Abstract 4264: Lysosome inhibition sensitizes pancreatic cancer to replication stress by aspartate depletion

Objective: Functional lysosomes are required for autophagy and macropinocytosis, the intra- and extracellular scavenging pathways cancer cells engage for nutrient acquisition. Pancreatic ductal adenocarcinoma (PDAC) tumors exhibit high basal lysosomal activity, and genetic or pharmacologic inhibition of lysosome function suppresses PDAC cell proliferation and tumor growth. However, the codependencies induced by lysosomal inhibition in PDAC have not been systematically explored. We hypothesized that identification and targeting the lysosomal inhibition-induced codependencies in PDAC cells would be an effective therapeutic strategy. Methods: A comprehensive pharmacological inhibition screen of the protein kinome was performed to identified lysosomal inhibition-induced codependency. LC-MS/MS-MRM and LC-MS methods were used to examine nucleotide biosynthesis and amino acid levels, respectively, to understand the mechanism of the identified codependency. A broad panel of PDAC cell lines, primary PDAC culture models, two- and three-dimentional culture models, a PDAC cell/stroma spheroid coculture model, and xenograft and syngeneic PDAC animal models were used to evaluate the treatment efficacy. Results: We found that replication stress response (RSR) inhibitors were synthetically lethal with chloroquine (CQ) and other lysosomal inhibitors in PDAC cells. CQ treatment reduced de novo nucleotide biosynthesis and induced replication stress. We found that CQ treatment caused mitochondrial dysfunction and depletion of aspartate, an essential precursor for de novo nucleotide synthesis, as an underlying mechanism. Supplementation with aspartate in PDAC cell culture and overexpression of the aspartate transporter SLC1A3 in xenograft PDAC tumors partially rescued the phenotypes induced by CQ. The synergy of CQ and the RSR inhibitor VE-822 was comprehensively validated in a broad panel of PDAC cell lines and primary PDAC cultures, in two- and three-dimentional PDAC cultures, in heterotypic spheroid culture with cancer-associated fibroblasts, and in in vivo xenograft and syngeneic PDAC mouse models. Conclusion: We discovered a codependency on functional lysosomes and an intact RSR pathway in PDAC, and developed the combination of CQ and RSR inhibitors as a translational therapeutic approach for PDAC. Citation Format: Shili Xu, Irmina A. Elliott, Amanda M. Dann, Stephanie S. Kim, Evan R. Abe, Woosuk Kim, Soumya Poddar, Alexandra Moore, Lei Zhou, Jennifer L. Williams, Joseph R. Capri, Razmik Ghukasyan, Cynthia Matsumura, D. Andrew Tucker, Wesley R. Armstrong, Anthony E. Cabebe, Nanping Wu, Luyi Li, Thuc M. Le, Caius G. Radu, Timothy R. Donahue. Lysosome inhibition sensitizes pancreatic cancer to replication stress by aspartate depletion [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4264.