Posttranslational regulation of macropinocytosis in pancreatic cancer - The majority of pancreatic ductal adenocarcinoma (PDAC) patients present with late-stage, metastatic disease that is often resistant to therapeutics, resulting in the lowest survival rate of all major cancers. Metabolic rewiring in response to oncogenic signals plays a critical role in PDAC cell survival, tumor growth, and metastasis. In contrast to normal epithelial cells, these metabolic alterations make PDAC tumors dependent on glutamine for survival, highlighting a unique metabolic vulnerability that can be therapeutically exploited. However, during times of nutrient stress, PDAC cells can circumvent this vulnerability by engulfing extracellular fluids to replenish amino acids, including glutamine, in a process called, macropinocytosis. Macropinocytosis occurs downstream of oncogenic KRAS, a small GTPase that is almost universally mutated in PDAC patients. The inhibition of macropinocytosis in vivo reduces PDAC tumor growth, underscoring the importance of this pathway to cancer cell survival. However, there remain gaps in our knowledge regarding the signaling mechanisms that regulate macropinocytosis and the contribution of macropinocytic signaling to aggressive cancer phenotypes. Protein phosphatase 2A (PP2A) is a heterotrimeric complex that regulates a wide variety of cell signaling pathways, including KRAS, and is commonly deregulated in human PDAC tumors. Recently, PP2A has been implicated as an important regulator of macropinocytosis, but the mechanism by which this occurs is unknown. We have shown that the genetic loss of the specific PP2A subunit, B56a, accelerates the formation of KRAS-driven pancreatic lesions in an in vivo mouse model, implicating this subunit as a critical negative regulator of KRAS phenotypes during PDAC progression. Here, we demonstrate that PP2A-B56a activation results in attenuated glutamine signaling, a significant accumulation of macropinocytic vesicles, and cell death. Furthermore, the therapeutic combination of PP2A activators and metabolic inhibitors leads to synergistic loss of PDAC tumor growth in vivo. Based on these findings, we hypothesize that activation of PP2A-B56a sensitizes PDAC tumors to glutamine antagonists by limiting metabolic plasticity through the blockade of macropinocytosis-dependent nutrient acquisition. We will test this hypothesis in the following Specific Aims: 1) Identify the posttranslational mechanisms by which PP2A-B56a regulates macropinosome-lysosome fusion, 2) Evaluate the impact of PP2A activation on PDAC progression and macropinocytosis in vivo, and 3) Determine if PP2A activation increases the efficacy of metabolic inhibitors, in vitro and in vivo. Successful completion of these studies will significantly increase our knowledge of the posttranslational mechanisms that govern PDAC nutrient scavenging, providing novel targets for drug discovery in a highly lethal cancer.
