1A methylation for CRC.nnnRESULTSnA total of 22 articles among 1736 CRC and 811 non-tumor samples were included in the current meta-analysis. Our results showed that RASSF1A hypermethylation was found more frequently in CRC than non-tumor samples (ORu202f=u202f6.02, 95% CIu202f=u202f4.57-7.93, Pu202f<u202f 0.001). Our SROC test showed that RASSF1A hypermethylation had an area under the curve (AUC) of 0.71 with a pooled sensitivity of 0.33 (95% CIu202f=u202f0.31-0.36), a pooled specificity of 0.86 (95% CIu202f=u202f0.84-0.89), a positive-likelihood ratio of 3.18 (95% CIu202f=u202f1.99-5.09), a negative-likelihood ratio of 0.71 (95% CIu202f=u202f0.63-0.80), and a diagnostic odds ratio of 5.53 (95% CIu202f=u202f3.40-9.00). Data mining study indicated that a trend of increased RASSF1A expression was found in the CRC cell line C2C12 after 5-AZA treatment.nnnCONCLUSIONSnOur study established that RASSF1A hypermethylation might have a potential value in the clinical diagnosis of CRC.

Hypermethylated Promoters of Secreted Frizzled-Related Protein Genes are Associated with Colorectal Cancer

Colorectal cancer (CRC) is one of the leading causes of death worldwide. Aberrant DNA methylation has been recognized as one of the most common molecular alterations in CRC. The goal of this study was to investigate the diagnostic value of SFRP1 and SFRP2 methylation for CRC. A total of 80 pairs of CRC patients were recruited to test the association of SFRP1 and SFRP2 promotor methylation with CRC. Methylation assay was performed using quantitative methylation-specific polymerase chain reaction (qMSP) method. In this study, we found the methylation levels of SFRP1 and SFRP2 in CRC tumor tissues were significantly higher than those in the adjacent non-tumor tissues (SFRP1: Pu2009=u20092E-5; SFRP2: Pu2009=u20090.014). Further bioinformatics analysis of TCGA data confirmed the association of the two genes with CRC (SFRP1: Pu2009=u20097E-21; SFRP2: Pu2009=u20095E-24). Luciferase reporter gene assay showed that the recombinant plasmids with SFRP1 and SFRP2 fragments could significantly enhance promoter activity (SFRP1: Pu2009=u20090.002; SFRP2: Pu2009=u20090.004). In addition, SFRP1 and SFRP2 methylation were inversely correlated with the mRNA expression displayed by TCGA data mining (SFRP1: ru2009=u2009−0.432, Pu2009=u20094E-11; SFRP2: ru2009=u2009−0.478, Pu2009=u20091E-13). GEO data analysis indicated that SFRP1 and SFRP2 expression were increased in three CRC cell lines (COLO320, HCT116 and HT29) after 5′-AZA-deoxycytidine treatment, suggesting that DNA methylation played an important role in regulating gene expression of the two genes. Our results confirmed that promoter methylation of SFRP1 and SFRP2 contributed to the risk of CRC.

Hypermethylation of MDFI promoter with NSCLC is specific for females, non‑smokers and people younger than 65

Non-small cell lung carcinoma (NSCLC) is a major subtype of lung cancer. Aberrant DNA methylation has been frequently observed in NSCLC. The aim of the present study was to investigate the role of MyoD family inhibitor (MDFI) methylation in NSCLC. Formalin-fixed paraffin-embedded tumor tissues and adjacent non-cancerous tissues were collected from a total of 111 patients with NSCLC. A methylation assay was performed using the quantitative methylation-specific polymerase chain reaction method. The percentage of methylated reference was used to represent the methylation level of the MDFI promoter. Data mining of a dataset from The Cancer Genome Atlas (TCGA) demonstrated that MDFI promoter methylation levels were significantly increased in 830 tumor tissues compared with 75 non-tumor tissues (P=0.012). However, the results on tissues obtained in the present study indicated that the MDFI promoter methylation levels in tumor tissues were not significantly different compared with those in the adjacent non-tumor tissues (P=0.159). Subsequent breakdown analysis identified that higher MDFI promoter methylation levels were significantly associated with NSCLC in females (P=0.031), but not in males (P=0.832). Age-based subgroup analysis demonstrated that higher MDFI promoter methylation levels were significantly associated with NSCLC in younger patients (≤65 years; P=0.003), but not in older patients (P=0.327). In addition, the association of MDFI methylation with NSCLC was significant in non-smokers (P=0.014), but not in smokers (P=0.832). Similar results also have been determined from subgroup analysis of the TCGA datasets. The Gene Expression Omnibus database indicated MDFI expression restoration in partial lung cancer cell lines (H1299 and Hotz) following demethylation treatment. However, it was identified that MDFI promoter hypermethylation was not significantly associated with prognosis of NSCLC (P>0.05). In conclusion, the present study indicated that the association of higher methylation of the MDFI promoter with NSCLC may be specific to females, non-smokers and people aged ≤65.

Aberrant methylation of mutL homolog 1 is associated with increased risk of non-small cell lung cancer

Non‐small cell lung cancer (NSCLC) is a common malignant tumor. DNA hypermethylation in the promoter region has been served as a potential molecular marker for several tumors. The goal of the current study was to assess the diagnostic ability of mutL homolog 1 (MLH1) promoter methylation in NSCLC.