Alternative polyadenylation (APA) is a gene regulatory process that dictates mRNA 3 ′ -UTR length, resulting in changes in mRNA stability and localization. APA is frequently disrupted in cancer and promotes tumorigenesis through altered expression of oncogenes and tumor suppressors. Pan-cancer analyses have revealed common APA events across the tumor landscape; however, little is known about tumor type-specific alterations that may uncover novel events and vulnerabilities. Here, we integrate RNA-sequencing data from the Genotype-Tissue Expression (GTEx) project and The Cancer Genome Atlas (TCGA) to comprehensively analyze APA events in 148 pancreatic ductal adenocarcinomas (PDACs). We report widespread, recurrent, and functionally relevant 3 ′ -UTR alterations associated with gene expression changes of known and newly identified PDAC growth-promoting genes and experimentally validate the effects of these APA events on protein expression. We find enrichment for APA events in genes associated with known PDAC pathways, loss of tumor-suppressive miRNA binding sites, and increased heterogeneity in 3 ′ -UTR forms of metabolic genes. Survival analyses reveal a subset of 3 ′ -UTR alterations that independently characterize a poor prognostic cohort among PDAC patients. Finally, we identify and validate the casein kinase CSNK1A1 (also known as CK1alpha or CK1a) as an APA-regulated therapeutic target in PDAC. Knockdown or pharmacological inhibition of CSNK1A1 attenuates PDAC cell proliferation and clonogenic growth. Our single-cancer analysis reveals APA as an underappreciated driver of protumorigenic gene expression in PDAC via the loss of miRNA regulation.

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Alternative polyadenylation (APA) is a gene regulatory process that dictates mRNA 3’-UTR length, resulting in changes in mRNA stability and localization. APA is frequently disrupted in cancer and promotes tumorigenesis through altered expression of oncogenes and tumor suppressors. Pan-cancer analyses have revealed common APA events across the tumor landscape; however, little is known about tumor type-specific alterations that may uncover novel events and vulnerabilities. Here we integrate RNA-sequencing data from the Genotype-Tissue Expression (GTEx) project and The Cancer Genome Atlas (TCGA) to comprehensively analyze APA events in 148 pancreatic ductal adenocarcinomas (PDAs). We report widespread, recurrent and functionally relevant 3’-UTR alterations associated with gene expression changes of known and newly identified PDA growth-promoting genes and experimentally validate the effects of these APA events on expression. We find enrichment for APA events in genes associated with known PDA pathways, loss of tumor-suppressive miRNA binding sites, and increased heterogeneity in 3’-UTR forms of metabolic genes. Survival analyses reveal a subset of 3’-UTR alterations that independently characterize a poor prognostic cohort among PDA patients. Finally, we identify and validate the casein kinase CK1α as an APA-regulated therapeutic target in PDA. Knockdown or pharmacological inhibition of CK1α attenuates PDA cell proliferation and clonogenic growth. Our single-cancer analysis reveals APA as an underappreciated driver of pro-tumorigenic gene expression in PDA via the loss of miRNA regulation.

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Background and Hypothesis:   Pancreatic ductal adenocarcinoma (PDAC) has a poor prognosis and the standard of care regimen, nab-paclitaxel (NPT) plus gemcitabine (Gem), leads to a dismal 8.5 months median survival. Targeted inhibition of Bromodomain and Extra-Terminal (BET) protein is currently under investigation for several cancers. We hypothesize that BET protein pathway inhibition by iBet-762 will enhance cytotoxic chemotherapy response in PDAC.  Experimental Design:  In vitro cell proliferation assays were performed using WST-1 reagent. Protein expressions were determined by Western Blot analysis. In vivo animal survival and tumor growth experiments were performed in NOD-SCID mice.   Results:  Inhibition in cell proliferation in human PDAC cells at 1 µM concentration in NPT+Gem, iBET-762, and NPT+Gem+iBet762 was 64%, 27%, 76% in AsPC-1; 43%, 13%, 69% in Panc-1; and 42%, 51%, 75% in MIA PaCa cells. iBET-762 decreased oncogenic proteins c-Myc, [Symbol]-catenin, Vimentin, and P-AKT while apoptosis related proteins such as cleaved PARP-1 and cleaved caspase-3 and cell cycle inhibitors proteins P21 & P27 were increased. In a peritoneal dissemination model, median animal survival compared to control (21 days) was increased after therapy with NPT+Gem (33 days, a 57% increase), iBet-762 (30 days, a 43% increase) and NPT+Gem+iBET-762 (44 days, a 110% increase). Effect of iBET-762 in combination with chemotherapy on local tumor growth is currently underway.    Conclusion and Potential Impact:   These findings suggest that the effects of standard chemotherapy can be enhanced through specific inhibition of BET proteins activity, and supports the clinical application of iBET-762 in combination with standard chemotherapy in PDAC patients.