Targeting specificity and pharmacokinetics of asialoorosomucoid, a specific ligand for asialglycoprotein receptor on hepatocyte.

Objective: To testify that the asialoorosomucoid (ASOR) prepared by us has liver-targeting specificity and to investigate its pharmacokinetic characteristics.

Methods: The distribution of 125I-ASOR in vivo was determined by single photon emission computed tomography (SPECT) and immunohistochemical technique after 125I-ASOR was injected into Sprague-Dawley (S-D) rats through their caudal veins. In vitro, different doses of pEGFP-N1 plasmid were transfected into both HepG2 cells and HT1080 cells with the use of ASOR-poly-L-lysine. At 24 and 48 h after transfection, the expression of green fluorescent protein (GFP) was determined under fluorescent microscope. Pharmacokinetic parameters were calculated according to two-compartment open system model with first-order kinetics.

Results: SPECT images showed that 125I-ASOR was located only in liver/stomach and root of caudal vein/bladder at 10 min after injection. The 125I-ASOR radioactivities of organs taken out from S-D rats were different at different times, and about 63% of 125I-ASOR was located in the liver at 10 min after injection. At 30 min after injection a peak of radioactivity was seen in stomach. The times of these two radioactivity peaks were different. Immunohistochemical study of liver frozen sections showed that ASOR was combined mainly with hepatocyte membrane, especially in areas with rich blood flow. In vitro study showed that ASOR targeted specifically cells with asialoglycoprotein receptor (ASGr). GFP expression was detected in HepG2 cells but not in HT1080 cells. Furthermore, the more quantity of pEGFP-N1 transfected and the longer expression time, the higher GFP expression level was in HepG2 cells. The 125I-ASOR pharmacokinetics equation for liver was Ct=662216e-3.362t+8896e-2343t. 125I-ASOR was excreted from liver slowly after an initial rapid decrease. The pharmacokinetic equation for stomach was Ct=-114815e-1.7t+1148153e-15t and the half-life of 125I-ASOR in stomach was 4.62 h.

Conclusions: ASOR prepared by us could be an efficient gene transfer vector, ASOR was distributed mainly in the liver and stomach and had high targeting specificity to hepatocytes or hepatic originating cells.