癌症中DNA甲基化基因模块筛选

肿瘤的发生受遗传学和表观遗传学修饰的共同影响。DNA甲基化是一种重要的表观遗传修饰,在癌症的发生与发展中起着重大的作用。因此找到癌症的甲基化标记物在癌症的诊断和治疗中具有重大意义。本文利用权重基因共表达网络分析的方法（WGCNA）筛选出甲基化基因模块,并分析模块向量基因,进行功能注释,最后对基因模块进行功能分析,得到DNA甲基化与肿瘤间的关系。结果显示,这些甲基化异常的基因模块与癌症的发生有着显著的关联。同时还发现某些甲基化异常的基因模块与多种癌症的发生都有着显著的关联。     

DGGE screening of PKD1 gene reveals novel mutations in a large cohort of 146 unrelated patients

Autosomal dominant polycystic kidney disease (ADPKD) is one of the most commonly inherited renal diseases. ADPKD is a genetically heterogeneous disorder involving at least three different genes. PKD1, the major locus mapped to chromosome 16p13.3 accounts for approximately 85% of ADPKD cases. The search for mutations is a very important step in understanding the molecular mechanisms underlying ADPKD. Despite intense screening by many groups, only a small number of mutations have been described so far. We undertook the first study using denaturing gradient gel electrophoresis (DGGE) to scan for mutations in the non-duplicated region of the PKD1 gene in a large cohort of 146 French unrelated ADPKD patients. We successfully identified novel mutations: 3 are frameshift mutations, 2 nonsense mutations, 2 missense mutations, 1 is an insertion in the frame of 9 nucleotides, 3 intronic variations and several polymorphisms. One of these mutations is the fourth de novo mutation described in this gene. We also describe a family with possible clinical anticipation. DGGE is an effective method for detecting nucleotide changes in the PKD1 gene.     

Germline mutations in the MYH gene in Swedish familial and sporadic colorectal cancer

Biallelic germline mutations in the base excision repair gene MYH have been shown to predispose to a proportion of multiple colorectal adenomas and cancer. To evaluate the contribution of MYH mutations to non- FAP, non-HNPCC familial colorectal cancer, 84 unrelated Swedish individuals affected with colorectal cancer from such families were screened for germline mutations in the coding sequence of the gene. None of the cases was found to carry any pathogenic sequence change. We then determined the prevalence of the two most common pathogenic MYH mutations found in Caucasians, Y165C and G382D, in 450 Swedish sporadic colorectal cancer cases and 480 Swedish healthy controls. The frequency of both variants in Swedish cases and controls was similar to those previously reported. In addition, we found that previously unknown sequence variations at the position of amino acid 423 (R423Q, R423P, and R423R) appear to occur more frequently in cases than in controls (p = 0.02), a finding that warrants future studies.     

Performance of a new, liquid-based cervical screening technique in the clinical setting of a large French laboratory

OBJECTIVE: To compare the performance of liquid-based cytology with the CYTO-screen System (SEROA) with that of conventional smears through a secondary analysis of a large database covering the activity of an independent French laboratory during the period 1998-2002. STUDY DESIGN: The study was performed with a retrospective, comparative, historical design on 2 subgroups of women having been screened by gynecologists who switched from conventional smears to the CYTO-screen System in the period 1998-2002. The first cohort population consisted of women who had at least 4 subsequent screening tests over the period with half conventional and half with the CYTO-screen System. A control group consisted of smears collected by gynecologists who fully maintained activity with a conventional method over the same period. The second group consisted of women who had their first screening test performed over the study period by gynecologists who modified their technique. Specimen adequacy, endocervical cell content and epithelial cell abnormality detection rates were compared between the groups. RESULTS: As compared with the conventional smear, the CYTO-screen System showed a reduction in unsatisfactory reports, especially in the second group of first-screened (0.14% versus 1.3%, P < .0001). The rate of atypical squamous cells of undetermined significance increased significantly after the switch to the CYTO-screen System (2.5% versus 1.3%, P = .004) and in the second group of first-screened women (2.05% versus 1.4%, P = .0014), with higher histologic confirmation in both situations. There was a non-significant increase in the detection rates of low and high grade squamous intraepithelial lesions after the switch to the CYTO-screen System and in the second group of first-screened women. CONCLUSION: The CYTO-screen System gives higher-quality specimens and has a higher detection rate for squamous intraepithelial lesions, but that rate was significant only for atypical squamous cells of undetermined significance.     

Analysis of Loss of heterozygosity and immunohistochemistry in BRCA1 gene in sporadic breast cancers

BRCA1 is a tumour suppressor gene (TSG), which predisposes cancer to both breast and ovary. The primary objective of the present study is to ascertain the involvement of BRCA1 gene in the pathogenesis of sporadic breast cancer women in Chennai (South India) by analysing its protein expression by immunohistochemistry (IHC) and loss of heterozygosity (LOH) for confirmation of the involvement of TSG in the study population. We found down regulation of BRCA1 protein (54%) in IHC and it was correlated with the clinicopathological parameters of the patients. We found near significant correlation (P < 0.063) between BRCA1 protein expression and clinicopathological parameters. We found 30% LOH in our study and it was also correlated with the clinicopathological parameters. No correlation was found between LOH and clinicopathological parameters. Though we found no correlation, the results revealed in this study support the involvement of BRCA1 TSG in the pathogenesis of sporadic breast cancer women in Chennai (South India).     

Evaluation of a colorectal cancer screening program composed of successive waves of different tests: The experience of the French Calvados County

BackgroundsThe value of colorectal cancer (CRC) screening program in a population with a limited participation rate is debated. This study assesses the real-life performances of different screening tests in a population benefiting from an organized program and included in a cancer registry.MethodsPatients who participated in at least one screening campaign between 2004 and 2016 were included. Four screening procedures were used: Hemoccult II, Magstream, Hemoccult and Magstream combined, and OC Sensor. Data were crossed with the Digestive Cancer Registry of Calvados to detect CRCs diagnosed during this period. The main outcomes were CRC detection and the incidence rate of interval cancers.ResultsScreening consisted of 325,083 tests in 134,498 patients. Of the 2580 CRCs detected in patients aged 50–74, 534 (20.7 %) were screen-detected. OC Sensor had the highest sensitivity for CRC detection (83.7 %, 95 % CI [76.8–89.1 %]) and the lowest interval cancer rate (2.0 per 10,000 person-years, 95 % CI [1.4–2.7]) compared with other screening tests, excluding combinations. The overall participation rate was 28.9 %.ConclusionReal-life differences in performance between different screening tests exist, and OC Sensor appears to be the best. The low participation rate suggests that the rate of screen-detected CRC could be higher.     

Hirschsprung disease in a large birth cohort

The incidence of Hirschsprung disease was studied in a series of almost 700,000 consecutive livebirths in British Columbia from 1964-1982, by means of the records of a health surveillance registry that uses multiple sources of ascertainment. The estimated liveborn incidence rate for Hirschsprung disease was 1 in 4,417 livebirths (156 cases out of 689,118 livebirths). Data pertaining to sex ratio, additional anomalies, recurrence, and mortality were also analyzed over the caseload period 1952 to 1983. A total of 29.8% of cases had some additional anomaly--the majority being nonregional anomalies in other systems or more distantly in the gastrointestinal tract. Cardiovascular and gastrointestinal anomalies not a direct consequence of Hirschsprung disease were the most frequent additional anomalies found, occurring in 10 and 12 of 178 cases, respectively. Sensorineural anomalies were also frequent, occurring in 12 of 178 cases. Clinical implications arising from the study regarding the neonatal assessment of infants with these anomalies are discussed.     

Retrospective study of the cystic fibrosis transmembrane conductance regulator (CFTR) gene mutations in Guthrie cards from a large cohort of neonatal screening for cystic fibrosis

The cystic fibrosis transmembrane conductance regulator (CFTR) gene encodes a cAMP-activated chloride channel, and in individuals with both alleles of the gene mutated, symptoms of CF disease are manifest. With more than 300 mutations so far described in the gene the profile of mutant alleles in a population is specific to its ethnic origin. For an analysis with an unbiased recruitment of the CF alleles in neonates of similar origin (Normandy, France), we have retrospectively analyzed the Guthrie cards of affected newborns, diagnosed by the immunoreactive trypsinogen (IRT) assay. Analysis of the 27 exons of the CFTR gene using a GC clamp denaturing gradient gel electrophoresis (DGGE) assay has enabled us to identify over 96% of the mutated alleles. Two of these were novel mutations. We would like to propose this strategy as an efficient method of retrospective molecular genetic diagnosis that can be performed wherever Guthrie cards can be obtained. Knowledge of rare alleles could be a prerequisite for CF therapy in the future.     

天津地区人群hMLH1和hMSH2基因多态性与散发性结直肠癌易感性的关系

为探讨错配修复基因hMLH1和hMSH2单核苷酸多态性(Single nucleotide polymorphism,SNP)与散发性结直肠癌(Sporadic colorectal caner,SCRC)发病易感性之间的关系,文章采用聚合酶链式反应-变性高效液相色谱方法和序列分析技术,检测了天津地区600例SCRC患者和600例健康对照个体hMLH1394G/C、hMSH2943-1G/A、hMSH21917T/G和hMSH22783C/A的基因型频率分布。结果显示:SCRC患者组hMSH22783C/A3种基因型C/C、C/A、A/A频率(90%、9%、1%)与对照组(95%、4.8%、0.2%)相比差异具有统计学意义(χ2=11.91,P0.01)。与hMSH22783C/C基因型相比,C/A和A/A基因型能增加SCRC发病风险(OR值分别为1.77和11.94,95%CI分别为1.03～3.03和1.38～103.2)。多态性位点联合分析显示,SCRC组与对照组单倍型分布差异有统计学意义(χ2=38.38,P0.01);与394G/943-1G/2783C单倍型相比,394G/943-1G/2783A单倍型显著增加SCRC的发病风险(OR=2.18,95%CI:1.40～3.40)。结果提示hMSH22783C/A多态性可能成为预测SCRC发病风险的独立因素,394G/943-1G/2783A单倍型可能增加SCRC的发病风险。     

The rabbit beta-globin gene contains a large large insert in the coding sequence

We have used the rabbit β-globin DNA plasmid PβG1 (Maniatis et al., 1976) labeled with ³²P as a filter hybridization probe for DNA fragments containing the β-globin gene in restriction endonuclease digests of rabbit liver DNA. The β-globin DNA fragments we detect appear to contain the gene, present in PβG1 DNA, which codes for adult rabbit β-globin. These fragments have been ordered into a physical map of cleavage sites within and neighboring the structural gene in the rabbit genome (Jeffreys and Flavell, 1977). A detailed analysis of β-globin DNA fragments produced by cleavage with restriction endonucleases which are known to cut the β-globin gene has now shown that the β-globin structural gene is not contiguous in rabbit liver DNA, but is interrupted by a 600 base pair DNA segment inserted somewhere within the coding sequence for amino acid residues 101–120 of the 146 residue β-globin chain. Otherwise, the map of cleavage sites within the gene is co-linear with that deduced from the sequence of rabbit β-globin messenger RNA. Preliminary analysis indicates that this insert is also present in the β-globin gene in rabbit brain, kidney, spleen, bone marrow and sperm, and in erythroid cells isolated from the marrow of an anemic rabbit. The insert appears, therefore, to be a general property of the rabbit β-globin gene, even in tissues in which this gene is active, which suggests that the insert is not involved in inactivating the gene in nonerythroid tissues.     

The effect of polymorphisms in the enhancer of split gene complex on bristle number variation in a large wild-caught cohort of Drosophila melanogaster

The Enhancer of split complex [E(spl)-C] in Drosophila encompasses a variety of functional elements controlling bristle patterning and on the basis of prior work is a strong candidate for harboring alleles having subtle effects on bristle number variation. Here we extend earlier studies identifying associations between complex phenotypes and polymorphisms segregating among inbred laboratory lines of Drosophila and test the influence of E(spl)-C on bristle number variation in a natural cohort. We describe results from an association mapping study using 203 polymorphisms spread throughout the E(spl)-C genotyped in 2000 wild-caught Drosophila melanogaster. Despite power to detect associations accounting for as little as 2% of segregating variation for bristle number, and saturating the region with single-nucleotide polymorphisms (SNPs), we identified no single SNP marker showing a significant (additive over loci) effect after correcting for multiple tests. Using a newly developed test we conservatively identify six regions of the E(spl)-C in which the insertion of transposable elements as a class contributes to variation in bristle number, apparently in a sex- or trait-limited fashion. Finally, we carry out all possible 20,503 two-way tests for epistasis and identify a slight excess of marginally significant interactions, although none survive multiple-testing correction. It may not be straightforward to extend the results of laboratory-based association studies to natural populations.     

Genetic variation screening of TNNT2 gene in a cohort of patients with hypertrophic and dilated cardiomyopathy

Mutations in troponin T (TNNT2) gene represent the important part of currently identified disease-causing mutations in hypertrophic (HCM) and dilated (DCM) cardiomyopathy. The aim of this study was to analyze TNNT2 gene exons in patients with HCM and DCM diagnosis to improve diagnostic and genetic consultancy in affected families. All 15 exons and their flanking regions of the TNNT2 gene were analyzed by DNA sequence analysis in 174 patients with HCM and DCM diagnosis. We identified genetic variations in TNNT2 exon regions in 56 patients and genetic variations in TNNT2 intron regions in 164 patients. Two patients were found to carry unique mutations in the TNNT2 gene. Limited genetic screening analysis is not suitable for routine testing of disease-causing mutations in patients with HCM and DCM as only individual mutation-positive cases may be identified. Therefore, this approach cannot be recommended for daily clinical practice even though, due to financial constraints, it currently represents the only available strategy in a majority of cardio-centers.     

Toward a comprehensive and systematic methylome signature in colorectal cancers

CpG Island Methylator Phenotype (CIMP) is one of the underlying mechanisms in colorectal cancer (CRC). This study aimed to define a methylome signature in CRC through a methylation microarray analysis and a compilation of promising CIMP markers from the literature. Illumina HumanMethylation27 (IHM27) array data was generated and analyzed based on statistical differences in methylation data (1st approach) or based on overall differences in methylation percentages using lower 95% CI (2nd approach). Pyrosequencing was performed for the validation of nine genes. A meta-analysis was used to identify CIMP and non-CIMP markers that were hypermethylated in CRC but did not yet make it to the CIMP genes’ list. Our 1st approach for array data analysis demonstrated the limitations in selecting genes for further validation, highlighting the need for the 2nd bioinformatics approach to adequately select genes with differential aberrant methylation. A more comprehensive list, which included non-CIMP genes, such as APC, EVL, CD109, PTEN, TWIST1, DCC, PTPRD, SFRP1, ICAM5, RASSF1A, EYA4, 30ST2, LAMA1, KCNQ5, ADHEF1, and TFPI2, was established. Array data are useful to categorize and cluster colonic lesions based on their global methylation profiles; however, its usefulness in identifying robust methylation markers is limited and rely on the data analysis method. We have identified 16 non-CIMP-panel genes for which we provide rationale for inclusion in a more comprehensive characterization of CIMP+ CRCs. The identification of a definitive list for methylome specific genes in CRC will contribute to better clinical management of CRC patients.     

The cost-effectiveness of screening for colorectal cancer

Background

Published decision analyses show that screening for colorectal cancer is cost-effective. However, because of the number of tests available, the optimal screening strategy in Canada is unknown. We estimated the incremental cost-effectiveness of 10 strategies for colorectal cancer screening, as well as no screening, incorporating quality of life, noncompliance and data on the costs and benefits of chemotherapy.

Methods

We used a probabilistic Markov model to estimate the costs and quality-adjusted life expectancy of 50-year-old average-risk Canadians without screening and with screening by each test. We populated the model with data from the published literature. We calculated costs from the perspective of a third-party payer, with inflation to 2007 Canadian dollars.

Results

Of the 10 strategies considered, we focused on three tests currently being used for population screening in some Canadian provinces: low-sensitivity guaiac fecal occult blood test, performed annually; fecal immunochemical test, performed annually; and colonoscopy, performed every 10 years. These strategies reduced the incidence of colorectal cancer by 44%, 65% and 81%, and mortality by 55%, 74% and 83%, respectively, compared with no screening. These strategies generated incremental cost-effectiveness ratios of $9159, $611 and $6133 per quality-adjusted life year, respectively. The findings were robust to probabilistic sensitivity analysis. Colonoscopy every 10 years yielded the greatest net health benefit.

Interpretation

Screening for colorectal cancer is cost-effective over conventional levels of willingness to pay. Annual high-sensitivity fecal occult blood testing, such as a fecal immunochemical test, or colonoscopy every 10 years offer the best value for the money in Canada.Colorectal cancer is the fourth most common cancer diagnosed in North America and the second leading cause of cancer death.^1,2 An effective population-based screening program is likely to decrease mortality associated with colorectal cancer^3–6 through earlier detection and to decrease incidence by allowing removal of precursor colorectal adenomas.^7,8 Professional societies and government-sponsored committees have released guidelines for screening of average-risk individuals for colorectal cancer by means of several testing options.^9–12 These tests vary in sensitivity, specificity, risk, costs and availability. With no published studies designed to directly compare screening strategies, decision analysis is a useful technique for examining the relative cost-effectiveness of these strategies.^13–21 Previous studies have shown that screening for colorectal cancer is cost-effective at conventional levels of willingness to pay, but no single strategy has emerged as clinically superior or economically dominant.²² The interpretations of economic evaluations in this area have been limited because investigators have not simultaneously accounted for the positive effects of screening on quality of life, the effect of noncompliance with screening schedules, and the greater efficacy and cost of more modern chemotherapy regimens for colorectal cancer. Furthermore, no study has included all of the strategies recommended in the 2008 guidelines of the US Multi-Society Task Force on Colorectal Cancer.¹⁰Our objective was to estimate the incremental cost-effectiveness of 10 strategies for colorectal cancer screening, as well as the absence of a screening program. The current study is more complete than earlier studies because we included information on quality of life, noncompliance with screening and the efficacy observed in recent randomized trials of colorectal cancer treatments. The complete model is available in Appendix 1 (available at www.cmaj.ca/cgi/content/full/cmaj.090845/DC1). This article focuses on the comparison of no screening and three screening strategies:¹ low-sensitivity guaiac fecal occult blood test,² performed annually; fecal immunochemical test,³ performed annually; and colonoscopy, performed every 10 years. These three tests are currently being used or considered for population-based screening of average-risk individuals in some Canadian provinces.     

Plasma lipid profiling in a large population-based cohort

We have performed plasma lipid profiling using liquid chromatography electrospray ionization tandem mass spectrometry on a population cohort of more than 1,000 individuals. From 10 μl of plasma we were able to acquire comparative measures of 312 lipids across 23 lipid classes and subclasses including sphingolipids, phospholipids, glycerolipids, and cholesterol esters (CEs) in 20 min. Using linear and logistic regression, we identified statistically significant associations of lipid classes, subclasses, and individual lipid species with anthropometric and physiological measures. In addition to the expected associations of CEs and triacylglycerol with age, sex, and body mass index (BMI), ceramide was significantly higher in males and was independently associated with age and BMI. Associations were also observed for sphingomyelin with age but this lipid subclass was lower in males. Lysophospholipids were associated with age and higher in males, but showed a strong negative association with BMI. Many of these lipids have previously been associated with chronic diseases including cardiovascular disease and may mediate the interactions of age, sex, and obesity with disease risk.     

A survey of tandem repeat instabilities and associated gene expression changes in 35 colorectal cancers

Background

Colorectal cancer is a major contributor to cancer morbidity and mortality. Tandem repeat instability and its effect on cancer phenotypes remain so far poorly studied on a genome-wide scale.

Results

Here we analyze the genomes of 35 colorectal tumors and their matched normal (healthy) tissues for two types of tandem repeat instability, de-novo repeat gain or loss and repeat copy number variation. Specifically, we study for the first time genome-wide repeat instability in the promoters and exons of 18,439 genes, and examine the association of repeat instability with genome-scale gene expression levels. We find that tumors with a microsatellite instable (MSI) phenotype are enriched in genes with repeat instability, and that tumor genomes have significantly more genes with repeat instability compared to healthy tissues. Genes in tumor genomes with repeat instability in their promoters are significantly less expressed and show slightly higher levels of methylation. Genes in well-studied cancer-associated signaling pathways also contain significantly more unstable repeats in tumor genomes. Genes with such unstable repeats in the tumor-suppressor p53 pathway have lower expression levels, whereas genes with repeat instability in the MAPK and Wnt signaling pathways are expressed at higher levels, consistent with the oncogenic role they play in cancer.

Conclusions

Our results suggest that repeat instability in gene promoters and associated differential gene expression may play an important role in colorectal tumors, which is a first step towards the development of more effective molecular diagnostic approaches centered on repeat instability.

Electronic supplementary material

The online version of this article (doi:10.1186/s12864-015-1902-9) contains supplementary material, which is available to authorized users.