Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.1.1.37 (DNA methyltransferase)
 4,983 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Recent studies show that stable expression of the human telomerase catalytic subunit, hTERT, alone can lead several types of normal human somatic cells to bypass replicative senescence and become immortal. The molecular mechanisms by which telomerase immortalizes cells are not fully understood, although a key function of telomerase is to maintain a critical length of telomeres in order to preserve the stability and integrity of the genome. Here we report that stable transfection of hTERT alone was sufficient to allow bovine capillary endothelial (BCE) cells to bypass senescence and acquire immortality. Surprisingly, telomere lengths in these stable transfectants were progressively shortened during an increasing number of population doublings (PDLs), despite high telomerase activity. The expression of the cyclin-dependent kinase inhibitors (CDKIs) p16INK4A and p21CIP1/WAF1 was concomitantly repressed, and the retinoblastoma protein (pRb) was maintained in a hyperphosphorylated state in the telomerase-expressing cells. Re-expression of p16INK4A in these cells by either treatment with a demethylating agent or by adenovirus-mediated expression reinduced a senescence-like phenotype, suggesting that the inactivation of p16INK4A was due to DNA methylation and was crucial for the immortalization process. In agreement with this finding, the expression levels of the prototypic DNA methyltransferase DNMT1 were elevated in the hTERT-positive cells.
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PMID:Immortalization of bovine capillary endothelial cells by hTERT alone involves inactivation of endogenous p16INK4A/pRb. 1258 45

DNA methylation at the 5th position of cytosine has been found to be correlated with tumorigenesis. An inhibitor of DNA methylase could, therefore, be used as an anticancer drug. However, only a few inhibitory compounds have been discovered due to the limitations for assaying the DNA methylation. In this study, we describe a modification of DNA cytosine-C5-methyltransferase assay system utilizing [(3)H]-labeled S-adenosyl-methionine (SAM) and Sephadex G-25 column. Pre-treatment of either lambda DNA or the promoter region of human telomerase (hTERT) with HaeIII methylase greatly reduced the digestion of the DNAs with the corresponding restriction enzyme HaeIII endonuclease (over 100-fold), and the result was further confirmed by agarose gel electrophoresis. Application of this column method to another modification/restriction system, EcoRI methylase/endonuclease, gave rise to the similar results. Our data suggest that the newly developed column method could be effective for rapid screening of large number of cytosine methylase inhibitors and could also be applicable to other DNA methylases.
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PMID:A column method for determination of DNA cytosine-C5-methyltransferase activity. 1476 31

5-Aza-2'-deoxycytidine (5azadC) inhibits DNA methyltransferase and subsequently induces the expression of genes silenced by methylation. While treatment with 5azadC downregulated hTERT and upregulated MGMT expression in two glioma cell lines, there was no change in the expression of these two genes in the normal cell line. However, cell viability was reduced as a result of 5azadC treatment in all three cell lines. 5azadC treatment reduced telomerase expression and activity and subsequently enhanced chemosensitivity towards cisplatin, taxol and tamoxifen but not with the alkylating agents temozolomide (TMZ), carmustine and chlorambucil. To further evaluate the effect of these findings, the level of hTERT and MGMT expression was measured in a recurrent anaplastic ependymoma, seven glioblastoma and two normal brain tissues. While four of eight gliomas and one of the normal tissues expressed MGMT, hTERT was expressed in all gliomas but not in the normal brain tissue. Results of this study suggest that taxol together with 5azadC may be a good therapeutic combination for glioma. In addition, the work on cell lines can be repeated on tissues utilizing hTERT as the therapeutic target for demethylation using 5azadC in glioma.
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PMID:Epigenetic silencing of telomerase and a non-alkylating agent as a novel therapeutic approach for glioma. 1802 53

Previously, we demonstrated that demethylation with 5-Aza-2'-deoxycytidine (5azadC) resulted in reduced levels of telomerase that led to telomere shortening, enhanced MGMT expression and enhanced chemosensitivity. Although the results were encouraging, the fact that 5azadC is highly toxic and nonspecific, thus is not favored as a therapeutic molecule. The aim of this research is to downregulate the DNA methyltransferase (DNMT1) gene using three sets of double-stranded RNA oligos designed to align different regions of DNMT1 sequence. Results showed the small-interfering RNA (siRNA) 1 and 3 demonstrated significant levels of silencing DNMT1 and hTERT transcription after 24-hour treatment (p = 0.01) and approximately 90% and 70% transcriptional downregulation of DNMT1 and hTERT, respectively after 48 hours. However, siRNA 2 downregulated DNMT1, hTERT, and MGMT in GOS-3 and U87-MG cells that was attributed to sequence homology between oligo 2 and MGMT complementary DNA. The siRNA-treated glioma cell lines GOS-3 and U87-MG were subjected to two chemotherapeutic agents; taxol and Temozolomide (TMZ). Results suggest that either a combination of siRNA 1 or 3 followed by taxol (2-6 muM) after 48 hours or a combination of siRNA 1 or 3 followed by TMZ (600-1000 microM) after 24 hours would be novel and effective glioma therapies.
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PMID:Silencing DNA methyltransferase (DNMT) enhances glioma chemosensitivity. 1892 31

Epigallocatechin-3-gallate (EGCG), a major component of green tea polyphenols (GTP), has been reported to downregulate telomerase activity in breast cancer cells thereby increasing cellular apoptosis and inhibiting cellular proliferation. However, the major concerns with GTPs are their bioavailability and stability under physiologic conditions. In the present study, we show that treatments with EGCG and a novel prodrug of EGCG (pro-EGCG or pEGCG) dose- and time-dependently inhibited the proliferation of human breast cancer MCF-7 and MDA-MB-231 cells but not normal control MCF10A cells. Furthermore, both EGCG and pro-EGCG inhibited the transcription of hTERT (human telomerase reverse transcriptase), the catalytic subunit of telomerase, through epigenetic mechanisms in estrogen receptor (ER)-positive MCF-7 and ER-negative MDA-MB-231 cells. The downregulation of hTERT expression was found to be because of hTERT promoter hypomethylation and histone deacetylations, mediated at least partially through inhibition of DNA methyltransferase and histone acetyltransferase activities, respectively. In addition, we also observed that EGCG and pEGCG can remodel chromatin structures of the hTERT promoter by decreasing the level of acetyl-H3, acetyl-H3K9, and acetyl-H4 to the hTERT promoter. EGCG and pEGCG induced chromatin alterations that facilitated the binding of many hTERT repressors such as MAD1 and E2F-1 to the hTERT regulatory region. Depletion of E2F-1 and MAD1 by using siRNA reversed the pEGCG downregulated hTERT expression and associated cellular apoptosis differently in ER-positive and ER-negative breast cancer cells. Collectively, our data provide new insights into breast cancer prevention through epigenetic modulation of telomerase by using pro-EGCG, a more stable form of EGCG, as a novel chemopreventive compound.
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PMID:A novel prodrug of epigallocatechin-3-gallate: differential epigenetic hTERT repression in human breast cancer cells. 2141 98

Telomerase activation is a critical step in human carcinogenesis through the maintenance of telomeres. Telomerase activity is primarily regulated by the human telomerase reverse transcriptase gene (hTERT), thus, an improved understanding of the transcriptional control of hTERT may provide potential therapeutic targets for the treatment of leukemia and other forms of cancer. Epigenetic modulation, a significant regulatory process in cell biology, has recently been shown to be involved in the regulation of the hTERT gene. Moreover, several epigenetic modifiers, including DNA methyltransferase (DNMT) and histone deacetylase (HDAC) inhibitors, are now in pre- and early clinical trials of leukemia as monotherapies or in combination with other drugs, and have achieved significant clinical success. In the present review, the epigenetic mechanisms associated with telomerase activity in leukemia, and the therapeutic potential of an antitelomerase strategy that combines epigenetic modifiers with telomerase hTR subunit small molecule inhibitors are discussed.
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PMID:Epigenetic regulation of the human telomerase reverse transciptase gene: A potential therapeutic target for the treatment of leukemia (Review). 2413 23

The maintenance cytosine DNA methyltransferase DNMT1 and de novo methyltransferase DNMT3b cooperate to establish aberrant DNA methylation and chromatin complexes to repress gene transcription during cancer development. The expression of DNMT3b was constitutively increased 5-20-fold in hTERT/CDK4-immortalized human bronchial epithelial cells (HBECs) before treatment with low doses of tobacco carcinogens. Overexpression of DNMT3b increased and accelerated carcinogen-induced transformation. Genome-wide profiling of transformed HBECs identified 143 DNMT3b-target genes, many of which were transcriptionally regulated by the polycomb repressive complex 2 (PRC2) complex and silenced through aberrant methylation in non-small-cell lung cancer cell lines. Two genes studied in detail, MAL and OLIG2, were silenced during transformation, initially through enrichment for H3K27me3 and H3K9me2, commonly methylated in lung cancer, and exert tumor suppressor effects in vivo through modulating cancer-related pathways. Re-expression of MAL and OLIG2 to physiological levels dramatically reduced the growth of lung tumor xenografts. Our results identify a key role for DNMT3b in the earliest stages of initiation and provide a comprehensive catalog of genes targeted for silencing by this methyltransferase in non-small-cell lung cancer.
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PMID:Global identification of genes targeted by DNMT3b for epigenetic silencing in lung cancer. 2446 50

Dietary compounds that possess the properties of altering epigenetic processes are gaining popularity as targets for cancer prevention studies. These compounds when administered at optimal concentrations and especially in combination can have enhanced effects in cancer prevention or therapy. It is important to study the interaction of two or more compounds in order to assess their role in enhancing prevention. Genistein (GEN), found in soy, has been extensively studied for its role as an epigenetic modifier especially as a DNA methyltransferase (DNMT) inhibitor and sulforaphane (SFN), found in cruciferous vegetables, is known as a histone deacetylase (HDAC) inhibitor. However, very little is known about the effects of these two compounds in conjunction in breast cancer prevention or therapy. In our current study, we determined that, at certain doses, the compounds have synergistic effects in decreasing cellular viability of breast cancer cell lines. Our results indicate that the combination of GEN and SFN is much more effective than their single doses in increasing the rate of apoptosis and lowering the colony forming potential of these cells. We determined that these compounds inhibit cell cycle progression to G2 phase in MDA-MB-231 and G1 phase in MCF-7 breast cancer cell lines. Additionally, we determined that the combination is effective as an HDAC and histone methyltransferase (HMT) inhibitor. Furthermore, we demonstrated that this combination downregulates the levels of HDAC2 and HDAC3 both at the mRNA and protein levels. We also found that these compounds have the potential to downregulate KLF4 levels, which plays an important role in stem cell formation. The combination of GEN and SFN is also effective in downregulating hTERT levels, which is known to be activated when KLF4 binds to its promoter region. Our hypothesis is further strengthened by in vivo studies, where the combination is administered to transgenic mice in the form of genistein and SFN-enriched broccoli sprouts. We have demonstrated that the combination is more effective in preventing or treating mammary cancer via extending tumor latency and reducing tumor volumes/sizes than either of these dietary components administered alone. These results are consistent with our in vitro study suggesting potential preventive and therapeutic effects of this novel dietary combinatorial approach against breast cancer.
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PMID:The Effects of Combinatorial Genistein and Sulforaphane in Breast Tumor Inhibition: Role in Epigenetic Regulation. 2989 71