Endoplasmic Reticulum Stress-Mediated Signaling in Pancreatic Cancer
2018-04
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Endoplasmic Reticulum Stress-Mediated Signaling in Pancreatic Cancer
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2018-04
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Abstract
Pancreatic ductal adenocarcinoma (PDAC) ranks among the poorest prognoses for cancer patients, with an estimated 5-year survival of just 8%. The stagnant survival rates are a result of late detection, chemoresistance, and an aggressive tumor phenotype. Too few patients are eligible for surgery, which results in an urgent need for more effective chemotherapeutic treatment options. One promising pharmacological advancement is currently undergoing a Phase II clinical trial and has been studied by our laboratory. Triptolide is a Chinese herb, which has shown to be very effective in eliminating pancreatic cancer cells in vitro and in vivo. In conjunction with the Medicinal Chemistry Department at the University of Minnesota, a prodrug of triptolide, named Minnelide™, has been synthesized. Our laboratory has since studied triptolide and Minnelide™ extensively, in order to determine the mechanisms of action. The initial study in this dissertation precipitated based on an earlier finding in the Saluja laboratory that triptolide not only downregulates heat shock protein 70 (HSP70) and specificity protein 1 (SP1), but also causes chronic endoplasmic reticulum (ER) stress and cell death. Our study shows that downregulating SP1, a transcription factor that is overexpressed in pancreatic cancer, activates the unfolded protein response (UPR) and results in chronic ER stress. We further show that inhibition of SP1, as well as inducing ER stress, leads to lysosomal membrane permeabilization (LMP), a sustained accumulation of cytosolic calcium, and eventually cell death in pancreatic cancer. Even though ER stress can result in cell death, it is initially a homeostatic mechanism, which aims to protect cells. This led us to ask what role acute ER stress and UPR plays in pancreatic cancer. We show that modulating glucose regulatory protein 78 (GRP78), the master regulator of the UPR, can have a profound effect on multiple pathways that mediate chemoresistance. Our study showed for the first time that knockdown of GRP78 can diminish efflux activity of ATP-binding cassette (ABC) transporters, and it can decrease the antioxidant response resulting in an accumulation of reactive oxygen species (ROS). We also show that these effects can be mediated by the activity of SP1. Our investigation into acute ER stress led to further studies to characterize the UPR signaling in pancreatic cancer. We show that shGRP78 dysregulates multiple transcriptomic and proteomic pathways important in cancer (proliferation, survival, fatty acid metabolism). GRP78 downregulation decreases stemness and self-renewal properties in vitro. In vivo studies demonstrate that GRP78 knockdown results in delayed tumor initiation, and decreased tumor growth. Further, downregulation of GRP78 results in fatty acid metabolism dysregulation. The last study in this dissertation focuses on the tumor microenvironment and SP1 oncogenic signaling. We evaluated the transcriptomic profiling conducted after treatment with triptolide revealed deregulation of the transforming growth factor beta (TGF-β) signaling pathway in cancer-associate fibroblasts (CAFs), resulting in an apparent reversal of their activated state to a quiescent, non-proliferative state. The neighboring epithelial cells exhibited a decrease in oncogenic signaling as manifested by downregulation of SP1. Our findings suggest that approaches to inactivate CAFs and prevent tumor-stromal crosstalk may offer a viable strategy to treat pancreatic cancer. These studies underscore the importance of ER stress and understanding the complex balance of adaptation versus cell death in pancreatic cancer. We have identified SP1 and GRP78 as potential targets for future PDAC therapies. These findings have clinical relevance as both SP1 and GRP78 are overexpressed in pancreatic cancer patients and increased expression of these proteins are indicative of poor prognosis.
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University of Minnesota Ph.D. dissertation. 2018. Major: Pharmacology. Advisor: Ashok Saluja. 1 computer file (PDF); 142 pages.
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Dauer, Patricia. (2018). Endoplasmic Reticulum Stress-Mediated Signaling in Pancreatic Cancer. Retrieved from the University Digital Conservancy, https://hdl.handle.net/11299/198397.
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