Alterations of androgen receptor-regulated enhancer RNAs (eRNAs) contribute to enzalutamide resistance in castrationresistant prostate cancer

Enzalutamide is a second-generation anti-androgen for treatment of castrationresistant prostate cancer (CPRC). It prolongs survival of CRPC patients, but its overall survival benefit is relatively modest (4.8 months) and by 24 months most patients progress on enzalutamide. To date, however, the molecular mechanisms underlying enzalutamide resistance remain elusive. Herein, we report enzalutamide treatmentinduced alterations of androgen receptor (AR)-regulated enhancer RNAs (AR-eRNAs) and their roles in enzalutamide-resistant growth and survival of CRPC cells. AR chromatin immunoprecipitation and high throughput sequencing (ChIP-seq) and RNAseq analyses revealed that 188 and 227 AR-eRNAs were differentially expressed in enzalutamide-resistant LNCaP and C4-2 cells, respectively. The AR-eRNAs upregulated in C4-2 cells and downregulated in LNCaP cells were selected through meta-analysis. Expression of AR-eRNAs and related mRNAs in the loci of FTO, LUZP2, MARC1 and NCAM2 were further verified by real-time RT-PCR. Silencing of LUZP2 inhibited, but silencing of MARC1 increased the growth of enzalutamide-resistant C4-2 cells. Intriguingly, meta-analysis showed that expression of LUZP2 mRNA increased in primary tumors compared to normal prostate tissues, but decreased again in metastatic CRPC. Our findings suggest that eRNA alteration profiling is a viable new approach to identify functional gene loci that may not only contribute to enzalutamide-resistant growth of CRPC, but also serve as new targets for CRPC therapy.