NADK regulation by oncogenic KRAS signaling is essential for PDAC Tanya Schild1,2,3, Melanie McReynolds4, Christie Shea1,2, Vivien Low1,2,3, John Asara5, Noah Dephoure3, Joshua D. Rabinowitz4, John Blenis1,2,6 and Ana P. Gomes1,2,6 1 Meyer Cancer Center, Weill Cornell Medicine, New York, NY, USA. 2 Department of Pharmacology, Weill Cornell Medicine, New York, NY, USA. 3 Department of Biochemistry, Weill Cornell Medicine, New York, NY, USA. 4 Department of Chemistry, Princeton University, Princeton, NJ, USA. 5 Department of Medicine, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, MA, USA. 6 Corresponding authors Diagnosis of pancreatic ductal adenocarcinoma (PDAC) carries a dismal prognostic outcome. In recent years, metabolic adaptations have been shown to be critical for PDAC maintenance and for its acquired resistance to therapies. Many of these adaptations rely heavily on NADPH availability—however our understanding of NADPH metabolism and how is it regulated to support PDAC is still in its infancy. NADPH is recycled through reduction of NADP+ by several enzymatic systems in cells. In conditions where NADPH availability is critical, cells have an evolutionarily conserved mode of increasing the global pools of NADPH through de novo synthesis of NADP+ catalyzed by the NAD+ kinase (NADK). Here we establish that NAD+ kinase (NADK) is a major contributor to NADPH pools in PDAC. NADK, which catalyzes the de novo synthesis of NADP+ from NAD+, becomes hyperactive when oncogenic KRAS is present and constitutes a vital mechanism for PDAC survival. We traced this hyperactivation to a KRAS-induced and PKC-mediated phosphorylation of NADK, which results in an increase in the de novo NADP+ synthesis, and consequently to a rise in NADPH levels. For the first time, we show that NADK can be regulated by oncogenic signaling. This regulatory mechanism is essential for PDAC survival and highlights NADK as a much-needed new therapeutic target for PDAC.