Extracellular-protein-enhanced cellular uptake of squalenoyl gemcitabine from nanoassemblies

The mechanism by which the squalenoyl prodrug of the anticancer agent gemcitabine was transferred from its nanoassemblies (NAs) to cancer cell lines was investigated by means of in vitro cell uptake experiments. The endocytosis of the NAs was proven not to constitute the main uptake pathway of the squalenoyl gemcitabine bioconjugate (SQdFdC), which indicated that the uptake occurred by passive diffusion. On the contrary, the cell uptake was found importantly influenced by the concentration and nature of the extracellular proteins: it was very limited in protein-free media, reached a maximum at intermediate protein concentrations, and then strongly diminished at high extracellular protein concentrations. This phenomenon was demonstrated not to stem from the alteration of the colloidal stability of the NAs that could have occurred upon modification of the protein concentration. Our results indicated that SQdFdC cell uptake happened through an albumin-enhanced diffusion of molecular SQdFdC across the aqueous medium, rather than through a direct interaction between NAs and cells. Noteworthy, in our in vitro experimental conditions, lipoproteins were detrimental to the cell uptake because they trapped the available SQdFdC molecules without mediating their diffusive capture by the cells.