The Immune Response Against Non-Viral Vectors
The immune response to non-viral DNA complexes is highly dependent upon the dose and route of administration. Low doses have little effect on organ function and tissue histology. Higher doses, especially of cationic liposomes, induce acute inflammation and profound tissue damage (Yew, 2005). Physical proper-ties of the complexes promote aggregation in the circulation, producing large particulates that are read-ily recognized by the RES (Dash, 1999). The most severe side effects occur after intravenous and intra-pulmonary delivery. Strategies to reduce inflammation and toxicity of non-viral vectors involve: removal of CpG motifs in plasmid DNA, minimizing interaction between the complex and the immune system and use of immunosuppressants in the complex.
Elimination of CpG motifs is achieved by: (a) amplifying plasmids in bacteria that express Sss I methylase to add a methyl group to cytosine residues, (b) use of CpG-free plasmids and/or removal of CpG motifs from plasmids by site-directed mutagenesis, and (c) removal of unnecessary prokaryotic sequences by site-specific recombination to create “minimal” plasmids (Yew, 2004; 2005). These actions have reduced the immune response, but have not eliminated it and compromise transduction efficiency. Covalent attachment of poly(ethylene) glycol to the surface of DNA complexes (PEGylation) has im-proved toxicity and promotes transduction efficiency by preventing aggregation and uptake by the RES (Lee, 2005). PEG molecules are also stabilizing, protective linkers that facilitate attachment of target-ing molecules and antibodies (Haag, 2006). Injection of lipids prior to administration of recombinant DNA has reduced cytokine production by 80% in mice by changing the tissue distribution of the plasmid (Zhang, 2006). Creation of “safeplexes”, lipo- or polyplexes that contain inflammatory suppressor molecules, has also markedly reduced inflammation and toxicity without compromising gene transfer (Liu, 2004). A combination of each of these approaches will most likely be needed for successful and safe clinical gene transfer.