Pancreatic ductal adenocarcinoma (PDA) is a devastating disease with a 5-year survival rate of roughly 7%. By 2020 it is predicted to be the second leading cause of cancer death in the United States. Two major contributors to these poor outcomes is the presence of metastasis and its high resistance to chemotherapy. Both metastasis and chemoresistance are associated with a process known as epithelial to mesenchymal transition (EMT) by which epithelial cells undergo a phenotypic switch to confer properties similar to mesenchymal cells. This endows tumor cells with the ability to invade, metastasize and resist chemotherapy. Current approaches to understanding cancer EMT have relied on in vitro studies and exogenous factors or known developmental pathways. However, our inability to isolate and analyze EMT tumor cells in vivo has limited our ability to identify and target pathways that drive this process in vivo. The goal of this proposal is to delineate the molecular mechanisms of EMT in PDA tumors and identify pathways that could ultimately be targeted to improve clinical outcomes. To study EMT in the context of stochastically-arising pancreatic tumors, our laboratory developed a mouse model of PDA (the ?KPCY? model) that employs a YFP lineage marker to label and track tumor cells in vivo. Using this model, we uncovered a novel ?non-classical EMT? program operating in a majority of tumors by which cancer cells use protein re-localization (as opposed to classically-defined programs of transcriptional repression) to shed their epithelial phenotype.
Aim 1 will focus on determining the mechanistic details of non-classical EMT while aim 2 represents a translational arm of our basic mechanistic study to unravel vulnerabilities of non-classical EMT and the potential use of calcium channel blockers to modulate sensitivity. In preliminary data, I found that calcium-calmodulin signaling is a driver of non-classical EMT. Treatment of PDA cells with a calcium ionophore, ionomycin, results in a loss of epithelial marker E-cadherin from the membrane and importantly occurs without mRNA repression. In addition, this process is dependent on calcium binding protein calmodulin.
Sub aims 1 a and 1b will delineate the role and impact of calmodulin on calcium-induced EMT, migration, invasion and metastasis. In addition, after treatment of PDA cells with ionomycin, I observed a 20kDa shift in E-cadherin protein suggesting the addition of a post-translational modification (PTMs) regulating protein re-localization.
Sub aim 1 c will identify calcium-induced PTMs regulating E-cadherin trafficking. Our lab has previously found that epithelial and mesenchymal tumor cells exhibit different sensitivities in vivo. Thus, the identification of a novel subtype of EMT (non-classical) allows for an opportunity to exploit vulnerabilities unique to this subtype (subaim 2a). In addition, given the role of EMT in chemoresistance, targeting EMT may improve the efficacy of anti-cancer drugs by altering the epithelial-mesenchymal plasticity of cancer cells. Therefore, the final aim of this proposal will determine the efficacy of calcium channel blockers on reversing EMT and sensitizing tumors to chemotherapeutic agents (subaim 2b).

Public Health Relevance

Epithelial-to-Mesenchymal Transition (EMT) is a key process during embryogenesis that is hijacked by tumors to promote invasion, metastasis, and chemoresistance. By using a novel and physiological mouse model of pancreatic cancer to study EMT in vivo, we found that most tumors lose their epithelial phenotype through a non- classical program involving protein re-localization instead of transcriptional repression. This proposal seeks to delineate the molecular mechanisms of non-classical EMT by calcium signaling and to explore translational therapeutics that may impact upon the clinical barriers of pancreatic cancer.

National Institute of Health (NIH)
National Cancer Institute (NCI)
Predoctoral Individual National Research Service Award (F31)
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Special Emphasis Panel (ZRG1)
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Schmidt, Michael K
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University of Pennsylvania
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