Fig. 5.

AR agonist treatment blocked Y537S mutant metastasis. A. Immunoblot analysis of MCF-7 parental (n=4) and YS1 (n=5) primary /metastatic pairs grown with E₂ withdrawal. B. Western blot analysis of MCF-7 parental, YS1, DG, and YS30 clones cultured in 5% CSS media for 4 days GAPDH was used as loading control. C. Mice with WHIM20 PDX tumors were supplemented with E₂, and randomized to –E₂, -E₂+DHT or –E₂+Enz for total of 4 months. Time to tumor doubling was calculated using Kaplan-Meier method. Data were analyzed using Log-rank (Mantel-Cox) Test. P= 0.10 for DHT vs vehicle control and P= 0.07 for MDV vs vehicle control. D. Macrometastasis frequency was calculated and displayed chi-square and fisher exact analyses, * P =0.002. E and F. ER and PR total score IHC staining two-sided Student’s t-test was performed to test significance. *P <0.05, **P <0.01,* P <0.001, NS=Not significant. G. Western blot analyses of WHIM20 primary tumors; GAPDH was used as a loading control. H. Western blot analyses of MCF-7 YS1 cells; performed using in vitro cell line protein lysis and GAPDH was used as a loading control. Cells were starved for 48 hours in 5% CSS, and then treated with hormones for 24 hours.