Liposomes for gene transfer and expression in vivo.

Abstract

A recombinant plasmid encoding rat preproinsulin I was encapsulated in large liposomes and injected intravenously into rats. Glycaemia and blood, splenic and hepatic insulin were assayed from 6 h after inoculation. Control animals received (i) empty liposomes, (ii) liposomes carrying the Escherichia coli pBR 322 plasmid, (iii) the free rat insulin I gene, or (iv) no injection. All controls showed unchanged glucose and insulin levels. Six hours after inoculation the treated rats had 72 +/- 5 mg glucose/100 ml of blood, compared with 107 +/- 2 mg/ml for controls. Radioimmunoassay of blood insulin gave 61 +/- 8 microunits/ml (43 +/- 5 microunits/ml for controls). Spleen and liver values were 242 +/- 22 and 204 +/- 20 microunits/g of tissue, respectively (112 +/- 20 and 87 +/- 15 microunits/g in controls). The kinetics and extent of uptake of liposomes by spleen and liver were studied by external gamma-camera imaging after injection of 111In-labelled liposomes. The results paralleled insulin synthesis in the two organs. The insulin gene was localized in liver cells after injection of liposomes containing the plasmid encoding the gene. Livers were processed 4 h after inoculation for isolation of hepatocytes, Kupffer cells and endothelial cells. DNA was purified and exogenous DNA detected by Southern blotting. Kupffer cells were the primary target for gene incorporation with liposomes consisting of phospholipids and cholesterol. Targeting of liposomes to other liver cells was attempted by including lactosylceramide in the liposomes. This increased the amount of the exogenous gene in hepatocytes and particularly in endothelial cells. The efficiency of liposome-mediated gene transfer in vivo is high, since a few per cent of the transferred DNA is taken up by the liver cells and detected.