RET-receptor mediated invasion in pancreatic ductal adenocarcinoma
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Authors
Thomson, Bryanna
Date
Type
thesis
Language
eng
Keyword
RET , Pancreatic ductal adenocarcinoma , Invasion , dorsal root ganglion
Alternative Title
Abstract
Pancreatic ductal adenocarcinoma (PDAC) is frequently associated with invasion along nerves, which generates severe pain. KRAS and TP53 are the most common driving mutations for PDAC, however, these tumours also overexpress the RET receptor which regulates cell proliferation and migration. Since nerves are known to secrete RET ligands, PDAC invasion towards nerves is potentially enhanced through RET signalling. We evaluated the dependency of PDAC invasion on RET using a collagen invasion assay and PDAC spheroids. We found that RET-selective and multikinase inhibitors, such as Selpercatinib and Vandetinib respectively, decreased or prevented invasion. We further determined RET pathways that are important to this process by treating with inhibitors or using shRNA knockdown cell lines for various proteins, required in the downstream signalling of RET, in our invasion assay. We found that proteins associated with RET migration, proliferation and recycling pathways, such as PI3Kinase, MEK, mTOR, GRB2 and ARF6, decrease invasion. However, proteins associated with RET-mediated proliferation and recycling, such as STAT3 and GGA3, showed no significant effects. Our data suggest that RET is required for PDAC invasion and therefore inhibiting RET, or certain downstream signals could aid in reducing PDAC invasion along the nerves.
Preliminary experiments were also performed to help build an experimental plan for determining the relevance of these proteins for chemotactic invasion of nerves. MiaPaCa2-EGFP cells and primary mouse neurons were cultured together to observe any interactions between the two cell types. Neurons were observed with axons growing in the direction of groups of MiaPaCa2 cells and two potential phenotypes of neurons were observed (axonal growth vs. no axonal growth). Neuron body area and axonal length measurements were taken to characterize morphological differences between the two. A correlation between axon length and neuronal body area was observed with smaller neurons having longer axons, and a significant difference between areas of neurons with axonal growth and those without. This indicates that there may be multiple subtypes of neurons present and that not all of them may react to the presence of PDAC cells, however further research is required.