Characterizing Gene Delivery Efficiency of An Adenovirus Coated with PLL-g-PEG Copolymer

Wednesday, November 10, 2010: 5:25 PM
255 B Room (Salt Palace Convention Center)
Ivy PAL and Joshua D. Ramsey, Chemical Engineering, OKLAHOMA STATE UNIVERSITY, Stillwater, OK

Immunogenicity and promiscuous tropism are two factors that limit the use of adenovirus as a gene therapy vector. Susceptibility of the virus to immune inactivation has been addressed to some extent by coating the virus with polyethylene glycol (PEG). While PEG provides some protection against inactivation, the polymer also severely affects the efficiency of the virus. Poly-L-lysine (PLL) used with PEG may alleviate the negative effects. In this study PLL grafted PEG copolymer (PLL-g-PEG) was synthesized using different molecular weights of PEG and PLL as well as different grafting ratios of PEG to PLL. Synthesis of the copolymer and grating ratios were verified using HNMR spectroscopy. The copolymer was next added to purified adenovirus to form PLL-g-PEG-Ad complexes, and the infectivity of the complex was tested on NIH-3T3 cells and SW480 cells. Both cell lines are deficient in expression of the coxsackie adenovirus receptor (CAR). Since CAR plays an important role in attachment and internalization of the virus, deficiency of the receptor hampers infectivity. Our assay results quantifying gene expression from infected NIH-3T3 cells, however, showed improved infectivity, illustrating that the grafted copolymer assists with attachment and internalization in a CAR-independent fashion. Similar results were obtained for SW480 cells, which are deficient in both CAR and integrin. These results indicate that the complex functions in both a CAR- and integrin-independent manner. The infectivity of the complex was optimized by varying the polymer and polymer concentration. The complexes were further characterized using dynamic light scattering to measure size and surface charge. Complex size was verified by transmission electron microscopy.

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See more of this Session: Nucleic Acid Delivery II
See more of this Group/Topical: Food, Pharmaceutical & Bioengineering Division