Asialofetuin liposome-mediated human alpha(1)-antitrypsin gene transfer invivo results in stationary long-term gene expression

The development of nonviral vectors for in vivo gene delivery to hepatocyte s is an interesting topic in view of their safety and tremendous gene thera py potential. Since cationic liposomes and liposome uptake by receptor-medi ated mechanisms could offer advantages in the efficacy of liposome-mediated gene transfer, we studied the effect of liposome charge (anionic vs, catio nic) and the covalently coupled asialofetuin ligand on the liposome surface in mediating human alpha (1)-antitrypsin (hAAT) gene transfer to mice in v ivo. The changes in liposome change were made by adding the following lipid s to the backbone liposomes: anionic phosphatidylserine, cationic N-[1-(2,3 -dioleoyloxy)propyl]-N,N,N-trimethylammonium methylsulfate or a lipopeptide synthesized from dipalmitoylphosphatidylethanolamine and covalently couple d to the cationic nuclear localization signal peptide. Two plasmids contain ing the hAAT gene were used: pTG7101, containing the complete genomic were quence of the human gene driven by the natural promoter, and p216, containi ng the human hAAT cDNA under the control of the CMV promoter. The results i ndicate that both untargeted anionic and cationic liposomes mediate plasma levels of hAAT that decline over time. However, asialofetuin liposomes incr ease the plasma levels of hAAT and can mediate long-term gene expression (> 12 months) with stationary plasma levels of protein. Results from quantitat ive and qualitative reverse transcriptase polymerase chain reaction match t hose from protein plasma levels and confirm both the human origin of the me ssage and the liver as source of the protein. The use of asialofetuin lipos omes in hepatic gene therapy may both increase and prolong in vivo gene exp ression of hAAT and other clinically important genes.