Pharmacokinetics, Tissue Distribution, and Expression Efficiency of Plasmid [33P]DNA Following Intravenous Administration of DNA/Cationic Lipid Complexes in Mice: Use of a Novel Radionuclide Approach

Abstract

The pharmacokinetics, tissue distribution, and efficacy of a systemic gene transfer method were examined in male BALB/c mice (6–8weeks old) using 33P-labeled plasmid DNA for luciferase. The DNA was delivered via tail vein injection in saline ([33P]DNA) or in a cationic lipid formulation ([33P]DNA/lipid). One group of mice received ≈1–3 µCi (45 µg of DNA) of either formulation, and mice were euthanized at 2 and 20 min, and 1 and 24h postdose (2 mice/time point). Blood and plasma radioactivity were quantified, and whole body autoradiographic (WBAR) images were obtained from 20-µm whole body sections. A tissue distribution (TD) study was conducted in a second group of mice, which received ≈4–6 µCi (45–60 µg of DNA) of [33P]DNA or [33P]DNA/lipid. Mice were euthanized at 1.5h (1 mouse; [33P]DNA/lipid) or 24h (2 mice/group), and organ radioactivity and luciferase expression were measured in lung, liver, kidney, spleen, thymus, and parotid salivary gland by direct quantitation methods. Microautoradiography (MAR) was performed on a third group of mice (n=2), which received 3 µCi (45 µg of DNA) of [33P]DNA/lipid and were euthanized at 24h postdose. For WBAR, the [33P]DNA/lipid tissue distribution (% dose equiv/g) at 2min was lung ≫ liver>spleen (red pulp)>kidney (cortex); at 24h the ranking was spleen (red pulp)>liver>lung, kidney (cortex). The [33P]DNA organ distribution observed at 2min was liver ≫ spleen (red pulp)>lung, blood>kidney (cortex); at 24h the ranking was liver, spleen (red pulp)>kidney (cortex)>lung, blood. High levels of radioactivity in bone (cortical, marrow, growth plate) in both groups may represent uptake of the 33P-labeled test articles by the cellular component of the bone marrow, particularly macrophages, as well as deposition of [33P]phosphate in the bone matrix following metabolism of the [33P]DNA. In the luciferase component of the study, no expression was observed in the [33P]DNA group at 24h. The [33P]-DNA/lipid group exhibited expression as early as 1.5h in the lung; at 24 h, expression was seen in all the organs examined. Microautoradiography of 24-h tissue samples revealed radioactivity in hepatic Kupffer cells, reticuloendothelial system cells in the marginal zone of the spleen, and diffusely along alveolar septae with scattered accumulations in alveolar macrophages. The results of the WBAR, TD, MAR, and luciferase assay show that the use of cationic lipids significantly altered the biodistribution and resulting expression of the DNA plasmid. Further, 33P (0.25MeV β, half-life=25days) was shown to be an excellent radionuclide for quantitative WBA and MAR, providing sharp images with less personal hazard and greater ease of handling than 32P (1.71MeV β, half-life=14.3days).