Design of a cell‐responsive de‐PEGylation DNA delivery system

Abstract

We are interested in formulating efficient non‐viral DNA delivery vehicles by incorporating multiple functional layers that can be sequentially cleaved off during the delivery process to expose new features. We have established a simplified model vehicle consisting of a polycation‐condensed DNA core attached via linker peptides to a layer of poly(ethylene) glycol (PEG). PEG has been shown to protect biomaterials from salt‐induced aggregation and immune recognition, but PEG coatings can inhibit DNA delivery at the target cell. This project is focused on validating cell‐responsive de‐PEGylation of the vehicle prior to delivery via the cleavage of linker peptides sensitive to matrix metalloproteinase‐1 (MMP‐1), a collagen‐degrading enzyme upregulated by fibroblasts as they migrate through the extracellular matrix. This migration is observed in tumor stroma and at sites of injury, making MMP‐1 secretion a useful targeting signal. Having established the conditions that stimulate MMP‐1 expression by human dermal fibroblasts via Western blotting and immunostaining, we are now investigating the efficiency of vehicle cleavage using light scattering in combination with salt aggregation assays (Figure 1) to detect PEG layer release. Further experiments in this area and cell studies are ongoing. Supported by HHMI, NSF, and UDRF.