Fig 1: Screening and validation of potential PR inducers selected by High throughput screening of FDA approved drug library. (A) Representative mCherry signal in the 3D cultured PR reporter gene-expressing cells treated with FDA approved drugs at 1µM. Romidepsin (100 nM) serves as positive control. (B) High throughput screening of the FDA approved drug library (Selleck Chemicals, Houston, TX). a: Scatter plot of all compounds in the screening. Vertical axis: the mCherry intensity (arbitrary unit as sum of mCherry intensity per well). Horizontal axis: area of the spheroids (mean µm2/well). Green as negative control (mock treatment), Red as positive control (romidepsin at 1 µM), and Blue as the compounds from the library. ?: Hits for mCherry intensity increase. ?: Hits for area of the spheroid decrease. ?: Hits for both mCherry intensity increase and area of the spheroid decrease. b: Histogram of mCherry intensity. c: Histogram of area of the spheroids. (C) mRNA expression of PAEP, total PR (PRA/B), and PRB after being treated with the initially selected FDA drugs. Cells were treated for 24 h with Carfilzomib (PR-171) [100 nM], Ponatinib (AP24534) [500 nM], alexidine hydrochloride [100 nM], CUDC-907 [20 nM], AR-42 [500 nM], Camptothecin [100 nM], Teniposide (Vumon) [500 nM], romidepsin (20 nM), daunorubicin (100 nM), or 1 µM of other drugs. (D,F) mRNA expression of PR and its downstream genes after the treatment with the 5 top-picked drugs, Carfilzomib [100 nM], CUDC-907 (20 nM), AR-42 (500 nM), romidepsin (20 nM), and daunorubicin (100 nM) for 24 h in Ishikawa cells (E) or in ECC1 cells (G). (E,G) Western blotting of PR and its downstream genes treated with 5 top-picked drugs for 24 h in Ishikawa cells (F) or in ECC1 cells (H). * p < 0.05 vs. control.