In vivo and in vitro anti-cancer activities and enhanced cellular uptakes of EGF fragment decorated doxorubicin nano-aggregates

Doxorubicin nano-aggregate was prepared for the purpose of epidermal growth factor receptor targeted anti-cancer therapy. An epidermal growth factor fragment composed of 11 amino acids was conjugated to an amino terminal of bi-functional poly(ethylene glycol) and doxorubicin was subsequently conjugated to the other carboxyl terminal of the conjugate. A mixture of the conjugate, free doxorubicin, and triethylamine spontaneously formed nano-sized aggregates in aqueous phase, which was confirmed by transmission electron microscopy and dynamic light scattering. A549 cells incubated with doxorubicin nano-aggregates with the epidermal growth factor fragment showed increased endocytic uptakes of the aggregates compared to unmodified aggregates. Pre-blocking of the epidermal growth factor receptor on the cell significantly decreased a degree of the cellular uptakes. Cytotoxicity of the nano-aggregates was significantly increased when epidermal growth factor fragments were decorated on the surface of doxorubicin nano-aggregates, which was confirmed by a live/dead cell assay and a MTT-based cytotoxicity assay. When doxorubicin nano-aggregates were administered to model animals bearing human lung carcinoma, the epidermal growth factor fragment significantly strengthened in vivo anti-cancer effects of doxorubicin nano-aggregates compared to native doxorubicin or unmodified nano-aggregates. Thus, doxorubicin nano-aggregates decorated with an epidermal growth factor fragment is expected to be a potent anti-cancer agent aiming to tumor tissue over-expressing epidermal growth factor receptors. (C) 2009 Elsevier B.V. All rights reserved.