Targeted gene delivery to liver sinusoidal endothelial cells by hyaluronanreceptor-mediated endocytosis

The liver sinusoidal endothelial cells (SECs) possess unique hyaluronan rec eptors that recognize and internalize hyaluronic acid(HA). This characteris tic was used in the development of a system for targeting foreign DNA to th e SEC. A gene carrier system was prepared by coupling HA (number-average molecular weight: 1.5 x 10(4)) to poly-l-lysine (PLL, number-average molecular weigh t: 4.6 x 10(4)) in a 1:1 weight ratio by reductive amination reaction. The resulting copolymer (hyaluronate-grafted poly-(L-Lysine): PLL-g-HA) was iso lated and lyophilized. The PI,L-g-HA thus formed was mixed with various amo unts of DNA in 154 mM NaCl at 4 degrees C. Neither turbidity nor precipitat ion was observed in PLL-g-HA/DNA mixtures over the copolymer (based on PLL) to DNA charge ratio from 0 to 2. This indicated that inter-polyelectrolyte complex formation between PLL-g-HA and DNA exhibited minimal self-aggregat ion, explaining the highly soluble nature of the complex. The agarose gel r etardation assay revealed that the titration point representing the minimum proportion of PLL-g-HA required to retard the DNA completely occurred at a 1:1 copolymer to DNA charge ratio. Intravenous injection of the [P-32]pSV beta-Gal plasmid (an expression plas mid encoding lacZ) complexed to PLL-g-HA in Wistar rats demonstrated specif ic hepatic targeting with >93% of the injected counts taken up by the liver in 1 h. Further using an FITC-labeled DNA, it was shown that the PLL-g-HA/ DNA complex was distributed exclusively in the SEC. Seventy-two hours after injection of 90 mu g of pSV beta-Gal in a PLL-g-HA-complexed form, a large number of SEC expressing beta-galactosidase were detected. The new PLL-g-H A/DNA carrier system permits targeted delivery of exogenous genes selective ly to the liver SEC.