Inhibition of DNA Methylation and Reactivation of Silenced Genes by Zebularine

Cheng, Jonathan; Matsen, Cindy B.; Gonzales, Felicidad A.; Ye, Wei; Greer, Sheldon; Márquez, Víctor E.; Jones, Peter A.; Selker, Eric U.

doi:10.1093/jnci/95.5.399

Cited by 482 publications

(415 citation statements)

References 35 publications

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“…Genomic DNA (4 μg) was treated with sodium bisulfite as previously described (14). The PCR primers used were 5′-TTAGTGGGGTATTTT-TATTTGTATGG-3′ (forward) and 5′-AAATATCCTAAAAATA-CAAACTACAC-3′ (reverse).…”

Section: Quantitation Of Dna Methylation By Ms-snupementioning

confidence: 99%

“…Zebularine, a novel inhibitor of DNA methylation, has been shown to have anticancer properties in vivo and in vitro (12)(13)(14)(15). Unlike 5-azacytidine and 5-aza-2′-deoxycytidine, which are chemically labile, zebularine is stable, making it possible to deliver the drug orally (14).…”

mentioning

confidence: 99%

“…Unlike 5-azacytidine and 5-aza-2′-deoxycytidine, which are chemically labile, zebularine is stable, making it possible to deliver the drug orally (14). However, chronic use of demethylating agents is of concern because genomewide hypomethylation has been associated with chromosomal instability and cancer in mice (16,17).…”

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confidence: 99%

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Long-term Epigenetic Therapy with Oral Zebularine Has Minimal Side Effects and Prevents Intestinal Tumors in Mice

Yoo

Chuang

Byun

et al. 2008

Cancer Prevention Research

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Recent successes in the application of epigenetic drugs for the treatment of myelodysplastic syndrome have raised questions on the safety of long-term administration of DNA methylation inhibitors. We treated preweaned cancer prone Apc Min/+ (Min) mice continuously with the DNA methylation inhibitor zebularine in their drinking water to determine the effects of the drug on normal mouse development as well as cancer prevention. Zebularine caused a tissue-specific reduction in DNA methylation at B1 short interspersed nucleotide elements in the small and large intestines of female Min mice but not in other organs examined after chronic oral treatment. No significant difference in the average weights of mice was observed during the treatment. In addition, analysis of global gene expression of colonic epithelial cells from the females indicated that only 3% to 6% of the genes were affected in their expression. We did not detect toxicity and abnormalities from the histopathologic analysis of liver and intestinal tissues. Lastly, we tested whether prevention of tumorigenesis can be achieved with chronic oral administration of zebularine in Min mice. The average number of polyps in Min females decreased from 58 to 1, whereas the average polyp number remained unaffected in Min males possibly due to differential activity of aldehyde oxidase. Taken together, our results show for the first time that long-term oral administration of zebularine causes a gender-specific abrogation of intestinal tumors while causing a tissuespecific DNA demethylation. Importantly, prolonged treatment of mice with epigenetic drugs resulted in only minor developmental and histologic changes.

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Section: Quantitation Of Dna Methylation By Ms-snupementioning

confidence: 99%

mentioning

confidence: 99%

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Long-term Epigenetic Therapy with Oral Zebularine Has Minimal Side Effects and Prevents Intestinal Tumors in Mice

Yoo

Chuang

Byun

et al. 2008

Cancer Prevention Research

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“…Although the mechanism of zebularine is similar to that of 5-aza-dC (13), a characteristic of zebularine is that it can be p.o. administered because of its stability in an aqueous solution (14). In addition, the sequential treatment with 5-aza-dC followed by zebularine has been shown to prevent remethylation, a common obstacle in antimethylation treatments (15).…”

Section: Introductionmentioning

confidence: 99%

Halogenated Thymidine Analogues Restore the Expression of Silenced Genes without Demethylation

et al. 2005

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Transcriptional silencing of tumor suppressor genes by aberrant DNA methylation is a characteristic frequently observed in cancer cells. Therefore, reversing this process is a therapeutic target against cancer. In this study, we established a screening system for silencing inhibitors with cell lines transfected by a retroviral vector containing a luciferase gene.More than 100 nucleosides were tested for antisilencing activity with a selected clone in which the silenced expression of luciferase could be recovered by 5-aza-2V -deoxycytidine. A group of halogenated thymidine analogues was found to reactivate transcription of not only the reporter retrovirus vector but also endogenous glutathione-S-transferase 1 gene, without influence to DNA hypermethylation. Gel mobility shift assay showed that 5-bromo-2V-deoxyuridine (BrdUrd) or 5-iodo-2V-deoxyuridine incorporation did not affect the binding of the methyl-CpG binding protein motif to methylated DNA. Finally, in the retroviral promoter, BrdUrd treatment increased the acetylated histone H3 level and decreased methylation of histone H3 Lys 9 in accordance with recovered transcription. This study shows that halogenated thymidines have an antisilencing effect without changing DNA methylation status by interfering with step(s) between DNA methylation and histone acetylation. (Cancer Res 2005; 65(15): 6927-33)

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“…(23) Our results, which demonstrate that phenotypic transition from VSMCs into osteoblast-like cells, can be prevented by demethylation are in harmony with other studies showing that demethylating agents (eg, 5-azacytidine and zebularine) help cells to preserve the myogenic phenotype. (27)(28)(29) The induction of methylation of the SM22a promoter with a methylating agent (S-adenosyl methionine) resulted in overexpression of Cbfa1 and mineral deposition despite no phosphate supplementation. S-Adenosyl methionine has been used previously to induce methylation of myogenic genes in myoblasts.…”

Section: Discussionmentioning

confidence: 99%

High-phosphate-induced calcification is related to SM22α promoter methylation in vascular smooth muscle cells

Madueño

Martínez-Moreno

et al. 2010

Journal of Bone and Mineral Research

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Hyperphosphatemia is closely related to vascular calcification in patients with chronic kidney disease. Vascular smooth muscle cells (VSMCs) exposed to high phosphate concentrations in vitro undergo phenotypic transition to osteoblast-like cells. Mechanisms underlying this transdifferentiation are not clear. In this study we used two in vitro models, human aortic smooth muscle cells and rat aortic rings, to investigate the phenotypic transition of VSMCs induced by high phosphate. We found that high phosphate concentration (3.3 mmol/L) in the medium was associated with increased DNA methyltransferase activity and methylation of the promoter region of SM22a. This was accompanied by loss of the smooth muscle cell-specific protein SM22a, gain of the osteoblast transcription factor Cbfa1, and increased alkaline phosphatase activity with the subsequent in vitro calcification. The addition of a demethylating agent (procaine) to the high-phosphate medium reduced DNA methyltransferase activity and prevented methylation of the SM22a promoter, which was accompanied by an increase in SM22a expression and less calcification. Additionally, downregulation of SM22a, either by siRNA or by a methyl group donor (S-adenosyl methionine), resulted in overexpression of Cbfa1.In conclusion, we demonstrate that methylation of SM22a promoter is an important event in vascular smooth muscle cell calcification and that high phosphate induces this epigenetic modification. These findings uncover a new insight into mechanisms by which high phosphate concentration promotes vascular calcification. ß

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Inhibition of DNA Methylation and Reactivation of Silenced Genes by Zebularine

Abstract: Zebularine is a stable DNA demethylating agent and the first drug in its class able to reactivate an epigenetically silenced gene by oral administration.

Cited by 482 publications

References 35 publications

Long-term Epigenetic Therapy with Oral Zebularine Has Minimal Side Effects and Prevents Intestinal Tumors in Mice

Long-term Epigenetic Therapy with Oral Zebularine Has Minimal Side Effects and Prevents Intestinal Tumors in Mice

Halogenated Thymidine Analogues Restore the Expression of Silenced Genes without Demethylation

High-phosphate-induced calcification is related to SM22α promoter methylation in vascular smooth muscle cells

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