肺癌组织中错配修复基因hMLH1 启动子甲基化状态分析

目的 探讨肺癌组织中错配修复基因ｈＭＬＨ１启动子甲基化状态及其与蛋白表达的关系。方法 用ＨｐａⅡ酶切ＰＣＲ及ＳＰ免疫组织化学方法,分析５０例肺癌标本ｈＭＬＨ１启动子甲基化状态及蛋白表达。结果 ｈＭＬＨ１启动子甲基化有１６例,占３２．０％（１６／５０）。ｈＭＬＨ１启动甲基化与性别、吸烟状态及临床病理无明显的相关性。ｈＭＬＨ１蛋白表达阴性的１４例中,１１例有甲基化（７８．６％）;而３２例蛋白表达阳性者中,只有５例甲基化（１５．６％）。结论 ｈＭＬＨ１启动子区在肺癌组织中有中等频率的甲基化,是ｈＭＬＨ１蛋白表达降低的主要原因之一。     

Clinical significance of RECK promoter methylation in pancreatic ductal adenocarcinoma

The aim of this study was to analyze the clinical significance of reversion-inducing-cysteine-rich protein with Kazal motifs (RECK) promoter methylation in pancreatic ductal adenocarcinoma (PDA). Methylation-specific polymerase chain reaction was used to examine the promoter methylation status of RECK in 60 pairs of PDA tissue samples and adjacent non-cancerous tissue samples. Statistical analyses were applied to test the associations between RECK promoter methylation status, clinicopathologic factors, and prognosis. The rate of RECK promoter methylation was significantly higher in PDA tissues than in adjacent non-cancerous tissues (P?<?0.001). RECK methylation status was significantly associated with clinical stage (P?=?0.017), histological differentiation (P?=?0.046), and lymph node metastasis (P?=?0.003), but was not associated with gender, age, and tumor location (all P?>?0.05). Additionally, RECK promoter methylation is associated with malignant behavior and poor prognosis. In conclusion, determination of RECK promoter methylation status in tumor tissues may assist in the identification of patients who require aggressive postoperative intervention in order to improve prognosis.     

妇科肿瘤与APC基因启动子甲基化研究进展

腺瘤性结肠息肉病（APC）基因是Wnt信号转导通路重要的抑癌基因,其启动子区域的异常甲基化可影响APC基因mRNA的转录和APC蛋白的表达,导致Wnt信号转导通路异常。近年来在妇科肿瘤的多项研究中发现了APC基因启动子区域异常甲基化,其与肿瘤的发生、发展有关。现就APC基因启动子甲基化与妇科肿瘤的研究进展进行综述。     

Expression and promoter methylation analysis of ATP-binding cassette genes in pancreatic cancer

We investigated the relationship of ABCB1/MDR1, ABCC1/MRP1 and ABCG2/BCRP expression and promoter methylation with pancreatic cancer tumorigenesis and drug resistance. Gemcitabine-resistant pancreatic cancer cells, SW1990/GZ (33.3-fold increased resistance), were obtained by treating SW1990 cells with gemcitabine. The expression of ABCB1/MDR1, ABCC1/MRP1 and ABCG2/BCRP was determined by quantitative real-time RT-PCR in the cell lines, 3 normal pancreatic tissues, 15 human pancreatic cancer samples and 15 adjacent tissues. Promoter methylation was determined in cell lines by bisulfite genomic sequencing. ABCB1/MDR1, ABCC1/MRP1 and ABCG2/BCRP were upregulated in SW1990 and SW1990/GZ compared with normal pancreatic tissue, and expression in SW1990/GZ was significantly higher than in SW1990 cells. ABCB1/MDR1, ABCC1/MRP1 and ABCG2/BCRP were upregulated in pancreatic cancer tissues, compared to adjacent tissues. The ABCB1/MDR1, ABCC1/MRP1 and ABCG2/BCRP promoter were hypomethylated in all the cell lines. ABCB1/MDR1, ABCC1/MRP1 and ABCG2/BCRP expression correlated with pancreatic cancer tumorigenesis and drug resistance in a mechanism that is independent of promoter methylation.     

乳腺癌患者血浆循环DNA中Sox17基因甲基化检测的临床意义

背景与目的：在乳腺癌发生、发展过程中,甲基化异常是导致抑癌基因失活的重要机制,是一种可用于肿瘤诊断及预后判断、有价值的生物标志物。本研究旨在通过检测乳腺癌组织及其相应的血浆循环DNA中Sox17基因的甲基化状况,探讨其在乳腺癌早期诊断和预后判断方面的应用价值。方法：采用甲基化特异性聚合酶链反应(methylation specific-PCR,MSP)法,对86例乳腺癌组织、36例乳腺良性肿瘤的癌旁正常组织及其配对的血浆循环DNA中Sox17基因启动子甲基化进行检测,并结合乳腺癌的主要临床病理特性进行分析。结果：86例乳腺癌组织中Sox17基因启动子的甲基化率为77.9%(67/86),与其相应血浆循环DNA中Sox17基因启动子的甲基化率为61.6%(53/86),36例癌旁正常乳腺组织及血浆中均未检测到Sox17基因异常甲基化。患者血浆循环DNA中Sox17基因启动子的甲基化与肿瘤组织中该基因的甲基化显著相关(r=0.502,P=0.000)。在乳腺癌组织标本中Sox17基因甲基化率与患者肿瘤分期(χ2=6.18,P=0.041)、淋巴结转移(χ2=13.54,P=0.001)显著相关,在血浆标本中,Sox17基因甲基化率与患者肿瘤分期(χ2=27.06,P=0.000)、肿瘤大小(χ2=9.65,P=0.007)及淋巴结转移(χ2=20.80,P=0.000)显著相关,与患者年龄、组织学分级及ER、PR、HER-2/neu等指标差异无统计学意义(P>0.05)。结论：Sox17基因启动子甲基化在乳腺癌的发生、发展中起着重要作用,可能与乳腺癌的预后相关。血浆中Sox17基因甲基化,是一个有潜在应用价值的生物标志物。     

GSTPl基因启动子甲基化与前列腺癌的相关性研究

背景与目的：谷胱甘S-转移酶P1(glutathione S-transferase P1,GSTP1)保护细胞避免DNA损伤和癌细胞形成,抑制GSTP1活性可以导致DNA损伤的敏感性增强和癌变发生的概率增加。该研究旨在研究前列腺癌组织中GSTPl基因甲基化与前列腺癌临床病理特征的关系。方法：收集2015年4月—2016年12月在海南省中医院和海口市人民医院住院的46例前列腺癌患者的前列腺癌组织及对应的包埋癌旁组织,应用实时荧光定量聚合酶链反应(real-time fluorescent quantitative polymerase chain reaction,RTFQ-PCR)检测GSTPl mRNA水平,通过甲基化特异性聚合酶链反应(methylation-specific polymerase chain reaction,MSP)检测GSTP1基因启动子区域CpG岛的甲基化表达水平,然后与性别、年龄及肿瘤TNM分期等临床数据进行关联分析。结果：与癌旁组织比较,前列腺癌组织中GSTP1 mRNA表达水平降低(P<0.01),并且GSTP1 mRNA降低与GSTPl基因甲基化呈负相关(P<0.05);前列腺癌和癌旁组织中甲基化阳性率分别为66.0%和23.5%,差异有统计学意义(P<0.05);GSTP1基因的启动子甲基化频率与肿瘤分期显著相关(r＝073,P<0.05),而与其他临床特征无明显相关性(P>0.05)。结论：GSTP1基因启动子甲基化可能造成GSTP1基因低表达,与前列腺癌发病明显相关,有望成为检测及诊断前列腺癌的新方法。     

卵巢癌中 CDK10基因启动子甲基化检测的临床意义

目的：分析卵巢癌中受体 O 型蛋白酪氨酸磷酸酶（CDK10）基因启动子甲基化状态及临床意义。方法：应用甲基化特异性 PCR 检测 CDK10基因在50例卵巢癌组织和27例正常卵巢组织中的甲基化状态并结合临床病理资料进行分析。结果：在卵巢癌组织中 CDK10甲基化阳性率为60．0％（30／50）,正常卵巢组织未出现 CDK10甲基化;CDK10甲基化与肿瘤临床分期、病理分型与 Ki －67表达相关（P ＜0．05）。结论：CDK10基因甲基化可能是参与卵巢癌发展的重要分子事件。     

Quantitative analysis of promoter methylation of the EDNRB gene in gastric cancer

Hypermethylation has been shown in the promoter region of the endothelin receptor B (EDNRB) gene in several human tumors. However, its role in gastric cancer formation is still unclear. In this study, the methylation status of the EDNRB gene in paired gastric cancer tissues and adjacent normal tissues from 96 patients was detected quantitatively using pyrosequencing. The results showed the methylation of promoter of EDNRB gene in gastric cancer (50.42 ± 9.03%) was significantly higher than in adjacent normal tissues (6.47 ± 2.98%) (P < 0.01). Among 96 tumor tissues, promoter hypermethylation of the EDNRB gene was correlated with tumor infiltration (T1: 47.4 ± 7.31% T2:48.2 ± 9.17% T3:52.9 ± 6.48% T4:53.2 ± 10.45%), lymph node metastasis (N0:45.4 ± 6.99% N1:49.0 ± 9.10% N2:52.0 ± 8.40% N3:53.7 ± 9.92%), and distant metastasis (M0:48.9 ± 6.99% M1:53.9 ± 11.98%) (P < 0.05), but it was not associated with other clinicopathological characteristics. In addition, the treatment of the human gastric cancer cell line, SGC-7901, with demethylation agent can restore the expression of EDNRB. Our results suggest that promoter hypermethylation of EDNRB gene is highly prevalent in gastric cancer, which may play a role in the pathogenesis of gastric cancer. Futhermore, hypermethylation of EDNRB gene was remarkably related to infiltration and metastasis of gastric cancer and may attribute to the tumor progression.     

卵巢癌中 ECRG4基因启动子甲基化的研究

目的：探讨人食管癌相关基因4（esophageal cancer related gene 4,ECRG4）基因启动子区域在卵巢癌组织中的甲基化状态及临床意义。方法：应用甲基化特异性 PCR 检测 ECRG4基因启动子在50例卵巢癌组织和10例正常卵巢组织中的甲基化状态并结合临床病理资料进行分析。结果：在卵巢癌组织中 ECRG4甲基化阳性率为36%（18/50）,正常卵巢组织未出现 ECRG4甲基化。ECRG4甲基化与肿瘤淋巴结转移及 Ki -67表达密切相关（P <0.05）。结论：ECRG4甲基化可能是参与卵巢癌发展的重要分子事件。     

NPTX2基因DNA甲基化与胰腺癌的研究进展

神经元正五聚蛋白Ⅱ（NPTX2)属于神经元正五聚体蛋白家族(NPs)的成员之一,参与细胞周期调控、细胞增殖、凋亡和致癌等生物学过程。近年来研究发现,在胰腺癌中DNA甲基化普遍存在,导致一些抑癌基因高甲基化而失活,NPTX2作为胰腺癌的一种抑癌基因,在胰腺癌癌前病变阶段就出现明显高甲基化。DNA甲基化可能是导致胰腺癌中NPTX2蛋白功能失活,表达沉默的机制之一。而DNA 甲基化是肿瘤的早期、多发、可逆事件,有望作为胰腺癌早期诊断的潜在标志物以及治疗靶点。文章对NPTX2 的 DNA 甲基化在胰腺癌中的最新研究进展做一综述。     

食管癌组织中hMSH2基因启动子区甲基化检测

目的检测食管癌组织中hMSH2基因启动子区甲基化状态,探讨与食管癌发生、发展的关系。方法32例食管癌患者术前均未接受放疗、化疗等其他治疗,手术切除后30min内,在癌组织及正常食管黏膜组织各取约1.0cm×1.0cm×1.0cm大小的组织块标本,立即储存于-80℃冰箱中,冻存备提DNA。采用甲基化特异PCR法(MSP法)检测食管癌及正常食管黏膜组织中错配修复基因hMSH2启动子区甲基化的表达。结果32例食管癌组织中,hMSH2启动子区甲基化发生率为34.4%(11/32),正常食管黏膜组织未发现甲基化,两组甲基化阳性率相比较,差异有统计学意义(P<0.01)。高龄患者(≥70岁)癌组织中启动子区hMSH2甲基化发生率(85.7%)明显高于较低龄患者(<70岁,20.0%;P<0.05)。病理组织学Ⅲ~Ⅳ级食管癌组织中,hMSH2启动子区甲基化发生率(70.0%)明显高于Ⅰ~Ⅱ级(18.2%,P<0.05)。结论食管癌组织中,hMSH2基因启动子区甲基化与年龄和病理组织学分级可能有关。     

PTPRO基因启动子甲基化与卵巢癌淋巴结转移的相关性

目的:分析卵巢癌中受体O型蛋白酪氨酸磷酸酶（PTPRO）基因启动子甲基化状态及临床意义。方法:应用甲基化特异性PCR检测PTPRO基因在50例卵巢癌组织和27例正常卵巢组织中的甲基化状态并结合临床病理资料进行分析。结果:在卵巢癌组织中PTPRO甲基化阳性率为66%（33/50）,正常卵巢组织未出现PTPRO甲基化;PTPRO甲基化与肿瘤淋巴结转移与FIGO分期密切相关（P＜0.05）。结论:PTPRO基因甲基化可能是参与卵巢癌发展的重要分子事件。     

DNA mismatch repair gene polymorphisms affect survival in pancreatic cancer

Purpose.

DNA mismatch repair (MMR) maintains genomic stability and mediates cellular response to DNA damage. We aim to demonstrate whether MMR genetic variants affect overall survival (OS) in pancreatic cancer.

Materials and Methods.

Using the Sequenom method in genomic DNA, we retrospectively genotyped 102 single-nucleotide polymorphisms (SNPs) of 13 MMR genes from 706 patients with pancreatic adenocarcinoma seen at The University of Texas MD Anderson Cancer Center. Association between genotype and OS was evaluated using multivariable Cox proportional hazard regression models.

Results.

At a false discovery rate of 1% (p ≤ .0015), 15 SNPs of EXO1, MLH1, MSH2, MSH3, MSH6, PMS2, PMS2L3, TP73, and TREX1 in patients with localized disease (n = 333) and 6 SNPs of MSH3, MSH6, and TP73 in patients with locally advanced or metastatic disease (n = 373) were significantly associated with OS. In multivariable Cox proportional hazard regression models, SNPs of EXO1, MSH2, MSH3, PMS2L3, and TP73 in patients with localized disease, MSH2, MSH3, MSH6, and TP73 in patients with locally advanced or metastatic disease, and EXO1, MGMT, MSH2, MSH3, MSH6, PMS2L3, and TP73 in all patients remained significant predictors for OS (p ≤ .0015) after adjusting for all clinical predictors and all SNPs with p ≤ .0015 in single-locus analysis. Sixteen haplotypes of EXO1, MLH1, MSH2, MSH3, MSH6, PMS2, PMS2L3, RECQL, TP73, and TREX1 significantly correlated with OS in all patients (p ≤ .001).

Conclusion.

MMR gene variants may have potential value as prognostic markers for OS in pancreatic cancer patients.     

Predicting gene promoter methylation in non-small-cell lung cancer by evaluating sputum and serum

The use of 5-methylcytosine demethylating agents in conjunction with inhibitors of histone deacetylation may offer a new therapeutic strategy for lung cancer. Monitoring the efficacy of gene demethylating treatment directly within the tumour may be difficult due to tumour location. This study determined the positive and negative predictive values of sputum and serum for detecting gene methylation in primary lung cancer. A panel of eight genes was evaluated by comparing methylation detected in the primary tumour biopsy to serum and sputum obtained from 72 patients with Stage III lung cancer. The prevalence for methylation of the eight genes in sputum (21-43%) approximated to that seen in tumours, but was 0.7-4.3-fold greater than detected in serum. Sputum was superior to serum in classifying the methylation status of genes in the tumour biopsy. The positive predictive value of the top four genes (p16, DAPK, PAX5 beta, and GATA5) was 44-72% with a negative predictive value for these genes > or =70%. The highest specificity was seen for the p16 gene, and this was associated with a odds ratio of six for methylation in the tumour when this gene was methylated in sputum. In contrast, for serum, the individual sensitivity for all genes was 6-27%. Evaluating the combined effect of methylation of at least one of the four most significant genes in sputum increased the positive predictive value to 86%. These studies demonstrate that sputum can be used effectively as a surrogate for tumour tissue to predict the methylation status of advanced lung cancer where biopsy is not feasible.     

MSP法检测胃癌组织中RASSF1A基因启动子区甲基化改变

目的：探讨抑癌基因RASSF1A启动子区CpG岛甲基化与胃癌及临床病理特征的关系。方法：采用甲基化特异性PCR（methylation—specific PCR,MSP）法检测60例胃癌组织及相应癌旁组织和30例对照组织中RASSF1A基因启动子区甲基化状态。结果：胃癌组织中RASSF1A基因启动子区CpG岛甲基化率为65．0％（39／60）,艋著高于癌旁组织6．7％（4／60）,及对照组0％（0／30）（P〈0．01）。胃癌组织中不同年龄、性别、分化程度及淋巴结转移与否的RASSF1A基因甲基化率的差异均无统计学意义。结论：胃癌中RASSF1A基因启动子区的高甲基化提示其与胃癌的发生密切相关,MSP法对RASSF1A基因启动子区甲基化的检测有望成为胃癌早期监测的重要方法。