Abstract Chronic inflammation is an important risk factor for the development of pancreatic ductal adenocarcinoma (PDAC). Dual oxidase 2 (DUOX2), one of the seven NADPH oxidase (NOX) family members, by generating extracellular H2O2, is intimately involved in both host defense and chronic inflammation in the gastrointestinal tract. Previously, we demonstrated that short (24 h) exposures of several human pancreatic cancer cell lines to pro-inflammatory cytokines (IFN-γ, IL-4/IL-13, IL-17A) can effectively enhance expression of DUOX2 and its maturation factor DUOXA2. To better mimic chronic inflammation in vivo, a panel of human PDAC cell lines was exposed to the TH2 cytokine IL-4 (50ng/ml) for longer times (7 days); and the effect of cytokine treatment on Stat6 activation and DUOX2 mRNA and protein expression was explored. In BxPC-3 and AsPC-1 cell lines, prolonged IL-4 stimulation results in a sustained activation of Stat6, dramatic induction of DUOX2 mRNA and protein expression, a 3-5-fold increase in extracellular H2O2 production compared to solvent-treated cells, and–most importantly–a robust DNA double strand break (DSB) response indicated by elevated levels of γH2AX. In marked contrast, for two JAK-Stat6 signaling defective PDAC lines, PANC-1 and CFPAC-1, no enhancement of DUOX2 expression with concomitant DSBs was observed under the same conditions. Moreover, when the anti-inflammatory drug dexamethasone (Dex) was co-administered with IL-4 (for 48h), cytokine-related DUOX2 induction and DNA damage in BxPC-3 and AsPC-1 cells was significantly diminished. Furthermore, in the KPC GEMM of PDAC, RT-PCR and immunohistochemistry revealed that DUOX2 mRNA and protein expression in pancreatic tumor tissues were significantly increased compared to adjacent normal tissues and were associated with areas of chronic inflammation. Finally, xenografts established with BxPC-3 cell clones in which DUOX2 mRNA was silenced by stable expression of DUOX2-specific shRNAs demonstrated severely delayed growth in vivo compared to clones stably expressing scrambled shRNAs. These studies, coupled with our previous data demonstrating elevated DUOX protein expression in pancreatitis, pancreatic intra-epithelial neoplasia (PanIN), and frank pancreatic cancers in humans, suggest that ROS- derived from cytokine-mediated DUOX2 up-regulation may initiate and promote pancreatic cancer progression through ROS-induced DNA damage. Citation Format: Yongzhong Wu, Jiamo Lu, Smitha Antony, Jennifer L. Meitzler, Agnes Juhasz, Guojian Jiang, Iris Dahan, James H. Doroshow. DUOX2-related H2O2 production contributes to pro-inflammatory cytokine-related DNA damage and tumor cell growth in models of pancreatic ductal adenocarcinoma [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 514.