All the mice were housed in the Animal Resource Facil ity of the University of thoroughly Alabama at Birmingham and were maintained under the following conditions 12 h dark/12 h light cycle, 24 2 C temperatures, and 50 10% humidity. Animal experimental designs Protocol 1. Tumor xenografts assay for treatment effects of GE After one week Inhibitors,Modulators,Libraries of acclimatization, Nu/Nu Nude mice were randomly divided into four groups and administered either control or GE diet as described above. Diets were provided from two weeks prior to in jection Inhibitors,Modulators,Libraries and the mice continued to receive the corre sponding experimental diets throughout the study. To determine the in vivo efficacy of GE on ER re activation and subsequent chemosensitization to estro gen antagonist, TAM, in human ER negative breast tumor xenografts, exponentially growing MDA MB 231 cells were mixed at a 1 1 ratio with Matrigel.

A 100 ul suspension containing 1 106 cells was injected orthotopically into the mammary fat pad of each mouse. The experimental groups were as follows Group. Control group Mice were fed with control diet as described previously. Group. GE group Mice were fed with GE diet . Group. TAM group Mice were fed with control Inhibitors,Modulators,Libraries diet plus TAM treatment for 3 wks after two wks of post injection . Group. GE TAM group Mice were fed a GE diet and received TAM treatment as described above. Protocol 2. Spontaneous breast cancer mouse model for preventive effects of GE The C3 SV40 Tag transgenic mouse model was used for prevention study of GE treatment because this mouse model can spontaneously develop breast cancer.

More importantly, this model tends to develop hormone independent invasive breast cancer, which is perfectly suitable to our in vestigation purpose for ER reactivation. The Tag genotypes were identified at 21 days of life by analysis of tail DNA using standard PCR techniques according to Inhibitors,Modulators,Libraries previous studies. The C3 SV40 Tag mice at 4 6 weeks of age were randomly divided to different experi mental groups and control and GE diets were administered at the indicated time and the diets were continued throughout the study. The experimental groups were as follows Group. Control group Mice were fed control diet as described previously. Group. GE group Mice were fed GE diet as described previously. Group. TAM group Mice were fed control diet and TAM tablet was implanted subcutane ously for 3 wks when tumor size reaches 400 mm3.

. GE TAM group Mice were administered with GE diet and TAM treatment as described above. Inhibitors,Modulators,Libraries Tumor parameters monitoring, experimental endpoint and tissue sample collection Tumor diameters and body weight were measured weekly. Tumor volumes were measured by a caliper and estimated using the following formula Dorsomorphin mw tumor volume 0. 523. For Protocol 1, the experiment was finished when the mean of tumor diameter in the control mice exceeded 1. 0 cm following the guidelines of Institutional Animal Care and Use Committee at the University of Alabama at Birmingham.