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Continuous zebularine treatment effectively sustains demethylation in human bladder cancer cells 总被引:18,自引:0,他引:18
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Cheng JC Weisenberger DJ Gonzales FA Liang G Xu GL Hu YG Marquez VE Jones PA 《Molecular and cellular biology》2004,24(3):1270-1278
During tumorigenesis, tumor suppressor and cancer-related genes are commonly silenced by aberrant DNA methylation in their promoter regions. Recently, we reported that zebularine [1-(beta-D-ribofuranosyl)-1,2-dihydropyrimidin-2-one] acts as an inhibitor of DNA methylation and exhibits chemical stability and minimal cytotoxicity both in vitro and in vivo. Here we show that continuous application of zebularine to T24 cells induces and maintains p16 gene expression and sustains demethylation of the 5' region for over 40 days, preventing remethylation. In addition, continuous zebularine treatment effectively and globally demethylated various hypermethylated regions, especially CpG-poor regions. The drug caused a complete depletion of extractable DNA methyltransferase 1 (DNMT1) and partial depletion of DNMT3a and DNMT3b3. Last, sequential treatment with 5-aza-2'-deoxycytidine followed by zebularine hindered the remethylation of the p16 5' region and gene resilencing, suggesting the possible combination use of both drugs as a potential anticancer regimen. 相似文献
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Kavita M. Pattani Ethan Soudry Chad A. Glazer Michael F. Ochs Hao Wang Juliana Schussel Wenyue Sun Patrick Hennessey Wojciech Mydlarz Myriam Loyo Semra Demokan Ian M. Smith Joseph A. Califano 《PloS one》2012,7(9)
Purpose
Although promoter hypermethylation has been an accepted means of tumor suppressor gene inactivation, activation of otherwise normally repressed proto-oncogenes by promoter demethylation has been infrequently documented.Experimental Design
In this study we performed an integrative, whole-genome analysis for discovery of epigenetically activated proto-oncogenes in head and neck cancer tumors. We used the 47K GeneChip U133 Plus 2.0 Affymetrix expression microarray platform to obtain re-expression data from 5-aza treated normal cell line and expression data from primary head and neck squamous cell carcinoma (HNSCC) tumor tissues and normal mucosa tissues. We then investigated candidate genes by screening promoter regions for CpG islands and bisulfite sequencing followed by QUMSP and RT PCR for the best candidate genes. Finally, functional studies were performed on the top candidate gene.Results
From the top 178 screened candidates 96 had CpG islands in their promoter region. Seven candidate genes showed promoter region methylation in normal mucosa samples and promoter demethylation in a small cohort of primary HNSCC tissues. We then studied the demethylation of the top 3 candidate genes in an expanded cohort of 76 HNSCC tissue samples and 17 normal mucosa samples. We identified MAGEB2 as having significant promoter demethylation in primary head and neck squamous cell carcinoma tissues. We then found significantly higher expression of MAGEB2 in tumors in a separate cohort of 73 primary HNSCC tissues and 31 normal tissues. Finally, we found that MAGEB2 has growth promoting effects on minimally transformed oral keratinocyte cell lines but not a definite effect on HNSCC cell lines.Conclusion
In conclusion, we identified MAGEB2 as activated by promoter demethylation in HNSCCand demonstrates growth promoting effects in a minimally transformed oral keratinocyte cell line. More studies are needed to evaluate MAGBE2''s exact role in HNSCC. 相似文献4.
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Nicola Amodio Dina Bellizzi Marzia Leotta Lavinia Raimondi Lavinia Biamonte Patrizia D’Aquila Maria Teresa Di Martino Teresa Calimeri Marco Rossi Marta Lionetti Emanuela Leone Giuseppe Passarino Antonino Neri Antonio Giordano Pierosandro Tagliaferri Pierfrancesco Tassone 《Cell cycle (Georgetown, Tex.)》2013,12(23):3650-3662
Epigenetic silencing of tumor suppressor genes frequently occurs and may account for their inactivation in cancer cells. We previously demonstrated that miR-29b is a tumor suppressor microRNA (miRNA) that targets de novo DNA methyltransferases and reduces the global DNA methylation of multiple myeloma (MM) cells. Here, we provide evidence that epigenetic activity of miR-29b leads to promoter demethylation of suppressor of cytokine signaling-1 (SOCS-1), a hypermethylated tumor suppressor gene. Enforced expression of synthetic miR-29b mimics in MM cell lines resulted in SOCS-1 gene promoter demethylation, as assessed by Sequenom MassARRAY EpiTYPER analysis, and SOCS-1 protein upregulation. miR-29b-induced SOCS-1 demethylation was associated with reduced STAT3 phosphorylation and impaired NFκB activity. Downregulation of VEGF-A and IL-8 mRNAs could be detected in MM cells transfected with miR-29b mimics as well as in endothelial (HUVEC) or stromal (HS-5) cells treated with conditioned medium from miR-29b-transfected MM cells. Notably, enforced expression of miR-29b mimics increased adhesion of MM cells to HS-5 and reduced migration of both MM and HUVEC cells. These findings suggest that miR-29b is a negative regulator of either MM or endothelial cell migration. Finally, the proteasome inhibitor bortezomib, which induces the expression of miR-29b, decreased global DNA methylation by a miR-29b-dependent mechanism and induced SOCS-1 promoter demethylation and protein upregulation. In conclusion, our data indicate that miR-29b is endowed with epigenetic activity and mediates previously unknown functions of bortezomib in MM cells. 相似文献
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Mayada Achour Marc Mousli Mahmoud Alhosin Abdulkhaleg Ibrahim Jean Peluso Christian D. Muller Valérie B. Schini-Kerth Ali Hamiche Sirano Dhe-Paganon Christian Bronner 《Biochemical and biophysical research communications》2013,430(1):208-212
Ubiquitin-like containing PHD and Ring finger 1 (UHRF1) contributes to silencing of tumor suppressor genes by recruiting DNA methyltransferase 1 (DNMT1) to their hemi-methylated promoters. Conversely, demethylation of these promoters has been ascribed to the natural anti-cancer drug, epigallocatechin-3-gallate (EGCG). The aim of the present study was to investigate whether the UHRF1/DNMT1 pair is an important target of EGCG action. Here, we show that EGCG down-regulates UHRF1 and DNMT1 expression in Jurkat cells, with subsequent up-regulation of p73 and p16INK4A genes. The down-regulation of UHRF1 is dependent upon the generation of reactive oxygen species by EGCG. Up-regulation of p16INK4A is strongly correlated with decreased promoter binding by UHRF1. UHRF1 over-expression counteracted EGCG-induced G1-arrested cells, apoptosis, and up-regulation of p16INK4A and p73. Mutants of the Set and Ring Associated (SRA) domain of UHRF1 were unable to down-regulate p16INK4A and p73, either in the presence or absence of EGCG. Our results show that down-regulation of UHRF1 is upstream to many cellular events, including G1 cell arrest, up-regulation of tumor suppressor genes and apoptosis. 相似文献
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天花粉蛋白对宫颈癌HeLa细胞TSLC1基因甲基化及其表达的影响 总被引:3,自引:0,他引:3
目的:检测人宫颈癌HeLa细胞中TSLC1基因甲基化的状况,研究在人宫颈癌HeLa细胞凋亡过程中TSLC1基因甲基化的变化情况,探讨肿瘤细胞凋亡与抑癌基因甲基化的相关性,并进一步证实天花粉蛋白(TCS)去甲基化作用是否存在普遍性,以促进天花粉蛋白的临床应用。方法:应用甲基化特异性PCR(MSP)法检测人宫颈癌HeLa细胞及其凋亡过程中TSLC1基因甲基化的状况;采用实时定量RT-PCR技术检测TCS处理前、后HeLa细胞TSLC1基因表达的变化。结果:肿瘤抑制基因TSLC1在人宫颈癌HeLa细胞中呈高度甲基化状态,经40μg/mL TCS处理48h后,TSLC1基因甲基化程度明显降低;RT-PCR检测结果显示,TCS处理组HeLa细胞中TSLC1 mRNA的表达量高于未处理组,提示TSLC1基因启动子区CpG岛甲基化是导致其低表达的重要机制。结论:肿瘤抑制基因TSLC1启动子甲基化在人宫颈癌癌变过程中可能是一种重要的分子调控机制;人宫颈癌HeLa细胞凋亡与抑癌基因的去甲基化之间可能存在某些密切的相关性;TCS对肿瘤抑制基因TSLC1有一定的去甲基化作用。 相似文献
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Francesca Crucianelli Rossella Tricarico Daniela Turchetti Greta Gorelli Francesca Gensini Roberta Sestini Laura Giunti Monica Pedroni Maurizio Ponz de Leon Serenella Civitelli Maurizio Genuardi 《Epigenetics》2014,9(10):1431-1438
Lynch syndrome (LS) is a tumor predisposing condition caused by constitutional defects in genes coding for components of the mismatch repair (MMR) apparatus. While hypermethylation of the promoter of the MMR gene MLH1 occurs in about 15% of colorectal cancer samples, it has also been observed as a constitutional alteration, in the absence of DNA sequence mutations, in a small number of LS patients. In order to obtain further insights on the phenotypic characteristics of MLH1 epimutation carriers, we investigated the somatic and constitutional MLH1 methylation status of 14 unrelated subjects with a suspicion of LS who were negative for MMR gene constitutional mutations and whose tumors did not express the MLH1 protein. A novel case of constitutional MLH1 epimutation was identified. This patient was affected with multiple primary tumors, including breast cancer, diagnosed starting from the age of 55 y. Investigation of her offspring by allele specific expression revealed that the epimutation was not stable across generations. We also found MLH1 hypermethylation in cancer samples from 4 additional patients who did not have evidence of constitutional defects. These patients had some characteristics of LS, namely early age at onset and/or positive family history, raising the possibility of genetic influences in the establishment of somatic MLH1 methylation. 相似文献
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Inga Hinrichsen Matthias Kemp Jan Peveling-Oberhag Sandra Passmann Guido Plotz Stefan Zeuzem Angela Brieger 《PloS one》2014,9(1)
Epigenetic silencing of tumour suppressor genes has been observed in various cancers. Looking at hepatocellular carcinoma (HCC) specific protein silencing was previously demonstrated to be associated with the Hepatitis C virus (HCV). However, the proposed HCV dependent promoter methylation of DNA mismatch repair (MMR) genes and thereby enhanced progression of hepatocarcinogenesis has been the subject of controversial discussion. We investigated promoter methylation pattern of the MMR genes MLH1, MSH2 and PMS2 as well as the cyclin-dependent kinase inhibitor 2A gene (p16) in 61 well characterized patients with HCCs associated with HCV, Hepatitis B virus infection or alcoholic liver disease. DNA was isolated from formalin-fixed, paraffin-embedded tumour and non-tumour adjacent tissue and analysed by methylation-specific PCR. Moreover, microsatellite analysis was performed in tissues showing methylation in MMR gene promoters. Our data demonstrated that promoter methylation of MLH1, MSH2, PMS2 and p16 is present among all considered HCCs. Hereby, promoter silencing was detectable more frequently in advanced-stage HCCs than in low-stage ones. However, there was no significant correlation between aberrant DNA methylation of MMR genes or p16 and HCV infection in related HCC specimens. In summary, we show that promoter methylation of essential MMR genes and p16 is detectable in HCCs most dominantly in pT3 stage tumour cases. Since loss of MMR proteins was previously described to be not only responsible for tumour development but also for chemotherapy resistance, the knowledge of mechanisms jointly responsible for HCC progression might enable significant improvement of individual HCC therapy in the future. 相似文献
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Xiang T Li L Yin X Yuan C Tan C Su X Xiong L Putti TC Oberst M Kelly K Ren G Tao Q 《PloS one》2012,7(1):e29783
Background
Breast cancer (BrCa) is a complex disease driven by aberrant gene alterations and environmental factors. Recent studies reveal that abnormal epigenetic gene regulation also plays an important role in its pathogenesis. Ubiquitin carboxyl- terminal esterase L1 (UCHL1) is a tumor suppressor silenced by promoter methylation in multiple cancers, but its role and alterations in breast tumorigenesis remain unclear.Methodology/Principal Findings
We found that UCHL1 was frequently downregulated or silenced in breast cancer cell lines and tumor tissues, but readily expressed in normal breast tissues and mammary epithelial cells. Promoter methylation of UCHL1 was detected in 9 of 10 breast cancer cell lines (90%) and 53 of 66 (80%) primary tumors, but rarely in normal breast tissues, which was statistically correlated with advanced clinical stage and progesterone receptor status. Pharmacologic demethylation reactivated UCHL1 expression along with concomitant promoter demethylation. Ectopic expression of UCHL1 significantly suppressed the colony formation and proliferation of breast tumor cells, through inducing G0/G1 cell cycle arrest and apoptosis. Subcellular localization study showed that UCHL1 increased cytoplasmic abundance of p53. We further found that UCHL1 induced p53 accumulation and reduced MDM2 protein level, and subsequently upregulated the expression of p21, as well as cleavage of caspase3 and PARP, but not in catalytic mutant UCHL1 C90S-expressed cells.Conclusions/Significance
UCHL1 exerts its tumor suppressive functions by inducing G0/G1cell cycle arrest and apoptosis in breast tumorigenesis, requiring its deubiquitinase activity. Its frequent silencing by promoter CpG methylation may serve as a potential tumor marker for breast cancer. 相似文献13.
Fabio Coppedè Francesca Migheli Angela Lopomo Alessandra Failli Annalisa Legitimo Rita Consolini Gabriella Fontanini Elisa Sensi Adele Servadio Massimo Seccia Giuseppe Zocco Massimo Chiarugi Roberto Spisni Lucia Migliore 《Epigenetics》2014,9(4):621-633
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. 相似文献
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Synergism between DNA methylation and macroH2A1 occupancy in epigenetic silencing of the tumor suppressor gene p16(CDKN2A) 总被引:1,自引:0,他引:1
Barzily-Rokni M Friedman N Ron-Bigger S Isaac S Michlin D Eden A 《Nucleic acids research》2011,39(4):1326-1335
Promoter hypermethylation and heterochromatinization is a frequent event leading to gene inactivation and tumorigenesis. At the molecular level, inactivation of tumor suppressor genes in cancer has many similarities to the inactive X chromosome in female cells and is defined and maintained by DNA methylation and characteristic histone modifications. In addition, the inactive-X is marked by the histone macroH2A, a variant of H2A with a large non-histone region of unknown function. Studying tumor suppressor genes (TSGs) silenced in cancer cell lines, we find that when active, these promoters are associated with H2A.Z but become enriched for macroH2A1 once silenced. Knockdown of macroH2A1 was not sufficient for reactivation of silenced genes. However, when combined with DNA demethylation, macroH2A1 deficiency significantly enhanced reactivation of the tumor suppressor genes p16, MLH1 and Timp3 and inhibited cell proliferation. Our findings link macroH2A1 to heterochromatin of epigenetically silenced cancer genes and indicate synergism between macroH2A1 and DNA methylation in maintenance of the silenced state. 相似文献
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Liang Jing Wang Xian Wang Jian Bin Zhang Francis K.L. Chan Joseph J.Y. Sung 《Biochemical and biophysical research communications》2009,379(4):959-231
As one of major epigenetic changes responsible for tumor suppressor gene inactivation in the development of cancer, promoter hypermethylation was proposed as a marker to define novel tumor suppressor genes. In the current study we identified ZIC1 (Zic family member 1, odd-paired Drosophila homolog) as a novel tumor suppressor gene silenced through promoter hypermethylation in gastric cancer, the second leading cause of cancer death worldwide. In all of gastric cancer cells lines examined, ZIC1 expression was downregulated and such downregulation was accompanied with the hypermethylation of ZIC1 promoter. Demethylation treatment with 5-aza-2′-deoxycytidine (Aza) reversed ZIC1 downregulation, highlighting the importance of promoter methylation to ZIC1 downregulation in gastric cancer cells. Notably, ZIC1 expression was significantly downregulated in primary gastric carcinoma tissues in comparison with non-tumor adjacent gastric tissues (p < 0.01). Accordingly, promoter methylation of ZIC1 was frequently detected in primary gastric carcinoma tissues (94.6%, 35/37) but not normal gastric tissues, indicating that promoter hypermethylation mediated ZIC1 downregulation may play an important role in gastric carcinogenesis. Indeed, ectopic expression of ZIC1 led to the growth inhibition of gastric cancer cells through the induction of S-phase cell cycle arrest (p < 0.01). Our results revealed ZIC1 as a novel candidate tumor suppressor gene downregulated through promoter hypermethylation in gastric cancer. 相似文献
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Rafael Guerrero-Preston Christina Michailidi Luigi Marchionni Curtis R Pickering Mitchell J Frederick Jeffrey N Myers Srinivasan Yegnasubramanian Tal Hadar Maartje G Noordhuis Veronika Zizkova Elana Fertig Nishant Agrawal William Westra Wayne Koch Joseph Califano Victor E Velculescu David Sidransky 《Epigenetics》2014,9(7):1031-1046
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Cui Y Ying Y van Hasselt A Ng KM Yu J Zhang Q Jin J Liu D Rhim JS Rha SY Loyo M Chan AT Srivastava G Tsao GS Sellar GC Sung JJ Sidransky D Tao Q 《PloS one》2008,3(8):e2990