Pancreatic ductal adenocarcinoma (PDAC) has a 5-year survival rate under 5% with 40,000 patients succumbing to it each year, and no improvement in first-line treatment has occurred since 1997. The most common mutation in PDAC is a constitutively-activating change in the small G protein KRAS. This mutation type, when targeted to the pancreas as a single residue change from glycine to aspartic acid at amino acid twelve using the pancreas-specific transcription factor Pdx1 and a LOX-Stop-LOX (LSL) cassette upstream of the KRAS gene (Pdx1-cre~ KRASG12D+/fl), accurately recapitulates the progression of PDAC from precursor lesions to invasive cancer. This mutation by itself, however, has a low level of penetrance, arguing for the idea that further lesions are needed to drive PDAC. One possible candidate for this role is p120catenin (p120ctn), an intracellular protein that is a component of the adherens junction. The adherens junction is a macromolecular complex required for normal organization of epithelial tissues, and one of its components, E-cadherin, is a potent tumor suppressor. p120ctn is a known binding partner to E-cadherin, and loss of p120ctn is known to lead to E-cadherin degradation. p120ctn has been implicated in several aspects of cancer, including inflammation, motility, and proliferation. Conditional knockout models in colon, esophagus, skin, and salivary gland show that p120ctn loss is able to lead to phenotypes ranging from hyperplasia to frank invasive cancer. Although p120ctn is known to be lost or mislocalized in almost all human carcinomas, including PDAC, it has not been shown whether p120ctn perturbation is incidental or central to tumor progression. Additionally, pancreatic development is in large part mediated by the small G protein Cdc42, a downstream target of p120ctn modulation. p120ctn likely therefore plays important roles in both development and tumorigenesis of the pancreas, and evidence suggests these roles may be isoform-specific. We therefore propose in this study to investigate p120ctn's role in pancreatic morphogenesis and tumorigenesis. Our overarching hypothesis is that p120ctn is critical in pancreatic development and its loss fosters PDAC progression. This hypothesis will be pursued through the following interrelated specific aims.
In Aim 1, we will use a combination of 3D organotypic culture and co-culture as well as senescence, invasion, migration, and proliferation assays to determine what effect p120ctn loss, as well as isolated p120ctn isoform overexpression, has on the morphogenesis of pancreatic ductal epithelium. In addition to these methods, we will utilize a full repertoire of molecular biology techniques including PCR/qPCR, Western blots, protein activity assays, viral vectors as well as confocal microscopy and immunofluoresence.
In Aim 2, we will utilize the Pdx1-cre~ LSL-KRASG12D+/fl and the Pdx1-cre~ p120ctnfl/fl mice to study the role of p120ctn both alone and in the context of a constitutively activating KRASG12D mutation. We also will utilize orthotopic transplantation, flow cytometry, histology, and biochemical techniques to characterize the role of p120ctn in these mice both on a gross, phenotypic level as well as cellular and biochemical levels. Our studies will provide new mechanistic insights into p120ctn biology and p120ctn's functional role in PDAC. Our complementary in vitro and in vivo approaches will provide a platform for potential translational applications.
Pancreatic ductal adenocarcinoma (PDAC) is the cause of nearly 40;000 deaths per year. No significant change in standard adjuvant therapy has taken place since 1997; suggesting a need for elucidating signaling pathways that might serve as putative drug targets. By investigating the role of p120catenin (p120ctn) in PDAC; we will be able to understand how this protein drives cancer progression.
|Takano, Shigetsugu; Reichert, Maximilian; Bakir, Basil et al. (2016) Prrx1 isoform switching regulates pancreatic cancer invasion and metastatic colonization. Genes Dev 30:233-47|