Abstract Pancreatic cancer (PanCa) is one of the leading causes of cancer-related mortality in the United States due to very limited therapeutic options. Thus, there is an urgent need to develop novel therapeutic strategies for the management of this disease. Paclitaxel (PTX), a natural product derived from the Pacific Yew tree, is an agent typically used in combination therapy for PanCa. Currently, PTX nanoparticles (Abraxane®) along with gemcitabine are in use for the treatment of advanced PanCa. Herein, we have generated and characterized a novel nano-formulation of PTX comprised of PLGA core, stabilized with polyvinyl alcohol (PVA) and coated with poly-L-lysine (PLL). Dynamic light scattering (DLS) characterization indicated that the average particle size of this formulation was approximately 160 nm, and exhibited Zeta potentials of ~ -6.02 mV in 10% serum, along with an outstanding PL loading efficiency. We also evaluated the uptake mechanism of PPNPs in PanCa cells using different endocytosis inhibitors (Genistein, amiloride, nocodazole, chlorpromazine, and methyl beta cyclodextrin). Our results demonstrated that PPNPs were taken up mainly by lipid raft mediated endocytosis pathway in an energy-dependent manner. Next, we evaluated its therapeutic efficacy and underlying molecular mechanisms in human PanCa cells (HPAF-II, Panc1, and MiaPaCa-2) by performing various in vitro experiments. We first performed MTS and colony formation assays to determine the effects of free PTX and PPNPs on growth of PanCa cells. In this experiment, cells were treated with various concentrations of free PTX or abraxane, or PPNPs (2.5-160 nM) for 48 hrs. As a result, the PPNPs formulation showed more therapeutic effect over free abraxane as determined by decrease in cell proliferation and enhanced apoptosis induction in PanCa cells. PPNPs also showed a significant (P<0.01) inhibition in colony formation compared to free PTX. Moreover, PPNPs showed a superior inhibition of the invasive and migratory potential of HPAF-II and Panc1 as determined by commercially available kits (Invitrogen). PPNPs more effectively inhibited expression of GLI-1 and SHH as determined by qPCR and Western blot analysis. In conclusion, our findings suggest that PPNPs formulation has more anti-cancer potential than free PTX/Abraxane against PanCa and could be a novel therapeutic modality for the treatment of PanCa. Citation Format: Bilal B. Hafeez, Andrew E. Massey, Vivek K. Kashyap, Mohammad Sikander, Advit Shetty, Mehdi Chaib, Hassan Mandil, Mohan Yallapu, Meena Jaggi, Subhash C. Chauhan. Novel nano-formulation of paclitaxel for pancreatic cancer therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr LB-011.