RNA-seq Analysis of Prostate Cancer Cell Lines

GE of all prostate cancer and normal prostate cell lines at baseline (no treatment) was assessed by RNA-seq. Furthermore, the effects of METRO-TOPO and CONV-TOPO exposure for 6 weeks on AR^Low mCRPC/NEPC PC-3 tumor model (3D spheroid) were assessed using RNA-seq. Pre- and post-drug exposure, as described above, tumor cells were harvested, and high-quality RNA was extracted using QIAshredder and RNeasy kit (Qiagen). RNA concentration and integrity were assessed using a Nanodrop-8000 spectrophotometer and Agilent 2100 Bioanalyzer. An RNA integrity number threshold >8 was applied, and RNA-seq libraries were constructed using Illumina TruSeq RNA Sample Preparation kit v2. Libraries were then size-selected to generate inserts of approximately 200 bp. RNA-seq was performed on Illumina's NovaSeq platform using a 150 bp paired-end protocol with a depth of >20 million reads per sample. Average quality scores were above Q30 for all libraries in both R1 and R2 (29 (link)). AA cell line RNA was isolated from cultured cells using TRIzol Reagent (Sigma Life Sciences) following the manufacturer's protocol (27 (link)). Library preparation, quality control, and sequencing of extracted RNA were performed by the Center for Pharmacogenomics and Single-Cell Omics (AUPharmGX).