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, 33 (12), 5325-33

Green Tea Polyphenol Epigallocatechin 3-gallate, Contributes to the Degradation of DNMT3A and HDAC3 in HCT 116 Human Colon Cancer Cells

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Green Tea Polyphenol Epigallocatechin 3-gallate, Contributes to the Degradation of DNMT3A and HDAC3 in HCT 116 Human Colon Cancer Cells

Vondina R Moseley et al. Anticancer Res.

Abstract

Background: Colon cancer is still the second leading cause of cancer deaths in the United States. Epigenetic gene silencing involving DNA methyltransferases (DNMTs) and histone deacetylases (HDACs) plays an important role in the progression of colon cancer.

Materials and methods: In the present study we found that the sensitivity of colon cancer cells to methylation plays a role in its response to alternative therapy involving the green tea polyphenol, epigallocatechin 3-gallate. HDAC and DNMT protein expression were reduced when methylation-sensitive HCT 116 human colon cancer cells was treated with EGCG, but was relatively stable in the HT-29 cell line. This decrease in expression may be partially explained by our finding that DNMT3A and HDAC3 are degraded in the methylation-sensitive colon cancer cells in part by inhibiting their association with the E3 ubiquitin ligase, UHRF1.

Conclusion: These findings provide a rationale for the development of a targeted therapy for methylation-sensitive colon cancer that can include EGCG in combination with other DNMT and HDAC inhibitors.

Keywords: DNMT3A; EGCG; Epigenetics; HDAC3; colon cancer; epigallocatechin 3-gallate; green tea.

Conflict of interest statement

Conflict of Interest Statement

None Declared.

Figures

Figure 1
Figure 1
Epigallocatechin 3-gallate reduces expression of some DNA methyltransferase (DNMT) transcripts. DNMT1, -3A and -3B transcript levels were reduced in HCT 116 cells at 72 hours at 150 μM EGCG (A). All studied transcripts were significantly reduced at 72 hours in HT-29 cells at the lower concentration of 50 μM (B). Error bars represent +/− one standard deviation (n=3). *p<0.05, **p<0.02, ***p<0.01 compared with the control.
Figure 2
Figure 2
DNMT3A protein levels decrease in response to EGCG in HCT 116 cell line while expression in HT-29 cells remained constant at 48 hours. Response to EGCG treatment is both time and dose- dependent in HCT 116 cells. There is no change in DNMT3a protein in response to EGCG in HT-29 cells at 48 hours but a significant decrease at 100 and 150 μM at 72 hours. Error bars represent +/− one standard deviation.
Figure 3
Figure 3
Histone deacetylase 1, -2, and -3 transcript levels decreased in HCT 116 and HT-29 cell lines when treated with EGCG. Significant results are seen at the higher concentration of 150 μM. Error bars represent +/− one standard deviation (n=3). *p<0.05, **p<0.02, ***p<0.01 compared with the control.
Figure 4
Figure 4
Protein expression of histone deacetylase 2 and -3 significantly decreased in a time- and dose- dependent manner in HCT 116 cells in response to EGCG treatment, while HDAC1 expression remained relatively constant except at the highest EGCG dosage and after 72 hours. In HT-29 cells, protein expression for all targets remained relatively constant.
Figure 5
Figure 5
Treatment with cycloheximide to inhibit protein synthesis lends decreased expression of histone deacetylase 3 and DNA methyltransferase 3A in extracts from HCT 116 cells co-treated with 100 μM EGCG in (5A). HT-29 cells co-treated with EGCG and cycloheximide showed no difference in protein expression (5B).
Figure 6
Figure 6
Immunoprecipitation of UHRF1 reveals an association of both histone deacetylase 3 and DNA methyltransferase 3A. HCT 116 cells treated with 100 μM EGCG exhibitied a decrease in association between UHRF1 and both HDAC3 and DNMT3a (6A). In HT-29 cells there is only a decerased association between UHRF1 and DMT3a in response to EGCG (6B).

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