Metabolic Reprogramming by Mutant GNAS Creates an Actionable Dependency in Intraductal Papillary Mucinous Neoplasms of the Pancreas

Abstract

ABSTRACTObjectiveOncogenic “hotspot” mutations ofKRASandGNASare two major driver alterations in Intraductal Papillary Mucinous Neoplasms (IPMNs), which arebona fideprecursors to pancreatic ductal adenocarcinoma. We previously reported that pancreas-specificKrasG12DandGnasR201Cco-expression in p48Cre;KrasLSL-G12D; Rosa26LSL-rtTA; Tg (TetO-GnasR201C) mice (“Kras;Gnas” mice) caused development of cystic lesions recapitulating IPMNs. Here, we aim to unveil the consequences of mutantGnasR201Cexpression on phenotype, transcriptomic profile, and genomic dependencies.DesignWe performed multimodal transcriptional profiling (bulk RNA sequencing, single cell RNA sequencing, and spatial transcriptomics) in the “Kras;Gnas”autochthonous model and tumor-derived cell lines (Kras;Gnascells), whereGnasR201Cexpression is inducible. A genome-wide CRISPR/Cas9 screen was conducted to identify potential vulnerabilities inKrasG12D;GnasR201Cco-expressing cells.ResultsInduction ofGnasR201C– and resulting G(s)alpha signaling – leads to the emergence of a gene signature of gastric (pyloric type) metaplasia in pancreatic neoplastic epithelial cells. CRISPR screening identified the synthetic essentiality of glycolysis-related genesGpi1andSlc2a1inKrasG12D;GnasR201Cco-expressing cells. Real-time metabolic analyses inKras;Gnascells and autochthonousKras;Gnasmodel confirmed enhanced glycolysis uponGnasR201Cinduction. Induction ofGnasR201CmadeKrasG12Dexpressing cells more dependent on glycolysis for their survival. Protein kinase A-dependent phosphorylation of the glycolytic intermediate enzyme PFKFB3 was a driver of increased glycolysis uponGnasR201Cinduction.ConclusionMultiple orthogonal approaches demonstrate thatKrasG12DandGnasR201Cco-expression results in a gene signature of gastric pyloric metaplasia and glycolytic dependency during IPMN pathogenesis. The observed metabolic reprogramming may provide a potential target for therapeutics and interception of IPMNs.SUMMARYWhat is already known on this topicActivating “hotspot” mutations ofKRASandGNASare found in a majority of Intraductal Papillary Mucinous Neoplasms (IPMNs).Expression of mutantKRASandGNASdrives development of IPMN-like cystic lesions in the murine pancreas that eventually progress to pancreatic ductal adenocarcinoma (PDAC).What this study addsMutantGNASand the resulting aberrant G(s)alpha signaling drives a transcriptional signature of gastric (pyloric type) metaplasia in IPMNs with mucin production.Aberrant G(s)alpha signaling enhances glycolysis via protein kinase A-dependent phosphorylation of the glycolytic enzyme PFKFB3.Enhanced glycolysis inKRAS;GNAS-mutated IPMN cells is validated via multiple orthogonal approachesin vitroandin vivoand represents an actionable metabolic vulnerability.How this study might affect research, practice or policyThe present study provides mechanistic insight into how aberrant G(s)alpha signaling alters the biology ofKras-mutant pancreatic epithelial neoplasia through metaplastic and metabolic reprogramming.Targeting glycolysis in IPMNs may represent both a therapeutic avenue as well as an opportunity for intercepting progression to invasive cancer.