Macropinocytosis Inhibition as a Glutamine Mimetic Sensitization Strategy in Pancreatic Cancer

PROJECT SUMMARY Pancreatic ductal adenocarcinoma (PDAC) cells depend on glutamine to meet their unique metabolic needs. Glutamine fuels a wide array of biosynthetic pathways; however, despite these established dependencies, the targeting of specific enzymes involved in glutamine utilization have yet to yield beneficial results. This proposal is based on our preliminary data demonstrating that broadly targeting glutamine utilization via the substrate analog 6-diazo-5-oxo-L-norleucine (DON) has a profound effect on PDAC tumor growth and metastasis. We have discovered that the effects of DON on PDAC cell fitness are directly linked to asparagine (Asn) supply. This observation is exciting because the ability of DON to exert its deleterious effects on tumors through blocking Asn biosynthesis has never been examined. Metabolic targeting can trigger adaptive responses that allow tumor cells to escape the effects of nutrient stress. We have found that PDAC cells robustly upregulate macropinocytosis as a metabolic adaptation to DON treatment. In this way, the protein-derived amino acids produced by this uptake pathway have the ability to dampen the effects of DON, suggesting that macropinocytosis inhibition might be a beneficial strategy to sensitize PDAC tumors to DON. To gain mechanistic insight into DON-induced uptake, we performed a high-throughput siRNA human kinome screen, identifying WNK lysine deficient protein kinase 4 (Wnk4) as a potent regulator of DON-stimulated macropinocytosis. Wnk4 plays an important role in ion transport in the kidney, but has not been linked to macropinocytosis and its role in cancer has not been elucidated. In this proposal, we will: (1) examine the role of macropinocytosis as a metabolic adaptation to DON in PDAC, and (2) decipher the function of Wnk4 in macropinocytosis and PDAC tumor growth. This proposed project is significant and innovative because it will establish the metabolic and molecular mechanisms underlying the functionality of glutamine mimetics in PDAC and decipher the therapeutic framework for new combination strategies for PDAC.