Amphiphilic polyaspartamide copolymer-based micelles for rivastigmine delivery to neuronal cells

A novel polysorbate-80 (PS80)-attached amphiphilic copolymer comprising a hydrophilic α,β-poly(N-2-hydroxyethyl)- d,l-aspartamide (PHEA) backbone and hydrophobic squalenyl-C17 (Sq17) portions was synthesized and characterized; the formation of polymeric micelles was also evaluated. Rivastigmine free-base (Riv), a hydrophobic drug employed to treat Alzheimer’s disease, was chosen as model drug to investigate micelle’s ability to incorporate hydrophobic molecules and target them to neuronal cells. Micelle formation was studied through analyses including fluorescence spectroscopy and 2D 1H-NMR NOESY experiments. Finally, the capacity of Riv-loaded micelles, versus free drug, to penetrate mouse neuroblastoma cells (Neuro2a) was evaluated. 2D 1H-NMR NOESY experiments demonstrated that the PHEA-EDA-Sq17-PS80 copolymer self-assembles into micelle structures in water, with a micelle core formed by hydrophobic interaction between Sq17 alkyl chains. Fluorescence probe studies revealed the CAC of PHEA-EDASq 17-PS80 micelles, which was 0.25 mg mL−1. The micelles obtained had a nanometric hydrodynamic diameter with narrow size distribution and negative surface charge. The PHEA-EDA-Sq17-PS80 micelles incorporated a large amount of Riv, and the system maintained the stability of Riv after incubation in human plasma. An in vitro biological assay evidenced no cytotoxic effects of either empty or loaded micelles on the neuronal cell lines tested. Moreover, the micelles are internalized by neuroblastoma cell lines with drug uptake depending on the micelles concentration.