Abstract The effects of dietary factors on pancreatic cancer have been proposed to augment etiology of disease, yet the mechanisms involved are poorly understood. Our previous work demonstrated that omega-6 fatty acid rich diets have been associated with increased severity of pancreatic neoplasia, but conversely, omega-3 fatty acid rich diets have chemoprotective effects against Kras-induced pancreatic neoplasia. Also, our more recent data indicates that docosahexaenoic acid (DHA, omega-3 fatty acid) and linoleic acid (LA, omega-6 fatty acid) have opposing effects on protein kinase B (Akt)-mediated mitogenic signaling in Human Pancreatic Duct Epithelial cells in vitro. Furthermore, published studies suggest that one mechanism of action may be through cyclooxygenase (COX)-2 derived prostaglandin E2 (PGE2) secretion leading to cell proliferation. In our current work, we demonstrate that DHA treated HPDE cells do not generate a change in phosphorylated Akt (pAkt) levels and proliferation between cells with or without COX-2 inhibition. As a major maker of proliferation, Akt is often activated by phosphatidylinositol 3 kinase (PI3K), which facilitates conversion of phosphatidylinositol bisphosphate (PIP2) to phosphatidylinositol trisphosphate (PIP3), and PIP3 phosphorylates Akt. Yet the effects of DHA are not directly at the level of PI3K, as HPDE cells treated with a PI3K inhibitor (LY294002), a significantly further reduction in DHA-induced decrease of pAkt. Therefore, it is likely that DHA may not mediate pAkt through PI3K, and we propose that due to its high lipid solubility, DHA may interact with membrane-tethered targets including PIP2. Citation Format: Lingji Zhu, Windel Emman T. Marsacarinas, Daniel R. Principe, Paul J. Grippo. Omega-3 fatty acids may regulate levels of pAkt through a mechanism independent of PI3K. [abstract]. In: Proceedings of the Twelfth Annual AACR International Conference on Frontiers in Cancer Prevention Research; 2013 Oct 27-30; National Harbor, MD. Philadelphia (PA): AACR; Can Prev Res 2013;6(11 Suppl): Abstract nr A31.