Abstract C36: Transcriptome sequencing identifies PCAT-1, a novel lncRNA implicated in prostate cancer progression and therapeutic response to PARP inhibitors

Abstract

Abstract High-throughput sequencing of polyA+ RNA (RNA-Seq) in human cancer shows remarkable potential to identify both novel disease-specific markers for clinical uses and uncharacterized aspects of tumor biology, particularly long non-coding RNA (lncRNA) species >250 bp in length. To illustrate this approach, we employed RNA-Seq on a cohort of 102 prostate tissues and cell lines and performed ab initio transcriptome assembly to discover unannotated ncRNAs with cancer-specific expression patterns. In total, we observed that ∼20% of the prostate cancer transcriptome represents unannotated, polyadenylated RNA species that were enriched for activate chromatin marks (H3K4me3, RNA polymerase II) defined by ChIP-Seq data. From 1,859 intergenic lncRNAs overall, we nominated 121 such Prostate Cancer Associated Transcripts (PCATs) that collectively performed as well as known biomarkers in differentiating benign, cancerous, and metastatic tissues. Among these, we identified an uncharacterized two-exon, polyadenylated lncRNA, PCAT-1, as a prostate cancer outlier and a prostate-specific driver of cancer cell proliferation. Mechanistically, microarray profiling following modulation of PCAT-1 expression in vitro defined a core set of genes repressed by PCAT-1, identifying this lncRNA as a transcriptional repressor of genes implicated in DNA maintenance processes, such as CENPF and BRCA2. Interestingly, when PCAT-1 expression is low, it is negatively regulated by the Polycomb Repressive Complex 2 (PRC2), which directly binds the PCAT-1 promoter in vitro. Supporting this, human tumor samples with high PCAT-1 or high PRC2 expression were exclusive and defined distinct molecular subtypes distinguished by gene expression signatures of PCAT-1-repressed target genes. PRC2-based repression of PCAT-1 was reversible by inhibition of PRC2. BRCA2 inactivation is known to sensitize cancer cells to small molecular inhibitors of the PARP1 DNA repair enzyme by impairing homologous recombation of double-stranded DNA breaks, suggesting that PCAT-1 expression may sensitize prostate cells to PARP1 inhibitors. To test this, we generated two isogenic overexpression models of PCAT-1, and one stable knockdown model, in prostate cell lines. PCAT-1-overexpressing cell lines (Du145 and RWPE) showed a marked increase in sensitivity to two PARP inhibitors, whereas the stable knockdown cells (LNCaP) showed decreased sensitivity to PARP inhibitors. This was accompanied by a decrease in RAD51 foci formation, a hallmark of DNA repair via homologous recombination, in PCAT-1 overexpressing cells, and an increase in RAD51 foci formation in PCAT-1 knockdown cells. MCF7 breast cancer cells overexpressing PCAT-1 did not recapitulate these phenotypes, supporting our observation that PCAT-1 is a prostate-specific gene. These data define PCAT-1 as a novel prostate cancer-associated lncRNA functionally involved in tumor cell proliferation and sensitivity to PARP inhibitor therapy by suppressing homologous recombination. High PCAT-1 expression may therefore represent a potential biomarker for patient response to PARP targeted therapy. The findings presented herein establish the utility of RNA-Seq to comprehensively identify disease-associated lncRNAs and suggest that clinical translation of these findings may improve the stratification of cancer subtypes. Citation Format: John R. Prensner, Hari K. Iyer, Christopher A. Maher, Felix Feng, Arul M. Chinnaiyan, Matthew K. Iyer, Wei Chen, O. Alejandro Balbin, Saravana M. Dhanasekaran, Qi Cao, Xuhong Cao, Xiaojun Jing, Daniel Robinson. Transcriptome sequencing identifies PCAT-1, a novel lncRNA implicated in prostate cancer progression and therapeutic response to PARP inhibitors [abstract]. In: Proceedings of the AACR Special Conference on Advances in Prostate Cancer Research; 2012 Feb 6-9; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2012;72(4 Suppl):Abstract nr C36.