Lipoplexes

Despite their relatively lower efficiency, nonviral approaches are emerging as safer alternatives in gene therapy to viral
vectors. Delivery of nucleic acids to the target site is an important factor for effective gene expression (plasmid DNA) or knockdown (siRNA) with minimal side effects. Direct deposition at the target site by physical methods, including ultrasound, electroporation and gene gun, is one approach for local delivery. For less accessible sites, the development of carriers that can home into the target tissue is required. Cationic peptides, lipoplexes, polyplexes and nanoplexes have been used as carriers for delivery of nucleic acids. Targeting
ligands, such as cell targeting peptides, have also been applied to decorate delivery vehicles in order to enhance their efficacy. This review focuses on delivery strategies and recent progress in non-viral carriers and their modifications to improve their performance in targeting and transfection.

Complexes between mRNA and GL67:DOPE:DMPE-PEG5000 (GL67) liposomes were formulated and 15 characterized. Subsequently, the in vitro and in vivo expression characteristics of mRNA/GL67 complexes and pDNA/GL67 16 complexes, each produced at their optimal ratio, were compared in respiratory cells. Transfection of A549 cells with mRNA/ 17 GL67 complexes resulted in a much faster expression than after transfection with pDNA/GL67 complexes. The percentage of 18 GFP-positive cells after mRNA and pDNA transfection peaked after 8 and 24 h, respectively. At these time points the percentage 19 of GFP-positive cells was two times higher after mRNA transfection than after pDNA transfection. Furthermore, the efficacy of 20 mRNA/GL67 complexes was independent of the cell cycle. This was in sharp contrast with pDNA/GL67 complexes that caused 21 only a weak expression in nondividing cells. This confirms that the nuclear barrier is a crucial obstacle for pDNA but not for 22