The long non-coding RNA <i>NEAR1</i> promotes neuroendocrine prostate cancer cell proliferation and survival

Neuroendocrine prostate cancer (NEPC) is an incurable, androgen receptor (AR)-negative disease that originates from AR-positive adenocarcinomas. Loss or mutation of the AR renders androgen deprivation therapy ineffective and presents a therapeutic problem. Currently, incidence of NEPC is rising and new therapeutic strategies are urgently needed. LncRNAs are some of the most differentially regulated transcripts in cancer, but are largely unexplored. To investigate the expression of lncRNAs in NEPC, we used patient-derived xenografts (PDXs) developed at the Living Tumour Laboratory (LTL). These PDXs retain the histology, genetic, and epigenetic features of their human counterparts. For this study, we employed the first-in-field animal model of trans-differentiation from hormone-sensitive prostate adenocarcinoma (LTL331) to NEPC (LTL331R). Using RNASeq we generated a shortlist of lncRNAs which were up-regulated in NEPC (fold change>4, FDR<.01), met a basal expression threshold, and had no coding potential. Based on these criteria, 26 lncRNAs were shortlisted (from 15,224). We then confirmed the expression of 7 lncRNAs in a panel of four prostate cell lines using qPCR (RWPE-1 (non-neoplastic); LNCaP (AR+); DU145 (AR-) and PC-3 (AR- and NEPC-like). From these data the lncRNA, neuroendocrine associated lncRNA 1 (NEAR1), was selected as the most suitable target. To determine the clinical relevance of NEAR1, publically available data from CBioPortal were used. NEAR1 was found to be up-regulated in NEPC samples compared to adenocarcinomas (p= 0.0003). Expression was also investigated by microarray analysis in several other LTL models and was found to be significantly up-regulated in NEPC (but not CRPC) compared to hormone-sensitive adenocarcinoma (p= 0.0161). We then determined the expression of NEAR1 in a panel of non-neoplastic tissues by qPCR and found an association with tissues of endodermal origin. However, the expression of NEAR1 observed in NEPC samples was higher than any non-neoplastic tissue tested. For functional studies of NEAR1 we employed three small-interfering RNAs (siRNAs) and one scrambled control (NC). Knockdown with siRNAs 1 and 2 significantly reduced cell proliferation (MTT assays carried out over 7 days) compared to the NC in proportion to the degree of target knockdown. Caspase 3/7 activity also significantly increased with siRNA 1 (p=0.0001) and siRNA 2 (p=0.0036) knockdown compared to the NC. These data suggest NEAR1 may represent a potential therapeutic target for NEPC