The long noncoding RNA landscape of neuroendocrine prostate cancer and its clinical implications

Abstract

BackgroundTreatment-induced neuroendocrine prostate cancer (tNEPC) is an aggressive variant of late-stage metastatic castrate-resistant prostate cancer that commonly arises through neuroendocrine transdifferentiation (NEtD). Treatment options are limited, ineffective, and, for most patients, result in death in less than a year. We previously developed a first-in-field patient-derived xenograft (PDX) model of NEtD. Longitudinal deep transcriptome profiling of this model enabled monitoring of dynamic transcriptional changes during NEtD and in the context of androgen deprivation. Long non-coding RNA (lncRNA) are implicated in cancer where they can control gene regulation. Until now, the expression of lncRNAs during NEtD and their clinical associations were unexplored.ResultsWe implemented a next-generation sequence analysis pipeline that can detect transcripts at low expression levels and built a genome-wide catalogue (n = 37,749) of lncRNAs. We applied this pipeline to 927 clinical samples and our high-fidelity NEtD model LTL331 and identified 821 lncRNAs in NEPC. Among these are 122 lncRNAs that robustly distinguish NEPC from prostate adenocarcinoma (AD) patient tumours. The highest expressed lncRNAs within this signature are H19, LINC00617, and SSTR5-AS1. Another 742 are associated with the NEtD process and fall into four distinct patterns of expression (NEtD lncRNA Class I, II, III, and IV) in our PDX model and clinical samples. Each class has significant (z-scores >2) and unique enrichment for transcription factor binding site (TFBS) motifs in their sequences. Enriched TFBS include (1) TP53 and BRN1 in Class I, (2) ELF5, SPIC, and HOXD1 in Class II, (3) SPDEF in Class III, (4) HSF1 and FOXA1 in Class IV, and (5) TWIST1 when merging Class III with IV. Common TFBS in all NEtD lncRNA were also identified and include E2F, REST, PAX5, PAX9, and STAF. Interrogation of the top deregulated candidates (n = 100) in radical prostatectomy adenocarcinoma samples with long-term follow-up (median 18 years) revealed significant clinicopathological associations. Specifically, we identified 25 that are associated with rapid metastasis following androgen deprivation therapy (ADT). Two of these lncRNAs (SSTR5-AS1 and LINC00514) stratified patients undergoing ADT based on patient outcome.DiscussionTo date, a comprehensive characterization of the dynamic landscape of lncRNAs during the NEtD process has not been performed. A temporal analysis of the PDX-based NEtD model has for the first time provided this dynamic landscape. TFBS analysis identified NEPC-related TF motifs present within the NEtD lncRNA sequences, suggesting functional roles for these lncRNAs in NEPC pathogenesis. Furthermore, select NEtD lncRNAs appear to be associated with metastasis and patients receiving ADT. Treatment-related metastasis is a clinical consequence of NEPC tumours. Top candidate lncRNAs FENDRR, H19, LINC00514, LINC00617, and SSTR5-AS1 identified in this study are implicated in the development of NEPC. We present here for the first time a gen...

Alternative RNA splicing of the GIT1 gene is associated with neuroendocrine prostate cancer

“…Whole‐transcriptome datasets of Beltran 2011, Beltran 2016, and VPC 2018 patient cohorts, LTL PDX models, and LnNE RNA‐sequenced datasets were analyzed to determine the expression of GIT1 splice variants in the various prostate cancer models. The software Integrative Genomics Viewer was used to visualize the coverage of RNA‐seq reads corresponding to the GIT1 gene.…”

“…Currently, there are no commercially available GIT1 antibodies to specifically detect each splice variant. As a result, we used RISH, a well‐established alternative technique, on a TMA to study and confirm the expression of GIT1‐A and GIT1‐C . We created probes to target the unique exon 7/8 junction of GIT1‐A , as well as the exon 7/9 junction of GIT1‐C , to detect RNA expression levels on the TMA.…”

“…Human CRPC TMA contains 64 cores: 52 AdPC, six AdNC, and six SCNC. Details of these prostate cancer subtype classifications have been previously published by our group . Briefly, AdPC is classified by the presence of glandular structures, large cells with a prominent nucleolus, and no NE cells.…”

“…RNA‐seq datasets from the Beltran 2011, Beltran 2016, and Vancouver Prostate Centre (VPC) 2018 clinical cohorts were used. Cohort composition for the Beltran 2011, Beltran 2016, and VPC 2018 was seven NEPC and 30 AdPC, 15 NEPC and 34 CRPC‐Ad, and five NEPC and 24 AdPC, respectively.…”

“…Red dots indicate positive RISH signals under 10× and 40× magnification. IHC assays on the CRPC TMA and scoring methods for RISH or IHC were carried out, as previously described . Briefly, RISH scores of 0, 1, and 2 indicate no positive signal, ≤20% positive signal, and >20% positive signal of cells within a tissue core, respectively.…”