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LINE-1 hypomethylation in cancer is highly variable and inversely correlated with microsatellite instability.

Estécio MR, Gharibyan V, Shen L, Ibrahim AE, Doshi K, He R, Jelinek J, Yang AS, Yan PS, Huang TH, Tajara EH, Issa JP - PLoS ONE (2007)

Bottom Line: Interestingly, some tumor samples in this group showed increase in LINE-1 methylation.Microarray analysis of repetitive element methylation confirmed this observation and showed a high degree of variability in hypomethylation between samples.We extended LINE-1 analysis to cancer cell lines from different tissues and found that 50/61 were hypomethylated compared to peripheral blood lymphocytes and normal colon mucosa.

View Article: PubMed Central - PubMed

Affiliation: Department of Leukemia, University of Texas M. D. Anderson Cancer Center, Houston, Texas, United States of America.

ABSTRACT

Background: Alterations in DNA methylation in cancer include global hypomethylation and gene-specific hypermethylation. It is not clear whether these two epigenetic errors are mechanistically linked or occur independently. This study was performed to determine the relationship between DNA hypomethylation, hypermethylation and microsatellite instability in cancer.

Methodology/principal findings: We examined 61 cancer cell lines and 60 colorectal carcinomas and their adjacent tissues using LINE-1 bisulfite-PCR as a surrogate for global demethylation. Colorectal carcinomas with sporadic microsatellite instability (MSI), most of which are due to a CpG island methylation phenotype (CIMP) and associated MLH1 promoter methylation, showed in average no difference in LINE-1 methylation between normal adjacent and cancer tissues. Interestingly, some tumor samples in this group showed increase in LINE-1 methylation. In contrast, MSI-showed a significant decrease in LINE-1 methylation between normal adjacent and cancer tissues (P<0.001). Microarray analysis of repetitive element methylation confirmed this observation and showed a high degree of variability in hypomethylation between samples. Additionally, unsupervised hierarchical clustering identified a group of highly hypomethylated tumors, composed mostly of tumors without microsatellite instability. We extended LINE-1 analysis to cancer cell lines from different tissues and found that 50/61 were hypomethylated compared to peripheral blood lymphocytes and normal colon mucosa. Interestingly, these cancer cell lines also exhibited a large variation in demethylation, which was tissue-specific and thus unlikely to be resultant from a stochastic process.

Conclusion/significance: Global hypomethylation is partially reversed in cancers with microsatellite instability and also shows high variability in cancer, which may reflect alternative progression pathways in cancer.

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Related in: MedlinePlus

LINE-1 methylation variability in cancer cell lines. DNA samples of normal peripheral blood lymphocyte, normal colon mucosa and sixty-one cell lines from eight different tissues types were investigated for LINE-1 methylation using bisulfite PCR followed by pyrosequencing. The normal tissues presented high levels of LINE-1 methylation (above 70% in average), and a large variation in methylation levels was observed for cancer cell lines, with a minimum methylation density of 6.5% being observed for the leukemia cell line K562. Taken as a group, leukemia cell lines were moderately demethylated (average 56.1%), followed by ovary, colon, prostate and lung cancer cell lines (variation from 49.7% to 35.1%). Central nervous system (CNS), breast and the one liver cancer cell lines tested were deeply demethylated (bellow 30% in average). Dotted line represents average methylation in normal controls.
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pone-0000399-g004: LINE-1 methylation variability in cancer cell lines. DNA samples of normal peripheral blood lymphocyte, normal colon mucosa and sixty-one cell lines from eight different tissues types were investigated for LINE-1 methylation using bisulfite PCR followed by pyrosequencing. The normal tissues presented high levels of LINE-1 methylation (above 70% in average), and a large variation in methylation levels was observed for cancer cell lines, with a minimum methylation density of 6.5% being observed for the leukemia cell line K562. Taken as a group, leukemia cell lines were moderately demethylated (average 56.1%), followed by ovary, colon, prostate and lung cancer cell lines (variation from 49.7% to 35.1%). Central nervous system (CNS), breast and the one liver cancer cell lines tested were deeply demethylated (bellow 30% in average). Dotted line represents average methylation in normal controls.

Mentions: To verify if the variability in genome-wide methylation is restricted to primary colorectal carcinomas or also occurs in other tumor types, we applied the LINE-1 bisulfite-pyrosequencing method to sixty-one cancer cell lines from eight different tissues types (breast, central nervous system, colon, leukemia, lung, ovary, prostate and liver). Interestingly, we found a marked decrease in LINE-1 methylation in most of the studied cell lines (Figure 4). Overall, 50/61 tested cancer cell lines were hypomethylated for LINE-1, with a relative demethylation of 15% or more (absolute methylation density lower than 60%) compared to peripheral blood lymphocytes and normal colon mucosa. Similarly to primary colorectal tumors, these cancer cell lines exhibited high variability in LINE-1 methylation, ranging from 6.5% (K562, a CML cell line with erythroleukemia features) to 74.2% (CEM, an acute lymphoblastic leukemia cell line). There is an apparent tissue-specificity for demethylation; the lowest levels of LINE-1 methylation were observed in liver (24.0%), followed by CNS (28.9%), breast (29.8%), lung (35.1%), prostate (41.9%), ovary (49.7%), colon (46.7%) and leukemia (56.1%). While an interesting finding, we caution generalization of the data because: (i) LINE-1 methylation was not studied for normal tissues except colon and peripheral blood; and (ii) a small number of cell lines were analyzed for liver and prostate cancer.


LINE-1 hypomethylation in cancer is highly variable and inversely correlated with microsatellite instability.

Estécio MR, Gharibyan V, Shen L, Ibrahim AE, Doshi K, He R, Jelinek J, Yang AS, Yan PS, Huang TH, Tajara EH, Issa JP - PLoS ONE (2007)

LINE-1 methylation variability in cancer cell lines. DNA samples of normal peripheral blood lymphocyte, normal colon mucosa and sixty-one cell lines from eight different tissues types were investigated for LINE-1 methylation using bisulfite PCR followed by pyrosequencing. The normal tissues presented high levels of LINE-1 methylation (above 70% in average), and a large variation in methylation levels was observed for cancer cell lines, with a minimum methylation density of 6.5% being observed for the leukemia cell line K562. Taken as a group, leukemia cell lines were moderately demethylated (average 56.1%), followed by ovary, colon, prostate and lung cancer cell lines (variation from 49.7% to 35.1%). Central nervous system (CNS), breast and the one liver cancer cell lines tested were deeply demethylated (bellow 30% in average). Dotted line represents average methylation in normal controls.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC1851990&req=5

pone-0000399-g004: LINE-1 methylation variability in cancer cell lines. DNA samples of normal peripheral blood lymphocyte, normal colon mucosa and sixty-one cell lines from eight different tissues types were investigated for LINE-1 methylation using bisulfite PCR followed by pyrosequencing. The normal tissues presented high levels of LINE-1 methylation (above 70% in average), and a large variation in methylation levels was observed for cancer cell lines, with a minimum methylation density of 6.5% being observed for the leukemia cell line K562. Taken as a group, leukemia cell lines were moderately demethylated (average 56.1%), followed by ovary, colon, prostate and lung cancer cell lines (variation from 49.7% to 35.1%). Central nervous system (CNS), breast and the one liver cancer cell lines tested were deeply demethylated (bellow 30% in average). Dotted line represents average methylation in normal controls.
Mentions: To verify if the variability in genome-wide methylation is restricted to primary colorectal carcinomas or also occurs in other tumor types, we applied the LINE-1 bisulfite-pyrosequencing method to sixty-one cancer cell lines from eight different tissues types (breast, central nervous system, colon, leukemia, lung, ovary, prostate and liver). Interestingly, we found a marked decrease in LINE-1 methylation in most of the studied cell lines (Figure 4). Overall, 50/61 tested cancer cell lines were hypomethylated for LINE-1, with a relative demethylation of 15% or more (absolute methylation density lower than 60%) compared to peripheral blood lymphocytes and normal colon mucosa. Similarly to primary colorectal tumors, these cancer cell lines exhibited high variability in LINE-1 methylation, ranging from 6.5% (K562, a CML cell line with erythroleukemia features) to 74.2% (CEM, an acute lymphoblastic leukemia cell line). There is an apparent tissue-specificity for demethylation; the lowest levels of LINE-1 methylation were observed in liver (24.0%), followed by CNS (28.9%), breast (29.8%), lung (35.1%), prostate (41.9%), ovary (49.7%), colon (46.7%) and leukemia (56.1%). While an interesting finding, we caution generalization of the data because: (i) LINE-1 methylation was not studied for normal tissues except colon and peripheral blood; and (ii) a small number of cell lines were analyzed for liver and prostate cancer.

Bottom Line: Interestingly, some tumor samples in this group showed increase in LINE-1 methylation.Microarray analysis of repetitive element methylation confirmed this observation and showed a high degree of variability in hypomethylation between samples.We extended LINE-1 analysis to cancer cell lines from different tissues and found that 50/61 were hypomethylated compared to peripheral blood lymphocytes and normal colon mucosa.

View Article: PubMed Central - PubMed

Affiliation: Department of Leukemia, University of Texas M. D. Anderson Cancer Center, Houston, Texas, United States of America.

ABSTRACT

Background: Alterations in DNA methylation in cancer include global hypomethylation and gene-specific hypermethylation. It is not clear whether these two epigenetic errors are mechanistically linked or occur independently. This study was performed to determine the relationship between DNA hypomethylation, hypermethylation and microsatellite instability in cancer.

Methodology/principal findings: We examined 61 cancer cell lines and 60 colorectal carcinomas and their adjacent tissues using LINE-1 bisulfite-PCR as a surrogate for global demethylation. Colorectal carcinomas with sporadic microsatellite instability (MSI), most of which are due to a CpG island methylation phenotype (CIMP) and associated MLH1 promoter methylation, showed in average no difference in LINE-1 methylation between normal adjacent and cancer tissues. Interestingly, some tumor samples in this group showed increase in LINE-1 methylation. In contrast, MSI-showed a significant decrease in LINE-1 methylation between normal adjacent and cancer tissues (P<0.001). Microarray analysis of repetitive element methylation confirmed this observation and showed a high degree of variability in hypomethylation between samples. Additionally, unsupervised hierarchical clustering identified a group of highly hypomethylated tumors, composed mostly of tumors without microsatellite instability. We extended LINE-1 analysis to cancer cell lines from different tissues and found that 50/61 were hypomethylated compared to peripheral blood lymphocytes and normal colon mucosa. Interestingly, these cancer cell lines also exhibited a large variation in demethylation, which was tissue-specific and thus unlikely to be resultant from a stochastic process.

Conclusion/significance: Global hypomethylation is partially reversed in cancers with microsatellite instability and also shows high variability in cancer, which may reflect alternative progression pathways in cancer.

Show MeSH
Related in: MedlinePlus