Abstract 2082: ERK activation mediated by Duox2 expression and reactive oxygen production is responsible for HIF-1α accumulation and VEGF induction in IFN-γ-treated BxPC-3 pancreatic cancer cells

Abstract

Abstract Reactive oxygen species (ROS) play an important role in tumor angiogenesis by up-regulating pro-angiogenic effectors such as HIF-1alpha and vascular endothelial growth factor (VEGF). However, the sources of ROS and the pro-angiogenic signaling pathways they target in tumor cells remain incompletely understood. We recently found that several members of the NADPH Oxidase (Nox) gene family are expressed in many human tumors; of these, Duox (Dual oxidase) 2 is highly expressed in tumors of the gastrointestinal tract. Recently, we demonstrated that IFN-gamma treatment of Bx-PC-3 cells upregulates the expression of Duox2 (and its maturation factor DuoxA2; but no other Nox isoform) leading to long-lived production of H2O2. Using this system, we investigated the role of Duox2-mediated ROS in the regulation of HIF-1alpha and VEGF expression. In BxPC-3 cells, IFN-gamma induction of Duox2/A2 expression was accompanied by both intra- and extracellular ROS accumulation, and by the simultaneous upregulation of HIF-1alpha and VEGF. However, in the MIA-PaCa-2 and PANC-1 lines that do not upregulate Duox2 following IFN-gamma exposure, no HIF-1alpha or VEGF induction was observed. In BxPC-3 cells, both the Nox inhibitor diphenylene iodonium and the ROS scavenger N-acetyl-L-cysteine suppressed IFN-gamma-induced HIF-1alpha accumulation and VEGF upregulation, suggesting that Duox2-derived ROS are involved in VEGF induction. Further experiments revealed that IFN-gamma induced a sustained activation of ERK1/2, which subsequently activated p90 ribosomal S6 kinases (RSKs) and the regulator of protein translation, ribosomal protein S6, resulting in enhanced synthesis of HIF-1alpha. ERK activation was required for IFN-gamma-induced HIF-1alpha and VEGF expression, since treatment with the MEK inhibitor U0126 completely abrogated IFN-gamma-induced HIF-1alpha and VEGF expression. Duox2-specific siRNA experiments showed that IFN-gamma-induced ROS, ERK activation, and HIF-1alpha accumulation were significantly attenuated following silencing of Duox2 expression. However, IFN-gamma-induced expression of VEGF was unchanged following Duox2 knockdown. Ongoing studies are evaluating the role of residual Duox2 or Duox2-independent pathways for VEGF activation following IFN-gamma exposure as well as other downstream, pro-angiogenic targets of Duox2-mediated HIF-1alpha expression under normoxic conditions. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2082. doi:10.1158/1538-7445.AM2011-2082