We hypothesize that dietary GE may have a similar effect on ER expression since both compounds are considered to mainly exert their Inhibitors,Modulators,Libraries anticancer properties via epigenetic control. We initiated our study to determine whether GE can impact ER expression and the optimal dose and time point that will induce ER activation. We treated ER negative breast cancer cells, MDA MB 231, with various concentrations of GE at different time points and observed ER transcription under these treatments. As shown in Figure 1A, Inhibitors,Modulators,Libraries a sig nificant increase of ER transcription was observed with 25 uM of GE and the ER reactivation was predominant at 3 days of treatment. This GE con centration is considered to be equivalent to the maximal consumption of soybean product per day or a pharma ceutically available GE supplementary tablet, suggesting a potential bioavailability of this treatment.

This result indicates that treatment with 25 Inhibitors,Modulators,Libraries uM GE at 3 days could serve as an optimal condition in regulating ER re expression in ER negative breast cancer cells. We also tested combination effects of GE with other epigenetic modulators such as the histone dea cetylase inhibitor, trichostatin A, and a demethylation agent, 5 aza 2 deoxycytidine, on ER re expression because epigenetic mechanisms such as histone modifications and DNA methylation were known to contribute to ER regulation. Both TSA and 5 aza have been reported to successfully acti vate ER transcription in human ER negative breast Inhibitors,Modulators,Libraries cancer cells, but have not previously been com bined with GE in ER studies.

Inhibitors,Modulators,Libraries Consistent with previous studies, our results indicated that 5 aza and TSA alone reactivated ER expression in MDA MB 231 cells. More importantly, we found that the combined treat ment of GE and TSA induced a significant synergistic effect on ER re expression, much more so than GE in combination with 5 aza. This effect was further confirmed by the results of ER protein levels in Figure 1E showing that combination treatment using GE and TSA led to more abundant ER re expression than the other treatments administered alone. To further verify the GE effects on ER reactivation on an ER negative breast cancer cell line other than MDA MB 231 cells, we performed similar experiments on ER negative MDA MB 157 cells. We found a dose dependent effect of ER up regulation in response to GE treatment and combin ation treatment of 25 uM of GE with TSA but not 5 aza resulted in a synergistic effect on ER reactivation. This similar response to GE treatment as seen in MDA MB 231 cells suggests that this combination regimen results in a prevalent effect cisplatin dna on ER reactivation in different ER negative breast cancer cells as well.