Differential expression profiling of the TamR cells following AR knockdown

The androgen receptor (AR) has emerged as a candidate target for the treatment of breast cancer. In this study, we sought to characterize the functional consequences of AR knockdown using a siRNA-mediated approach in a tamoxifen-resistant (TamR) derivative of MCF7 breast cancer cell line. We evaluated the role of AR in endocrine-resistance using tamoxifen resistant (TamR) cells derived from the MCF7 breast cancer cell line. The TamR line was continuously cultured in RPMI 1640 media (Thermo Fisher) supplemented with 10% fetal bovine serum (GE Healthcare), 20 mM HEPES (Thermo Fisher) and 0.28 IU/ml insulin (Novo Nordisk). Base media for TamR cells was also supplemented with 5 µM 4-hydroxytamoxifen (Sigma). Gene expression profiling was performed on the TamR cells 48 hours post transfection with nonsense (NS) control or AR siRNA (5'-GGAACUCGAUCGUAUCAUU-3' , Thermo Fisher Silencer® Select Pre-Designed siRNA #4390824) using 3 biological replicates. Microarray hybridization was performed on the Affymetrix PrimeView™ Human Gene Expression Array following the manufacturer's protocol. CEL files were further processed in the R software (v 3.4.3) environment using affy (Gautier et al., 2004) and limma (Ritchie et al., 2015) packages from Bioconductor.