Abstract
Gene therapy is a promising new approach for medicine, and able to target a wide variety of diseases. [1]. Cationic liposome (CL)-DNA complexes (also known as lipoplexes) are desirable non-viral gene vectors because of low immunogenicity, ease of scale-up procedures, etc.. To be viable for in vivo applications, CL-DNA complexes need to be stable in circulation, which can be achieved by addition of poly-(ethylene glycol) (PEG)-lipids. Unfortunately, PEG-lipids interfere with complex-cell membrane interactions that are vital for particle attachment and uptake and can inhibit membrane fusion required for endosomal escape. The benefit of increased stability from PEG-lipids, thus, comes at the cost of a reduction in transfection efficiency (TE).To recover the resulting loss in TE, we have designed and synthesized a hydrolysable acid-labile PEG-lipid (HPEG-lipid, PEG MW 2000) which is stable at physiological pH, but is cleaved at low pH. The natural acidification process of endosomes will liberate the PEG from the lipoplexes. We have studied the colloidal stability and transfection efficiency of these HPEG-lipoplexes in mammalian cell cultures. The acid sensitivity and the colloidal stability were characterized by TLC and dynamic light scattering, respectively. The HPEG-lipid is stable at neutral pH for more than 24 h, but degrades completely within 1 h at pH 4, leading to particle aggregation. HPEG-lipoplexes show lower toxicity and enhanced TE in comparison to lipoplexes stabilized with pH-stable PEG-lipids. Live-cell images show that both pH-sensitive and pH-stable PEG-lipoplexes were internalized to quantitatively similar particle distributions within the first 2 h of incubation. Thus, the increased TE of the HPEG-lipoplexes can be attributed to efficient endosomal escape, enabled by the novel HPEG-lipid. Funded by NIH-GM-59288 and NSF-DMR-1101900.[1] Ewert, K. K. et al, Topics Curr. Chem. 2010, 296, 191-226
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