Formulation of highly soluble poly(ethylene glycol)-peptide DNA condensates

Two poly(ethylene glycol) (PEG)-peptides were synthesized and tested for their ability to bind to plasmid DNA and form soluble DNA condensates with reduced spontaneous gene expression. PEG-vinyl sulfone or PEG-orthopyridyl disulfide were reacted with the sulfhydryl of Cys-Trp-Lys(18) (CWK18) resulting in the formation of nonreducible (PEG-VS-CWK18) and reducible (PEG-VS-CWK18) PEG-peptides. Both PEG-peptides were prepared on a micromole scale, purified by RP-HPLC in >80% yield, and characterized by H-1 NMR and MALDI-TOF. PEG-peptides bound to plasmid DNA with an apparent affinity that was equivalent to alkylated (Alk)CWK18, resulting in DNA condensates with a mean diameter of 80-90 nm and xi (zeta) potential of +10 mV. The particle size of PEG-peptide DNA condensates was constant throughout the DNA concentration range of 0.05-2 mg/mL, indicating these to be approximately 20-fold more soluble than AlkCWK(18) DNA condensates. The spontaneous gene transfer to HepG2 cells mediated by PEG-VS-CWK18 DNA condensates was over two orders of magnitude lower than PEG-SS-CWK18 DNA condensates and three orders of magnitude lower than AlkCWK(18) DNA condensates. PEG-VS-CWK18 efficiently blocked in vitro gene transfer by reducing cell uptake. The results indicate that a high loading density of PEG on DNA is necessary to achieve highly soluble DNA condensates that reduce spontaneous in vitro gene transfer by blocking nonspecific uptake by HepG2 cells. These two properties are important for developing targeted gene delivery systems to be used in vivo.