Teine sulfinic acid, while it may promote apoptosis by increasing reactive oxygenPLOS ONE | DOI:10.1371/journal.pone.0157866 June 29,15 /Bayer 41-4109 chemical information methylation Landscape of Breast RG7666 web cancer Cells in Response to ResveratrolFig 8. DNA methylation and mRNA expression of selected genes during course of time. Schematic representation of methylation status and mRNA levels of eight modulated genes in MDA-MB-231 cells treated for 24 h and 48 h with resveratrol (100 M) relative to control non-treated cells. The y-axis represents the methylation levels of gene promoters (peak score) and the differential fold change in mRNA expression relative to control. Black bars indicate the methylation level of specific gene promoters. White bars indicate the gene expression level as obtained from transcriptome analysis [24]. doi:10.1371/journal.pone.0157866.gspecies through suppression of glutathione generation. CDO1 plays a tumor suppressive role in human carcinogenesis and is frequently inactivated by promoter methylation in breast, esophagus, lung, bladder, gastric and colorectal tumors [38]. These data suggested that AURKA, CCNB1, HK2, SOX17, SLIT3 and CDO1 might represent novel potential targets forPLOS ONE | DOI:10.1371/journal.pone.0157866 June 29,16 /Methylation Landscape of Breast Cancer Cells in Response to Resveratrolepigenetic therapy in breast cancer. In agreement with a recent report here we identified hypomethylated oncogenes (HK2, FGFR4, DNMT3A) and methylated tumor suppressor genes (CDO1, SLIT3, and SOX17), which were previously identified as candidate genes that comprise part of the emerging “cancer methylome” from 22 cancer cell lines derived from several cancer types as lung, breast, colon, liver, skin, prostate, and cervical cancer, among others [20?2]. On the other hand, we found a good correlation between DNA methylation and gene expression changes in a specific set of cancer-related genes that further highlighted the biological significance of resveratrol-induced methylation alterations in breast cancer cells. These concordant changes in DNA methylation and gene expression were found in oncogenes (AURKA, CCNB1, DDIT4, DLGAP5, EYS, FAM83D, HIST1H2BM, IL24, LPXN, NFIL3, PFKFB3, SLC14A1, STC1, GPR110, HIST1H3F, HK2, MMP9, NFIL3, PSMD11, RUNX2, SH3KBP1) and tumor suppressor genes (AMY2A, IL18, SLIT3, MPHOSPH9, SLC27A2, TMOD2, TTI1 and XYLB). In contrast, other genes such as PEG10 showed an inverse correlation between promoter DNA methylation and gene expression suggesting that additional mechanisms of gene regulation are operating in response to resveratrol. Finally, although our study demonstrates a systematic and compelling effect of resveratrol on DNA methylation in breast cancer cells, data requires further animal and human studies in order to validate in vivo these results. Although speculative, our findings permit us to propose that resveratrol may modulate gene expression and exert anti-proliferative activities based on its ability to modify the DNA methylation of well-known cancer genes suggesting that it may be useful as a novel epigenetic therapeutic tool. Additional in vivo studies are needed to evaluate the precise molecular mechanisms of resveratrol-mediated methylation changes and estimate the potential of resveratrol for inducing epigenetic modulations in preclinical models of breast cancer.Supporting InformationS1 Table. Methylation raw data for MDA-MB-231 control cells. (XLS) S2 Table. Methylation raw data for MDA-MB-231 cells treated.Teine sulfinic acid, while it may promote apoptosis by increasing reactive oxygenPLOS ONE | DOI:10.1371/journal.pone.0157866 June 29,15 /Methylation Landscape of Breast Cancer Cells in Response to ResveratrolFig 8. DNA methylation and mRNA expression of selected genes during course of time. Schematic representation of methylation status and mRNA levels of eight modulated genes in MDA-MB-231 cells treated for 24 h and 48 h with resveratrol (100 M) relative to control non-treated cells. The y-axis represents the methylation levels of gene promoters (peak score) and the differential fold change in mRNA expression relative to control. Black bars indicate the methylation level of specific gene promoters. White bars indicate the gene expression level as obtained from transcriptome analysis [24]. doi:10.1371/journal.pone.0157866.gspecies through suppression of glutathione generation. CDO1 plays a tumor suppressive role in human carcinogenesis and is frequently inactivated by promoter methylation in breast, esophagus, lung, bladder, gastric and colorectal tumors [38]. These data suggested that AURKA, CCNB1, HK2, SOX17, SLIT3 and CDO1 might represent novel potential targets forPLOS ONE | DOI:10.1371/journal.pone.0157866 June 29,16 /Methylation Landscape of Breast Cancer Cells in Response to Resveratrolepigenetic therapy in breast cancer. In agreement with a recent report here we identified hypomethylated oncogenes (HK2, FGFR4, DNMT3A) and methylated tumor suppressor genes (CDO1, SLIT3, and SOX17), which were previously identified as candidate genes that comprise part of the emerging “cancer methylome” from 22 cancer cell lines derived from several cancer types as lung, breast, colon, liver, skin, prostate, and cervical cancer, among others [20?2]. On the other hand, we found a good correlation between DNA methylation and gene expression changes in a specific set of cancer-related genes that further highlighted the biological significance of resveratrol-induced methylation alterations in breast cancer cells. These concordant changes in DNA methylation and gene expression were found in oncogenes (AURKA, CCNB1, DDIT4, DLGAP5, EYS, FAM83D, HIST1H2BM, IL24, LPXN, NFIL3, PFKFB3, SLC14A1, STC1, GPR110, HIST1H3F, HK2, MMP9, NFIL3, PSMD11, RUNX2, SH3KBP1) and tumor suppressor genes (AMY2A, IL18, SLIT3, MPHOSPH9, SLC27A2, TMOD2, TTI1 and XYLB). In contrast, other genes such as PEG10 showed an inverse correlation between promoter DNA methylation and gene expression suggesting that additional mechanisms of gene regulation are operating in response to resveratrol. Finally, although our study demonstrates a systematic and compelling effect of resveratrol on DNA methylation in breast cancer cells, data requires further animal and human studies in order to validate in vivo these results. Although speculative, our findings permit us to propose that resveratrol may modulate gene expression and exert anti-proliferative activities based on its ability to modify the DNA methylation of well-known cancer genes suggesting that it may be useful as a novel epigenetic therapeutic tool. Additional in vivo studies are needed to evaluate the precise molecular mechanisms of resveratrol-mediated methylation changes and estimate the potential of resveratrol for inducing epigenetic modulations in preclinical models of breast cancer.Supporting InformationS1 Table. Methylation raw data for MDA-MB-231 control cells. (XLS) S2 Table. Methylation raw data for MDA-MB-231 cells treated.
