Analysis of novel enzalutamide-resistant cells: Upregulation of testis-specific Y-encoded protein gene promotes the expression of androgen receptor splicing variant 7

Abstract

Background: Enzalutamide, a second-generation antiandrogen, is an approved medicine for the treatment of metastatic castration-resistant prostate cancer (CRPC); however, the mechanisms behind the resistance are not completely understood. In the present study, we established enzalutamide-resistant cells derived from lymph node carcinoma of the prostate (LNCaP) cells and characterized their androgen receptor (AR) status and changes in the gene expression with an aim to elucidate these mechanisms. Methods: SAS MDV No. 3-14 enzalutamide-resistant cells were established from LNCaP xenograft castrated male mice under continuous administration of enzalutamide. Then, the AR status and expression of AR target genes were evaluated by western blotting or real-time polymerase chain reaction analysis. The role of AR in the proliferation was also analyzed using the AR siRNA approach. The gene expression profiling in SAS MDV No. 3-14 cells was evaluated by microarray analysis. The role of testis-specific Y-encoded protein (TSPY), one of the upregulated genes, in the expression of AR and AR target genes and cell growth was also verified using siRNA. Results: SAS MDV No. 3-14 cells expressed AR-v7, leading to the increased expression of AR target genes. Gene silencing of AR showed that both AR-FL and AR-v7 function as proliferative drivers in SAS MDV No. 3-14 cells. Microarray analysis revealed that TSPY is upregulated genes in these cells. TSPY siRNA inhibited cell proliferation, decreased the expression of AR-v7 and AR-v7 targeted genes. Conclusions: This study demonstrated that SAS MDV No. 3-14 cells increase the expression of AR-v7 by upregulating TSPY, leading to acquired resistance to enzalutamide.