Evaluating the therapeutic potential of the PAK1 and TBK1 kinases in pancreatic ductal adenocarcinoma Public Deposited

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  • March 20, 2019
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  • Baker, Nicole
    • Affiliation: School of Medicine, Department of Pharmacology
Abstract
  • Pancreatic ductal adenocarcinoma (PDAC) is an extremely lethal cancer characterized by a high frequency (>95%) of activating mutations in the KRAS oncogene, which is a well-validated driver of PDAC growth. However, to date, no successful anti-KRAS therapies have been developed. Inhibitors targeting components of KRAS downstream signaling pathways, when used as monotherapy or in combination, have been ineffective for long-term treatment of KRAS-mutant cancers. Decidedly, the most studied and most targeted KRAS effector pathways have been the RAF-MEK-ERK mitogen-activated protein kinase (MAPK) cascade and the PI3K-AKT-mTOR lipid kinase pathway. The apparent lack of success exhibited by these inhibitors is due, in part, to an underestimation of the importance of other effectors in KRAS-dependent cancer growth. Additionally, compensatory mechanisms reprogram these signaling networks to overcome the action of inhibitors of the ERK MAPK and PI3K pathways. Consequently, the central hypothesis of my dissertation research is that a better understanding of the role of less-studied KRAS effector signaling pathways may lead to more effective therapeutic strategies to block KRAS effector signaling and PDAC growth. Although the TIAM1-RAC1 small GTPase effector pathway has been validated as a driver of KRAS-mutant cancer growth, how RAC1 mediates this role has not been established. My studies address a possible critical role for the p21-activated kinase 1 (PAK1) in this effector pathway. In support of this, I found that PAK1 protein levels are overexpressed both in a subset of pancreatic cancer cell lines and in primary patient tumor samples. Moreover, I determined that stable shRNA-mediated suppression of PAK1 protein expression inhibited the anchorage-dependent and -independent growth of PDAC cell lines in vitro. I also observed that a pharmacologic inhibitor of PAK1 recapitulated the reduced growth phenotypes observed upon genetic ablation of PAK1. As KRAS-mutant tumors are known to upregulate certain cellular processes in order to support the increased metabolic demands of uncontrolled cellular proliferation, I sought to determine whether PAK1 signaling was partially accountable for ensuring that these metabolic needs were met. My studies confirmed a role for PAK1 in regulating macropinocytosis, a mechanism by which PDAC cells acquire macromolecules (e.g., proteins, polysaccharides, and lipids) from the extracellular environment as a source of nutrients. I found that both pharmacologic inhibition and genetic ablation of PAK1 resulted in markedly decreased macropinocytosis in PDAC cells. These data suggest inhibition of PAK1 in KRAS-mutant PDAC could interfere with PDAC metabolism and reduce tumor cell growth. I observed a further reduction in macropinocytosis upon inhibition of PAK1 together with concurrent ERK1/2 or PI3K inhibition. In summary, my results support PAK1 as a promising therapeutic target for pancreatic cancer. My lab and others have provided strong evidence for the key role of a second less-studied KRAS effector pathway, the RALGEF-RAL small GTPase effector pathway, in the growth of pancreatic and other cancers. One critical effector of RAL is the SEC5 component of the exocyst complex. How SEC5 contributes to the role of RAL in cancer remains unresolved. White and colleagues initially identified a SEC5 function independent of exocyst regulation that involved the TANK-binding kinase 1 (TBK1). When a study that searched for synthetic lethal partners of mutant KRAS identified TBK1, these findings suggested that TBK1 may be a critical mediator of RalGEF-RAL effector-driven cancer growth. However, a subsequent study questioned the role of TBK1 in the growth of KRAS-mutant cancers. When my lab obtained a novel pharmacologic inhibitor of TBK1, I embarked on studies to determine whether inhibition of TBK1 kinase activity could be an efficacious treatment strategy for PDAC. My studies revealed that inhibition of TBK1 alone showed limited growth inhibition in PDAC cell lines. Furthermore, I found that concurrent inhibition of TBK1 did not enhance the growth inhibitory activity of an ERK inhibitor. However, loss of TBK1 protein via shRNA or pharmacologic inhibition prompted the development of large, intracellular vesicles that appeared to be swollen autolysosomes and the product of non-productive autophagy. This work suggests that TBK1 may play a role in PDAC autophagy and provides a rationale for pairing TBK1 inhibition with other targeted therapies or chemotherapies to drive these tumor cells towards death. In summary, my studies support my hypothesis that concurrent inhibition of multiple KRAS effector pathways may provide more effective therapeutic strategies for PDAC. They emphasize that single agent therapies targeting KRAS will not be effective, a reality that is emerging from ongoing clinical trials. While my studies took a rational approach to identifying these combinations, unbiased chemical library screens with PAK1 and TBK1 inhibitors will also likely identify additional combinations of effector inhibitors for PDAC.
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  • In Copyright
Advisor
  • Cox, Adrienne
  • Johnson, Gary
  • Major, Michael
  • Der, Channing
  • Graves, Lee
Degree
  • Doctor of Philosophy
Degree granting institution
  • University of North Carolina at Chapel Hill Graduate School
Graduation year
  • 2016
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