Figure 2. AR protein and gene regulation differs in WT versus mutant ER expressing MCF-7 breast cancer cells and patient derived xenografts (PDX), particularly after long-term estrogen deprivation (LTED).

A. Western blot of whole cell lysates from WT, D538G, and Y537S MCF7 cells grown in full serum media (left) or charcoal stripped serum containing media (LTED, right) were probed for AR, ER, GR, PR-A/B, and tubulin. B. ER+ patient-derived xenografts (PDX) tissue microarray from Dr. Alana Welm, Huntsman Cancer Institute (HCI) and ER+ PDX from University of Colorado (UCD) with ESR1 mutation status indicated, were stained for AR by IHC with representative images at 100X (left) and 400X (right). For the estrogen independent version of HCI-013, HCI-013-EI, an image representative of primary tumor (PT) and lung metastasis (Met) are shown. C. Heatmap of known AR regulated genes in MCF7 (left) and T47D (right) depicting relative expression of genes displayed as a Z-score across WT and mutant cells after 5 days of growth in hormone-depleted media followed by treatment with either 10nM E2 or vehicle (DMSO) for 8 hours. Significant overlap was determined using hypergeometric tests with a p-value cut off of <0.05: AR genes vs D538G up-regulated genes = 5.5x10⁻⁰⁵; AR genes vs D538G down-regulated genes = 1.0x10⁻⁰⁸; AR genes vs Y537S up-regulated genes = 3.2x10⁻⁰⁸; AR genes vs Y537S down-regulated genes = 9.3x10⁻¹¹.