Gene promoter methylation in colorectal cancer and healthy adjacent mucosa specimens: Correlation with physiological and pathological characteristics,and with biomarkers of one-carbon metabolism

We evaluated the promoter methylation levels of the APC, MGMT, hMLH1, RASSF1A and CDKN2A genes in 107 colorectal cancer (CRC) samples and 80 healthy adjacent tissues. We searched for correlation with both physical and pathological features, polymorphisms of folate metabolism pathway genes (MTHFR, MTRR, MTR, RFC1, TYMS, and DNMT3B), and data on circulating folate, vitamin B12 and homocysteine, which were available in a subgroup of the CRC patients. An increased number of methylated samples were found in CRC respect to adjacent healthy tissues, with the exception of APC, which was also frequently methylated in healthy colonic mucosa. Statistically significant associations were found between RASSF1A promoter methylation and tumor stage, and between hMLH1 promoter methylation and tumor location. Increasing age positively correlated with both hMLH1 and MGMT methylation levels in CRC tissues, and with APC methylation levels in the adjacent healthy mucosa. Concerning gender, females showed higher hMLH1 promoter methylation levels with respect to males. In CRC samples, the MTR 2756AG genotype correlated with higher methylation levels of RASSF1A, and the TYMS 1494 6bp ins/del polymorphism correlated with the methylation levels of both APC and hMLH1. In adjacent healthy tissues, MTR 2756AG and TYMS 1494 6bp del/del genotypes correlated with APC and MGMT promoter methylation, respectively. Low folate levels were associated with hMLH1 hypermethylation. Present results support the hypothesis that DNA methylation in CRC depends from both physiological and environmental factors, with one-carbon metabolism largely involved in this process.     

Gene promoter methylation in colorectal cancer and healthy adjacent mucosa specimens

We evaluated the promoter methylation levels of the APC, MGMT, hMLH1, RASSF1A and CDKN2A genes in 107 colorectal cancer (CRC) samples and 80 healthy adjacent tissues. We searched for correlation with both physical and pathological features, polymorphisms of folate metabolism pathway genes (MTHFR, MTRR, MTR, RFC1, TYMS, and DNMT3B), and data on circulating folate, vitamin B12 and homocysteine, which were available in a subgroup of the CRC patients. An increased number of methylated samples were found in CRC respect to adjacent healthy tissues, with the exception of APC, which was also frequently methylated in healthy colonic mucosa. Statistically significant associations were found between RASSF1A promoter methylation and tumor stage, and between hMLH1 promoter methylation and tumor location. Increasing age positively correlated with both hMLH1 and MGMT methylation levels in CRC tissues, and with APC methylation levels in the adjacent healthy mucosa. Concerning gender, females showed higher hMLH1 promoter methylation levels with respect to males. In CRC samples, the MTR 2756AG genotype correlated with higher methylation levels of RASSF1A, and the TYMS 1494 6bp ins/del polymorphism correlated with the methylation levels of both APC and hMLH1. In adjacent healthy tissues, MTR 2756AG and TYMS 1494 6bp del/del genotypes correlated with APC and MGMT promoter methylation, respectively. Low folate levels were associated with hMLH1 hypermethylation. Present results support the hypothesis that DNA methylation in CRC depends from both physiological and environmental factors, with one-carbon metabolism largely involved in this process.     

Detection of MGMT, RASSF1A, p15INK4B, and p14ARF promoter methylation in circulating tumor-derived DNA of central nervous system cancer patients

Despite the growing understanding of the mechanisms of carcinogenesis, cancers of the central nervous system are usually associated with unfavorable prognosis. The use of an appropriate molecular marker may improve the treatment outcome by allowing early diagnosis and treatment susceptibility monitoring. Since methylation of tumor-derived DNA can be detected in the serum of cancer patients, this makes DNA methylation-based biomarkers one of the most promising diagnostic strategies. In this study, the methylation profiles of MGMT, RASSF1A, p15INK4B, and p14ARF genes were evaluated in serum free-circulating DNA and the corresponding tumor tissue in a group of 33 primary or metastatic central nervous system cancer patients. Gene promoter methylation was assessed using methylation-specific polymerase chain reaction (PCR). All the tested genes were found to be methylated to a different extent in both serum and tumor samples. In comparison to metastatic brain tumor patients, the patients with glial tumors were characterized by a higher frequency of gene hypermethylation. The hypermethylation of RASSF1A differentiated primary from metastatic brain cancers. Moreover, the gene methylation profiles observed in serum, in most cases, matched the methylation profiles detected in paired tumor samples.     

Aberrant promoter methylation of RASSF1A gene may be correlated with colorectal carcinogenesis: a meta-analysis

We conducted a meta-analysis of cohort studies to evaluate the potential role of RASSF1A promoter methylation in colorectal carcinogenesis. A range of electronic databases were searched: PubMed (1966–2013), the Cochrane Library Database (Issue 12, 2013), EMBASE (1980–2013), CINAHL (1982–2013), Web of Science (1945–2013) and the Chinese Biomedical Database (CBM) (1982–2013) without language restrictions. Meta-analysis was conducted using the STATA 12.0 software. Crude odds ratio (OR) with 95 % confidence interval (95 % CI) was calculated. Eleven clinical cohort studies with a total of 1,505 colorectal cancer (CRC) patients that met all inclusion criteria were included in our meta-analysis. The results of our meta-analysis revealed that the frequency of RASSF1A promoter methylation was strongly correlated with clinical stage (OR = 1.69, 95 % CI 1.16–2.44, P = 0.006), histological grade (OR = 1.92, 95 % CI 1.22–3.04, P = 0.005) and distant metastasis (OR = 2.59, 95 % CI 1.46–4.60, P = 0.037) of CRC patients. However, we observed no positive correlations of RASSF1A promoter methylation with gender (OR = 1.04, 95 % CI 0.74–1.46, P = 0.842), age (OR = 1.70, 95 % CI 0.98–2.93, P = 0.057) and lymph node metastasis (OR = 1.65, 95 % CI 0.87–3.14, P = 0.127) of CRC patients. Further subgroup analysis by ethnicity demonstrated that RASSF1A promoter methylation was correlated with clinicopathological characteristics of CRC patients among Asians (clinical stage: OR = 2.55, 95 % CI 1.55–4.20, P < 0.001; histological grade: OR = 2.70, 95 % CI 1.44–5.06, P = 0.002; lymph node metastasis: OR = 4.09, 95 % CI 1.49–11.26, P = 0.006; distant metastasis: OR = 5.38, 95 % CI 1.73–16.70, P = 0.004), but not among Caucasians and Africans (all P > 0.05). Our meta-analysis has shown positive correlations between aberrant promoter methylation of RASSF1A gene and clinicopathological characteristics of CRC patients, especially among Asians.     

Halplotypes of the ovine ADRB3 gene (ADRB3) and their association with post-weaning growth in New Zealand Suffolk sheep

The β₃-adrenergic receptor (ADRB3) regulates thermogenesis and lipolysis in brown and white adipose tissue. Previously, sixteen ovine ADRB3 haplotypes have been defined. In this study, the relationship between these ADRB3 haplotypes and variation in post-weaning growth was investigated in 797 New Zealand Suffolk lambs from 38 sire lines and eight studs, using PCR-SSCP and General Linear Mixed-effects Models. Seven haplotypes were found in these sheep and they comprised five previously reported intron sequences and four previously reported 3′UT sequences. The frequencies of the various diplotypes ranged from 0.1 to 17.6 % and individual haplotypes from 0.8 to 32.5 %. The presence of haplotype A-b was associated with a decreased weaning-weight (P = 0.001). Sheep with the B-c/F-e diplotype had a higher mean weaning-weight than those with A-b/B-c or A-b/E-e (P < 0.05). The presence of C-a was found to be associated with increased post-weaning growth (P = 0.008), while the presence of B-c was associated with decreased post-weaning growth (P = 0.005). Sheep with A-b/C-a had higher mean post-weaning growth than those with A-b/A-b, A-b/B-c, B-c/B-c, B-c/E-e or B-c/F-e (P < 0.05). Sheep with B-c/B-c had lower mean post-weaning growth than those with A-b/C-a or B-c/C-a (P < 0.05). Additive effects for the different forms of the B-c haplotypes on post-weaning growth were identified. The effects of the ovine ADRB3 haplotypes on weaning-weight and post-weaning growth confirm that they could be used as a candidate gene-marker for improving sheep growth.     

Aberrant Gene Promoter Methylation Associated with Sporadic Multiple Colorectal Cancer

Background

Colorectal cancer (CRC) multiplicity has been mainly related to polyposis and non-polyposis hereditary syndromes. In sporadic CRC, aberrant gene promoter methylation has been shown to play a key role in carcinogenesis, although little is known about its involvement in multiplicity. To assess the effect of methylation in tumor multiplicity in sporadic CRC, hypermethylation of key tumor suppressor genes was evaluated in patients with both multiple and solitary tumors, as a proof-of-concept of an underlying epigenetic defect.

Methodology/Principal Findings

We examined a total of 47 synchronous/metachronous primary CRC from 41 patients, and 41 gender, age (5-year intervals) and tumor location-paired patients with solitary tumors. Exclusion criteria were polyposis syndromes, Lynch syndrome and inflammatory bowel disease. DNA methylation at the promoter region of the MGMT, CDKN2A, SFRP1, TMEFF2, HS3ST2 (3OST2), RASSF1A and GATA4 genes was evaluated by quantitative methylation specific PCR in both tumor and corresponding normal appearing colorectal mucosa samples. Overall, patients with multiple lesions exhibited a higher degree of methylation in tumor samples than those with solitary tumors regarding all evaluated genes. After adjusting for age and gender, binomial logistic regression analysis identified methylation of MGMT2 (OR, 1.48; 95% CI, 1.10 to 1.97; p = 0.008) and RASSF1A (OR, 2.04; 95% CI, 1.01 to 4.13; p = 0.047) as variables independently associated with tumor multiplicity, being the risk related to methylation of any of these two genes 4.57 (95% CI, 1.53 to 13.61; p = 0.006). Moreover, in six patients in whom both tumors were available, we found a correlation in the methylation levels of MGMT2 (r = 0.64, p = 0.17), SFRP1 (r = 0.83, 0.06), HPP1 (r = 0.64, p = 0.17), 3OST2 (r = 0.83, p = 0.06) and GATA4 (r = 0.6, p = 0.24). Methylation in normal appearing colorectal mucosa from patients with multiple and solitary CRC showed no relevant difference in any evaluated gene.

Conclusions

These results provide a proof-of-concept that gene promoter methylation is associated with tumor multiplicity. This underlying epigenetic defect may have noteworthy implications in the prevention of patients with sporadic CRC.     

DNA methylation at promoter regions of interleukin 1B, interleukin 6, and interleukin 8 in non-small cell lung cancer

Epidemiologic and experimental evidences support the concept that inflammation promotes the development and progression of cancers. Interleukins (ILs) regulate the expression of several molecules and signaling pathways involved in inflammation. High expression of some ILs in the tumor microenvironment has been associated with a more virulent tumor phenotype. To examine the role of IL-1β, IL-6, and IL-8 in non-small cell lung cancer, we measured mRNA levels and promoter DNA methylation in a panel of cultured human lung cells (n = 23) and in matched pair lung tumor versus adjacent non-tumorous tissues (n = 24). We found that lung cancer cells or tissues had significantly different DNA methylation and mRNA levels than normal human bronchial epithelial cells or adjacent non-tumorous tissues, respectively. High DNA methylation of ILs promoters in lung cancer cells or tissues was associated with low mRNA levels. We found an inverse correlation between DNA methylation of IL1B, IL6, and IL8 gene promoters and their corresponding mRNA levels, such inverse correlation was more significant for IL1B (i.e., all cancer cell lines used in this study had a hypermethylated IL1B promoter which was associated with silencing of the gene). Our results underline for the first time the role of epigenetic modifications in the regulation of the expression of key cytokines involved in the inflammatory response during lung cancer development.     

Investigating the potential role of genetic and epigenetic variation of DNA methyltransferase genes in hyperplastic polyposis syndrome

Background

Hyperplastic Polyposis Syndrome (HPS) is a condition associated with multiple serrated polyps, and an increased risk of colorectal cancer (CRC). At least half of CRCs arising in HPS show a CpG island methylator phenotype (CIMP), potentially linked to aberrant DNA methyltransferase (DNMT) activity. CIMP is associated with methylation of tumor suppressor genes including regulators of DNA mismatch repair (such as MLH1, MGMT), and negative regulators of Wnt signaling (such as WIF1). In this study, we investigated the potential for interaction of genetic and epigenetic variation in DNMT genes, in the aetiology of HPS.

Methods

We utilized high resolution melting (HRM) analysis to screen 45 cases with HPS for novel sequence variants in DNMT1, DNMT3A, DNMT3B, and DNMT3L. 21 polyps from 13 patients were screened for BRAF and KRAS mutations, with assessment of promoter methylation in the DNMT1, DNMT3A, DNMT3B, DNMT3L MLH1, MGMT, and WIF1 gene promoters.

Results

No pathologic germline mutations were observed in any DNA-methyltransferase gene. However, the T allele of rs62106244 (intron 10 of DNMT1 gene) was over-represented in cases with HPS (p<0.01) compared with population controls. The DNMT1, DNMT3A and DNMT3B promoters were unmethylated in all instances. Interestingly, the DNMT3L promoter showed low levels of methylation in polyps and normal colonic mucosa relative to matched disease free cells with methylation level negatively correlated to expression level in normal colonic tissue. DNMT3L promoter hypomethylation was more often found in polyps harbouring KRAS mutations (p = 0.0053). BRAF mutations were common (11 out of 21 polyps), whilst KRAS mutations were identified in 4 of 21 polyps.

Conclusions

Genetic or epigenetic alterations in DNMT genes do not appear to be associated with HPS, but further investigation of genetic variation at rs62106244 is justified given the high frequency of the minor allele in this case series.     

Mutation and expression analysis of the IDH1, IDH2, DNMT3A, and MYD88 genes in colorectal cancer

Colorectal cancer (CRC) is one of the leading causes of death around the world. Its genetic mechanism was intensively investigated in the past decades with findings of a number of canonical oncogenes and tumor-suppressor genes such as APC, KRAS, and TP53. Recent genome-wide association and sequencing studies have identified a series of promising oncogenes including IDH1, IDH2, DNMT3A, and MYD88 in hematologic malignancies. However, whether these genes are involved in CRC remains unknown. In this study, we screened the hotspot mutations of these four genes in 305 CRC samples from Han Chinese by direct sequencing. mRNA expression levels of these genes were quantified by quantitative real-time PCR (RT-qPCR) in paired cancerous and paracancerous tissues. Association analyses between mRNA expression levels and different cancerous stages were performed. Except for one patient harboring IDH1 mutation p.I99M, we identified no previously reported hotspot mutations in colorectal cancer tissues. mRNA expression levels of IDH1, DNMT3A, and MYD88, but not IDH2, were significantly decreased in the cancerous tissues comparing with the paired paracancerous normal tissues. Taken together, the hotspot mutations of IDH1, IDH2, DNMT3A, and MYD88 gene were absent in CRC. Aberrant mRNA expression of IDH1, DNMT3A, and MYD88 gene might be actively involved in the development of CRC.     

DNMT1, DNMT3A and DNMT3B gene variants in relation to ovarian cancer risk in the Polish population

Studies have demonstrated that changes in DNA methylation of cancer related genes can be an elementary process accounting for ovarian tumorigenesis. Therefore, we evaluated the possible association of single nucleotide polymorphisms (SNPs) of DNA methyltransferases (DNMTs) genes, including DNMT1, DNMT3B, and DNMT3A, with ovarian cancer development in the Polish population. Using PCR–RFLP and HRM analyses, we studied the prevalence of the DNMT1 rs8101626, rs2228611 and rs759920, DNMT3A rs2289195, 7590760, rs13401241, rs749131 and rs1550117, and DNMT3B rs1569686, rs2424913 and rs2424932 SNPs in patients with ovarian cancer (n = 159) and controls (n = 180). The lowest p values of the trend test were observed for the DNMT1 rs2228611 and rs759920 SNPs in patients with ovarian cancer (p _trend = 0.0118 and p _trend = 0.0173, respectively). Moreover, we observed, in the recessive inheritance model, that the DNMT1 rs2228611 and rs759920 SNPs are associated with an increased risk of ovarian cancer development [OR 1.836 (1.143–2.949), p = 0.0114, p _corr = 0.0342, and OR 1.932 (1.185–3.152), p = 0.0078, p _cor=0.0234, respectively]. However, none of other nine studied SNPs displayed significant contribution to the development of ovarian cancer. Furthermore, haplotype and multifactor dimensionality reduction analysis of the studied DNMT1, DNMT3B, and DNMT3A polymorphisms did not reveal either SNP combinations or gene interactions to be associated with the risk of ovarian cancer development. Our results may suggest that DNMT1 variants may be risk factors of ovarian cancer.     

A Colorimetric Method for Detection of K-ras Codon 12 Point Mutations in DNA Extracted from Tissue and Peripheral Blood in Pancreatic Disorders

Molecular-based methods to monitor point mutations require special and expensive equipment unavailable in most hospitals. Colorimetric-based analysis is an ideal platform for K-ras codon 12 gene point mutations because it uses commonly found hospital equipment. The colorimetric assay is sensitive and specific, detecting mutated DNA levels as low as 1% in a wild-type background. Paired genomic DNA extracts of fixed tissue and cellular fractions of peripheral blood are more sensitive and accurate than unpaired samplings. This approach has the potential to improve K-ras point mutation scans as well as to detect micrometastases in circulating tumor cells.     

The Correlation of MGMT Promoter Methylation and Clinicopathological Features in Gastric Cancer: A Systematic Review and Meta-Analysis

The silencing of the tumor suppressor gene O-6-methylguanine-DNA methyltransferase (MGMT) by promoter methylation commonly occurs in human cancers. The relationship between MGMT promoter methylation and gastric cancer (GC) remains inconsistent. This study aimed to evaluate the potential value of MGMT promoter methylation in GC patients. Electronic databases were searched to identify eligible studies. The pooled odds ratio (OR) and the corresponding 95% confidence interval (95% CI) were used to evaluate the effects of MGMT methylation on GC risk and clinicopathological characteristics. In total, 31 eligible studies including 2988 GC patients and 2189 nonmalignant controls were involved in meta-analysis. In the pooled analysis, MGMT promoter methylation was significantly associated with GC risk (OR = 3.34, P < 0.001) and substantial heterogeneity (P < 0.001). Meta-regression and subgroup analyses based on the testing method, sample material and ethnicity failed to explain the sources of heterogeneity. Interestingly, MGMT methylation showed a trend associated with gender, and methylation is lower in males compared with females (OR = 0.76, 95% CI = 0.56–1.03). We did not find a significant association in relation to tumor types, clinical stage, age status or H. pylori status in cancer (all P > 0.1). MGMT promoter methylation may be correlated with the prognosis of GCs in disease free survival (DFS) or overall survival (OS) for univariate analysis. MGMT promoter methylation may play a crucial role in the carcinogenesis and prognosis of GC. MGMT methylation was not correlated with tumor types, clinical stage, age status, H. pylori status. However, the result of the association of MGMT methylation and gender should be considered with caution.     

KRAS mutations in colorectal cancer from Tunisia: relationships with clinicopathologic variables and data on TP53 mutations and microsatellite instability

Mutations in KRAS gene are among the critical transforming alterations occurring during CRC tumorigenesis. Here we screened 51 primary CRC tumors from Tunisia for mutations in KRAS (codons 12 and 13) using PCR-direct sequencing. Our aim was to analyze tumor mutation frequencies and spectra in Tunisian patients with CRC. KRAS status and mutation site/type were than correlated with familial and clinicopathologic variables and data on TP53 mutations and nuclear protein accumulation and microsatellite instability (MSI). A KRAS somatic mutation has been detected in the CRC tumor of 31.5 % (16/51) of the patients. 81.2 % had a single mutation at codon 12 and 23 % had a single mutation at codon 13. The most common single mutation (50 %) was a G>A transition in codon 12 (c.35G>A; p.Gly12Asp). 81.25 % of the KRAS mutations were transitions and 23 % were transversions. All the mutations in codon 13 were a c.38G>A transition, whereas both G>A transitions and G>T and G>C transversions were found in codon 12. The mutation spectrum was different between MSS and MSI-H tumors and more varied mutations have been detected in MSS tumors. Some amino acid changes were detected only in MSS tumors, i.e. p.Gly12Ser, p.Gly12Cys and p.Gly12Ala. Whereas, the KRAS mutation p.Gly13Asp have been detected only in MSI-H. 43.75 % of the patients harboured combined mutations in KRAS and TP53 genes and the tumor of 71.42 % of them showed TP53 overexpression. In conclusion, the frequency and types of KRAS mutations were as reported for non-Tunisian patients. However, no significant associations have been detected between KRAS mutations and clinicopathologic variables and MSI in Tunisian patients as previously reported.     

Comprehensive Biostatistical Analysis of CpG Island Methylator Phenotype in Colorectal Cancer Using a Large Population-Based Sample

Background

The CpG island methylator phenotype (CIMP) is a distinct phenotype associated with microsatellite instability (MSI) and BRAF mutation in colon cancer. Recent investigations have selected 5 promoters (CACNA1G, IGF2, NEUROG1, RUNX3 and SOCS1) as surrogate markers for CIMP-high. However, no study has comprehensively evaluated an expanded set of methylation markers (including these 5 markers) using a large number of tumors, or deciphered the complex clinical and molecular associations with CIMP-high determined by the validated marker panel.

Metholodology/Principal Findings

DNA methylation at 16 CpG islands [the above 5 plus CDKN2A (p16), CHFR, CRABP1, HIC1, IGFBP3, MGMT, MINT1, MINT31, MLH1, p14 (CDKN2A/ARF) and WRN] was quantified in 904 colorectal cancers by real-time PCR (MethyLight). In unsupervised hierarchical clustering analysis, the 5 markers (CACNA1G, IGF2, NEUROG1, RUNX3 and SOCS1), CDKN2A, CRABP1, MINT31, MLH1, p14 and WRN were generally clustered with each other and with MSI and BRAF mutation. KRAS mutation was not clustered with any methylation marker, suggesting its association with a random methylation pattern in CIMP-low tumors. Utilizing the validated CIMP marker panel (including the 5 markers), multivariate logistic regression demonstrated that CIMP-high was independently associated with older age, proximal location, poor differentiation, MSI-high, BRAF mutation, and inversely with LINE-1 hypomethylation and β-catenin (CTNNB1) activation. Mucinous feature, signet ring cells, and p53-negativity were associated with CIMP-high in only univariate analysis. In stratified analyses, the relations of CIMP-high with poor differentiation, KRAS mutation and LINE-1 hypomethylation significantly differed according to MSI status.

Conclusions

Our study provides valuable data for standardization of the use of CIMP-high-specific methylation markers. CIMP-high is independently associated with clinical and key molecular features in colorectal cancer. Our data also suggest that KRAS mutation is related with a random CpG island methylation pattern which may lead to CIMP-low tumors.     

DNA Methylation Combinations in Adjacent Normal Colon Tissue Predict Cancer Recurrence: Evidence from a Clinical Cohort Study

Accumulating evidence has suggested the requirement for further stratification of patients in the same tumor stage according to molecular factors. We evaluate the combination of cancer stage and DNA methylation status as an indicator of the risk of recurrence and mortality among patients with colorectal cancer (CRC). A cohort study of 215 patients with CRC (mean age 64.32 years; 50.5% of men) from Tri-Service General Hospital in Taiwan examined the association between cancer stage and risk of CRC recurrence and mortality. A Cox proportional hazard model was used to analyze patient methylation status and clinical information at study entry, and their associations with CRC recurrence and mortality during follow-up. The advanced stage patients with p16, hMLH1, and MGMT methylation were associated with higher risk of CRC recurrence compared with the local stage patients with unmethylation status in tumor tissues, with adjusted hazard ratios (HRs) (95% confidence interval [CI]) of 9.64 (2.92–31.81), 8.29 (3.40–20.22), and 11.83 (3.49–40.12), respectively. When analyzing normal tissues, we observed similar risk of CRC recurrence with adjusted HRs (95% CI) of 10.85 (4.06–28.96), 9.04 (3.79–21.54), and 12.61 (4.90–32.44), respectively. For combined analyses, the risk of recurrence in the patients in advanced stage with DNA methylation in both normal and tumor tissues, compared with local stage with unmethylation, was increased with adjusted HR (95% CI) of 9.37 (3.36–26.09). In the advanced stage patients, methylation status and tissue subtype were associated with increased risk of 5-year cumulative CRC recurrence (p < 0.001). This study demonstrates that clustering DNA methylation status according to cancer stage and tissue subtype is critical for the assessment of risk of recurrence in CRC patients and also indicated an underlying mechanism.     

Role of RASSF1A promoter methylation in the pathogenesis of hepatocellular carcinoma: a meta-analysis of 21 cohort studies

We carried out the current meta-analysis aiming to comprehensively assess the potential role of RASSF1A aberrant promoter methylation in the pathogenesis of hepatocellular carcinoma (HCC). A range of electronic databases were searched: Web of Science (1945–2013), the Cochrane Library Database (Issue 12, 2013), PubMed (1966–2013), EMBASE (1980–2013), CINAHL (1982–2013) and the Chinese Biomedical Database (CBM) (1982–2013) without language restrictions. Meta-analysis was conducted using the STATA 12.0 software. Crude risk difference (RD) with their 95 % confidence interval (95 % CI) was calculated. In the present meta-analysis, 21 clinical cohort studies with a total of 1,205 HCC patients were included. The results of our meta-analysis illustrated that the frequency of RASSF1A promoter methylation in cancer tissues were significantly higher than those of normal, adjacent and benign tissues (cancer tissues vs. normal tissues: RD = 0.63, 95 % CI 0.53–0.73, P < 0.001; cancer tissues vs. adjacent tissues: RD = 0.43, 95 % CI 0.33–0.53, P < 0.001; cancer tissues vs. benign tissues: RD = 0.48, 95 % CI 038–0.58, P < 0.001; respectively). Further subgroup by ethnicity demonstrated that RASSF1A aberrant promoter methylation was correlated with the pathogenesis of HCC among both Asians and Caucasians (all P < 0.05). The current meta-analysis suggests that RASSF1A aberrant promoter methylation may be implicated in the pathogenesis of HCC. Thus, detection of RASSF1A promoter methylation may be a helpful and valuable biomarker for diagnosis and prognosis of HCC.     

Polymorphisms in Alcohol Metabolism Genes ADH1B and ALDH2, Alcohol Consumption and Colorectal Cancer

Background

Colorectal cancer (CRC) is a leading cause of cancer death worldwide. Epidemiological risk factors for CRC included alcohol intake, which is mainly metabolized to acetaldehyde by alcohol dehydrogenase and further oxidized to acetate by aldehyde dehydrogenase; consequently, the role of genes in the alcohol metabolism pathways is of particular interest. The aim of this study is to analyze the association between SNPs in ADH1B and ALDH2 genes and CRC risk, and also the main effect of alcohol consumption on CRC risk in the study population.

Methodology/Principal Findings

SNPs from ADH1B and ALDH2 genes, included in alcohol metabolism pathway, were genotyped in 1694 CRC cases and 1851 matched controls from the Molecular Epidemiology of Colorectal Cancer study. Information on clinicopathological characteristics, lifestyle and dietary habits were also obtained. Logistic regression and association analysis were conducted. A positive association between alcohol consumption and CRC risk was observed in male participants from the Molecular Epidemiology of Colorectal Cancer study (MECC) study (OR = 1.47; 95%CI = 1.18-1.81). Moreover, the SNPs rs1229984 in ADH1B gene was found to be associated with CRC risk: under the recessive model, the OR was 1.75 for A/A genotype (95%CI = 1.21-2.52; p-value = 0.0025). A path analysis based on structural equation modeling showed a direct effect of ADH1B gene polymorphisms on colorectal carcinogenesis and also an indirect effect mediated through alcohol consumption.

Conclusions/Significance

Genetic polymorphisms in the alcohol metabolism pathways have a potential role in colorectal carcinogenesis, probably due to the differences in the ethanol metabolism and acetaldehyde oxidation of these enzyme variants.     

Chimeric promoter mediates guard cell-specific gene expression in tobacco under water deficit

The engineering of stomatal activity under water deficit through guard cell-specific gene regulation is an effective approach to improve drought tolerance of crops but it requires an appropriate promoter(s) inducible by water deficit in guard cells. We report that a chimeric promoter can induce guard cell-specific gene expression under water deficit. A chimeric promoter, p4xKST82-rd29B, was constructed using a tetramer of the 82 bp guard cell-specific regulatory region of potato KST1 promoter (4xKST82) and Arabidopsis dehydration-responsive rd29B promoter. Transgenic tobacco plants carrying p4xKST82-rd29B:mGFP-GUS exhibited GUS expression in response to water deficit. GUS enzyme activity of p4xKST82-rd29B:mGFP-GUS transgenic plants increased ~300 % by polyethylene glycol treatment compared to that of control plant but not by abscisic acid (ABA), indicating that the p4xKST82-rd29B chimeric promoter can be used to induce the guard cell-specific expression of genes of interest in response to water deficit in an ABA-independent manner.     

Effect of Δ9-stearoyl-ACP-desaturase-C mutants in a high oleic background on soybean seed oil composition

Key message

Two new sources of elevated seed stearic acid were identified and the feasibility of an elevated stearic acid, high oleic acid germplasm was studied.

Abstract

Soybean [Glycine max (L.) Merr.] oil typically contains 2–4 % stearic acid. Oil with at least 20 % stearic acid is desirable because of its improved baking properties and health profile. This study identifies two new sources of high stearic acid and evaluates the interaction of high stearic and oleic acid alleles. TCHM08-1087 and TCHM08-755, high stearic acid ‘Holladay’ mutants, were crossed to FAM94-41-3, a line containing a point mutation in a seed-specific isoform of a Δ9-stearoyl-acyl carrier protein-desaturase (SACPD-C). F₂-derived lines were evaluated for fatty acid content in four field environments. Sequencing of SACPDs in TCHM08-1087 and TCHM08-755 revealed distinct deletions of at least one megabase encompassing SACPD-C in both lines. After genotyping, the additive effect for stearic acid was estimated at +1.8 % for the SACPD-C point mutation and +4.1 % for the SACPD-C deletions. Average stearic acid in lines homozygous for the deletions was 12.2 %. A FAM94-41-3-derived line and TCHM08-1087-11, a selection from TCHM08-1087, were crossed to S09-2902-145, a line containing missense mutations in two fatty acid desaturases (FAD2-1A and FAD2-1B). F₁ plants were grown in a greenhouse and individual F₂ seed were genotyped and phenotyped. No interaction was observed between either FAD2-1A or FAD2-1B and any of the SACPD-C mutant alleles. Seed homozygous mutant for SACPD-C/FAD2-1A/FAD2-1B contained 12.7 % stearic acid and 65.5 % oleic acid while seed homozygous for the SACPD-C deletion and mutant for FAD2-1A and FAD2-1B averaged 10.4 % stearic acid and 75.9 % oleic acid.