Fig. 6

Pharmaceutical inhibition of Akt suppresses epithelial-mesenchymal transition and cell proliferation preventing Brca1-deficient tumor progression. a p18^−/−;Brca1^MGKO tumor cells were treated with dimethyl sulfoxide (DMSO) or 5 nM E2 in the presence or absence of different dosage of AZD5363 for 2 h, the expression of p-mTor, p-4E-bp1, p-Gsk3β, p-Fra1 and vimentin (Vim) were analyzed by western blot. b p18^−/−; Brca1^MGKO tumor cells were treated with DMSO or 5 nM E2 in the presence or absence of different dosage of AZD5363, and the numbers of viable cells were determined on day 1, day 3, and day 5. *p < 0.05 between E2-treated and E2 + AZD5363-treated groups at the time points (Student t test). Data are represented as mean ± SD (n = 4). c p18^−/−;Brca1^MGKO tumor cells were treated with 5 nM E2 with or without 1 μM AZD5363 for 24 h, and bromodeoxyuridine (Brdu) incorporation was then determined by fluorescence-activated cell sorting (FACS). d-g We transplanted 1 × 10⁶ p18^−/−; Brca1^MGKO tumor cells into the mammary fat pads of NSG mice along with E2 pellet under the skin, and tumors were allowed to reach ~ 250 mm³ in size. Mice were then treated with AZD5363 at 150 mg/kg body weight or vehicle daily by oral gavage. The tumor size was determined and plotted (d). Data in d are represented as mean ± SD of four tumors in each group. *p < 0.05 between two groups at each time point (Student t test). p18^−/−; Brca1^MGKO tumors treated with AZD5363 or vehicle for 7 days (d) were analyzed by western blot (e), immunohistochemical staining (f), and immunofluorescent staining (g). Samples in e were derived from eight different tumors developed in eight individual mice. The percentages of Ki67⁺ and/or Ck14⁺ cells (g) were quantitated in four randomly selected fields for each section of a tumor, and the results represent the mean ± SD of three individual tumors per group