Fig. 1: Prostate cancer cells respond differently to different doses of androgens.

LNCaP (a) or VCaP (b) cells plated in media containing charcoal-stripped serum (CFS) or full-serum (FBS) (c, d) were treated with increasing concentrations of R1881. Cell growth was assessed by DNA quantification at day 7. Data are shown as mean ± SD as representative results from three independent experiments, n = 3 wells of cells. RFU relative fluorescence units. Castrated male NSG mice bearing subcutaneous LNCaP tumors were administered increasing doses of testosterone in (e) (Placebo, n = 10 mice; 0.5 mg/kg/d, n = 9 mice; 1 mg/kg/d, n = 10 mice; 2 mg/kg/d, n = 10 mice; 4 mg/kg/d, n = 9 mice; 8 mg/kg/d, n = 9 mice) or DHT in (f) Placebo, n = 10; 0.1 mg/kg/d, n = 10 mice; 0.2 mg/kg/d, n = 10 mice; 0.4 mg/kg/d, n = 10 mice; 0.8 mg/kg/d, n = 10 mice. Data are shown as mean ± SD. p values were determined by two-way ANOVA followed by Tukey’s multiple comparison test (P < 0.0001 for (e) and P < 0.0004 for (f)). g Significant Hallmark pathways of selected Gene Set Enrichment Analysis (GSEA) differentially regulated in VCaP cells in response to low (0.06 nM R1881) as compared to high dose (R1881 10 nM) androgens are represented (FDR q value < 0.001; NES normalized enrichment score). NES < 0 represents downregulation of specified pathway in LD vs HD. NES > 0 represents upregulation of specified pathway in LD vs HD. h VCaP cells plated in CFS-supplemented media were treated with increasing concentrations of R1881 for 24 h. The mRNA expression for PSA and E2F1 were assessed using qRT-PCR. Data are shown as mean ± SD as representative results from three independent experiments, n = 3 technical replicates. i Heatmap presentation of gene expression as assessed using the Nanostring nCounter platform from VCaP cells treated in CFS-supplemented media with either vehicle (Veh), LD or HD R1881. j VCaP cells plated in media supplemented with CFS were treated with increasing concentrations of either R1881, Testosterone (T) or Dihydrotestosterone (DHT) for 48 h. Whole cell extracts were probed for RB1 phosphorylation (pRB), E2F1, FOXM1, PSA or β-Actin using western blot. Representative images are shown from n = 3 biologically independent experiments. Source data are provided as a Source Data file.