Abstract 3432: A genome-wide shRNA screen for suppressors of prostate cancer cell invasion

Abstract

Metastasis in prostate cancer is caused by genetic reprogramming leading to increased cell motility and the formation of invasive structures which allow cells to invade out of the primary tumour in a process known as intravasation. In order to search for novel genes inhibiting invasive phenotypes responsible for intravasation, a genome-wide high throughput short hairpin RNA screen was used to identify genes which suppress aggressive phenotypes in BPH (Benign Prostatic Hyperplasia) prostate epithelial cells when grown in 3D matrigel culture conditions. Using the The RNAi Consortium (TRC) lentiviral shRNA library that covers the entire human transcriptome (80,000 unique constructs), BPH cells were infected such that each cell would integrate a single shRNA construct that will target mRNA expression of a known human gene. Cells were plated at 4500 cells per cm2 until a 2.5X genome-wide coverage was achieved. In 3D matrigel culture conditions, BPH colonies exhibit normal epithelial (round) colony morphology. Hence, we screened for clonogenic BPH colonies that appeared fibroblastic and spindle-shaped. Collection of these “hits” yield gene identities that suppress intravasation. In this ongoing study, we have achieved 2.5x coverage of the TRC shRNA library in BPH cells. After 11 days of growth, 7 “hits” from the BPH screen that exhibit an invasive phenotype were isolated, and were validated again in 3D matrigel culture. These “hits” have revealed 12 potential invasion suppressors involved in pathways including ER stress, cortactin phosphorylation signalling, cell differentiation and genome stability. These hits are currently being re-validated with other shRNAs specific for each gene. Overall, we provide a high-throughput and high-content screen for discovering novel genes that inhibit prostate cancer progression. These functional genomics screens which focus on altered prostate cell colony phenotypes may open new doors for potential drug treatments and a greater understanding of prostate cancer and BPH. Genome-wide shRNA screen invasion suppressor genesHit #GeneSymbolCellular Function1Interleukin 19IL19Upregulates IL6 and TNFA1Polyhomeotic-like protein 3PHC3Part of polycomb group complex (repressor)2Zinc finger protein 397ZNF397Repressor/activator transcription factor2Transmembrane protein 189-ubiquitin-cojugating enzyme E2 variant 1TMEM189-UBE2V1Cytosolic ubiquitination protein3Nuclear protein localization 4 homologNPLOC4Involved in ER stress (retro-translocation of proteins for degradation)4Flavin adenine dinucleotide synthetase 1FADSConverts FMN to FADH25Selenoprotein SSELSInvolved in ER stress (retro-translocation of proteins for degradation)5Serine/threonine-protein kinase 1OSR1Phosphorylates PAK1 (interacts with cortactin)6Ecotropic viral integration site protein 2BEVI2BInvolved in melanocyte and keratinocyte differentiation6Coiled-coil domain containing 111CCDC111Involved in DNA integrity and genome stability7Serine hydrolase-like 1SERHLPeroxisomal protein7Growth factor-induced 1GFI1Transcriptional repressor; involved in hematopoesis and oncogenesis Citation Format: Sean J. Leith, Susan E. Kuruvilla, Jason Moffat, Ann F. Chambers, Eva A. Turley, Joseph L. Chin, Hon S. Leong. A genome-wide shRNA screen for suppressors of prostate cancer cell invasion. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3432. doi:10.1158/1538-7445.AM2014-3432