Preclinical assessment of hepatocyte-targeted MR contrast agents in stable human liver cell cultures.

Abstract

Much effort has been expended in the search for hepatocyte-specific MR contrast agents to improve the detection and characterization of liver tumors. The purpose of this study was to establish human hepatocyte cell cultures to preclinically assess hepatocyte-targeted magnetopharmaceuticals. Cultured human hepatocytes were sandwiched between two layers of collagen preserving both hepatocyte function and morphology over prolonged period of time. Cultures (n = 37) were subsequently used to test different fluorescinated MR contrast agents. Plain and rhodaminated monocrystalline iron oxide particles (MION and MION-rh) and asialoglycoprotein-receptor-specific rhodaminated asialofetuin coupled to MION (MION-ASF-rh) were prepared. Competition experiments of these agents were performed with D(+)-galactose to study the specificity of galactose-mediated cell uptake. To assess the impact of cell integrity on cell uptake, functional experiments with CCl4 were performed. Normal cell cultures showed significantly higher fluorescence light emission after incubation with hepatocyte-directed ASF-MION-rh than after incubation with MION-rh. Competition experiments of ASF-MION-rh with galactose showed a dose-dependent decrease of calibrated fluorescence light emission. Cell cultures treated with CCl4 demonstrated a dose-dependent significant reduction of calibrated fluorescence light emission, indicating reduced uptake of ASF-MION-rh. Our data demonstrate that stable human hepatocyte cell cultures can be used to preclinically assess novel magnetopharmaceuticals. Different contrast agents may be directly compared to each other and may accelerate their preclinical design. Because the assay can be applied to cells from any species, it may represent an ideal test system before clinical trials of new cell-directed MR contrast agents.