Pancreatic ductal adenocarcinoma (PDA) is the third leading cause of cancer related deaths in the U.S., while colorectal cancer (CRC) is the third most common cancer. The RNA binding protein HuR (ELAVL1), supports a pro-oncogenic network in gastrointestinal (GI) cancer cells through enhanced HuR expression. Using a publically available database, HuR expression levels were determined to be increased in primary PDA and CRC tumor cohorts as compared to normal pancreas and colon tissues, respectively. CRISPR/Cas9 technology was successfully used to delete the HuR gene in both PDA (MIA PaCa-2 and Hs 766T) and CRC (HCT116) cell lines. HuR deficiency has a mild phenotype, in vitro, as HuR-deficient MIA PaCa-2 (MIA.HuR-KO(−/−)) cells had increased apoptosis when compared to isogenic wild-type (MIA.HuR-WT(+/+)) cells. Using this isogenic system, mRNAs were identified that specifically bound to HuR and were required for transforming a 2D culture into 3D (i.e., organoids). Importantly, HuR-deficient MIA PaCa-2 and Hs 766T cells were unable to engraft tumors in vivo compared to control HuR-proficient cells, demonstrating a unique xenograft lethal phenotype. While not as a dramatic phenotype, CRISPR knockout HuR HCT116 colon cancer cells (HCT.HuR-KO(−/−)) showed significantly reduced in vivo tumor growth compared to controls (HCT.HuR-WT(+/+)). Finally, HuR deletion affects KRAS activity and controls a subset of pro-oncogenic genes. Implications The work reported here supports the notion that targeting HuR is a promising therapeutic strategy to treat GI malignancies.

Ras-dependent signaling is an important regulator of cell cycle progression, proliferation, senescence, and apoptosis. Several of the downstream effectors of Ras play dual roles in each of these processes. Under one set of conditions, they promote cell cycle progression and proliferation; yet, in a different paradigm, they drive cell cycle arrest and apoptosis. Furthermore, there is cross talk between certain downstream effectors of Ras including the PI3K-AKT and Raf-MEK-ERK pathways. Here we describe a series of experiments used to dissect the effect of different Ras-dependent signaling pathways on cell cycle progression, proliferation, senescence, and apoptosis. Furthermore, we highlight the importance of consistent growth conditions of cells in culture when studying Ras-dependent signaling as we show that the activation of downstream effectors of Ras changes with the confluency at which the cells are grown.

Increased expression of the chemokine CXCL1 and its sole receptor, CXCR1 have been correlated with poor pancreatic cancer patient survival and time to recurrence, as well as with pancreatic perineural invasion. We have previously shown that metastasis of prostate and breast cancer is in part driven by CXCL1, and have developed small molecule inhibitors against the CXCR1 receptor that diminish metastatic burden. Here we ask if inhibition of this chemokine receptor affects the phenotype of PDAC tumor cells. Our findings demonstrate that motility, invasion, and contact-independent growth of PDAC cells all increase following CXCL1 exposure, and that antagonism of CXCR1 by the inhibitor JMS-17-2 reduces each of these phenotypes and correlates with a downregulation of AKT phosphorylation. These data suggest that PDAC tumor cell migration and growth, elements critical in metastatic progression, may susceptible to pharmacologic intervention.

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