Spr4594

T47D and BT474 cells were acquired from ATCC and cultured in DMEM (Cellgro) supplemented with 5% FBS and 1% penicillin/streptomycin. PR, STAT1, and STAT2 shRNA knockdown cells were created using viral particles (GE/Dharmacon) targeting three different regions of each respective gene. Viral transduction protocol was followed as per manufacturer’s instructions. Transduced, stable cell line pools expressing NS, PR, STAT1, or STAT2 shRNA were created in T47D cells following 14 d of selection in 2.5 ug/ml Puromycin (MP Biomedicals). Target shRNA sequences are listed in Supplementary Table 1. RFP and STAT1 overexpressing cells were created using lentiviral ORF particles (GE/Dharmacon). Transduced, stable cell line pools overexpressing RFP (control) or STAT1 were created in T47D cell lines stably expressing NS or STAT2 shRNA following 14 d of selection in 15 ug/mL Blasticidin S HCL (Corning). Sequences are shown in Supplementary Table 1. Cells were treated with the following reagents where indicated: R5020 (10 nM, Sigma), human rIFN-α (1000 IU/mL, IFN-α2A, SPR4594; Sigma-Aldrich, vehicle-H20), onapristone (10uM, provided through an MTA with Context Therapeutics), cycloheximide (200ug/mL, Calbiochem), and MG132 (5uM, Selleck Chemical).