The cytoplasmic escape and nuclear accumulation of endocytosed and microinjected HPMA copolymers and a basic kinetic study in Hep G2 cells.

The development of macromolecules as drugs and drug carriers requires knowledge of their fate in cells. To this end, we studied the internalization and subcellular Fate of N-(2-hydroxypropyl)methacrylamide (HPMA) copolymers in Hep G2 (human hepatocellular carcinoma) cells. Semiquantitative fluorometry confirmed that galactose was an effective ligand for receptor-mediated endocytosis for Hep G2 cells. The rate of internalization of a galactose-targeted copolymer was almost 2 orders of magnitude larger than that of the nontargeted copolymer. Confocal fluorescent microscopy of both fixed and live cells revealed that the polymer entered the cells by endocytosis. After longer incubation times (typically >8 hours), polymer escaped from small vesicles and distributed throughout the cytoplasm and nuclei of the cells. Polymer that entered the cytoplasm tended to accumulate in the nucleus. Microinjection of the HPMA copolymers into cells' cytoplasm and nuclei indicated that the polymers partitioned to the nucleus. The data from fixed cells was confirmed by microscopy of live, viable cells. To examine the effect of the fluorescent dye on the intracellular fate, polymers with fluorescein, Oregon Green 488, Lissamine rhodamine B, and doxorubicin were tested; no significant differences were observed.