2008 NSTI Nanotechnology Conference and Trade Show - Nanotech 2008 - 11th Annual

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TechConnect Summit
Clean Technology 2008

Epidermal Growth Factor Receptor-Targeted Engineered Gelatin Nanovectors for Gene Delivery and Transfection

P. Magadala, M. Amiji
Northeastern University, US

Keywords:
EGFR, targeted gene delivery, Pancreatic cancer

Abstract:
Purpose: Over 80% of human pancreatic cancer expresses epidermal growth factor family of receptors on the cell surface. In this study, we have examined the potential of EGFR-targeted gelatin-based engineered nanovectors (GENS) for plasmid DNA delivery and transfection in pancreatic cancer cells. Methods: Thiol-modified type B gelatin was synthesized by reacting gelatin with 2-aminothiolane (Traut’s reagent). EGFP-N1 plasmid DNA, expressing enhanced green fluorescent protein (EGFP), was encapsulated in thiolated gelatin nanoparticles by the solvent displacement method. The nanoparticle surface was modified with EGF receptor binding peptide (-AAEEIYAARRG-) through a poly(ethylene glycol) (PEG) spacer. DNA encapsulation efficiency was measured using PicoGreen® fluorescence assay and the stability was evaluated by agarose gel electrophoresis. Panc-1 human pancreatic adenocarcinoma cells were established in culture. The cytotoxicity of control and surface modified GENS was measured using the MTS (formazan) assay. GFP transfection efficiency was quantitatively evaluated by flow cytometry and qualitatively by fluorescence microscopy. Results: Surface-modified thiolated gelatin nanoparticles of ~200 nm in diameter were reproducibly synthesized. SHGel-5 and SHGel-10, made by reacting 1 gram of gelatin with 5 mg and 10 mg, respectively, of 2-iminothiolane showed relative cell viability of 97% and 95% at the highest concentration of 200 g/mL. In comparison, poly(ethyleneimine), a cationic cytotoxic polymer, showed relative cell viability of only 40% at the same concentration. The DNA loading efficiency of control and surface-modified GENS was >95%. With unmodified GENS, the GFP transfection efficiency in Panc-1 cells was 19%. Upon PEG-modification, the transfection efficiency increased to 30%. EGFR-targeted peptide conjugation to PEG modified nanoparticles led to further increase in the transfection efficiency of GFP. Conclusions: The results of this study showed that EGFR-targeted gelatin-based nanovectors can serve as a safe and efficiency carrier for potential in vivo gene therapy in pancreatic cancer.


Nanotech 2008 Conference Program Abstract