ABSTRACT: We have designed an amphiphilic prodrug of gemcitabine (dFdC) by its covalent coupling to a derivative of squalene, a natural lipid. The resulting bioconjugate self-assembled spontaneously in water as nanoparticles that displayed a promising in vivo anticancer activity. The aim of the present study was to provide further insight into the in vitro subcellular localization and on the metabolization pathway of the prodrug. Cells treated with radiolabelled squalenoyl gemcitabine (SQdFdC) were studied by differential detergent permeation, and microautography coupled to fluorescent immunolabeling and confocal microscopy. This revealed that the bioconjugate accumulated within cellular membranes, especially in those of the endoplasmic reticulum. Radio-chromatography analysis proved that SQdFdC delivered dFdC directly in the cell cytoplasm. Mass spectrometry studies confirmed that gemcitabine was then either converted into its biologically active triphosphate metabolite or exported from the cells through membrane transporters. To our knowledge, this is the first description of such an intracellular drug delivery pathway. In vitro cytotoxicity assays revealed that SQdFdC was more active than dFdC on a transporter-deficient human resistant leukemia model, which was explained by the subcellular distribution of the drugs and their metabolites. The squalenoylation drug delivery strategy might, therefore, dramatically improve the efficacy of gemcitabine on transporter-deficient resistant cancer in the clinical context.
Journal of Controlled Release 10/2010; 147(2):163-70. · 5.73 Impact Factor