Ternary Polypeptide Nanoparticles with Improved Encapsulation, Sustained Release, and Enhanced In Vitro Efficacy of Carfilzomib

Purpose To develop a new nanoparticle formulation for a proteasome inhibitor Carfilzomib (CFZ) to improve its stability and efficacy for future in vivo applications. Methods CFZ-loaded ternary polypeptide nanoparticles (CFZ/tPNPs) were prepared by using heptakis(6-amino-6-deoxy)-β-cyclodextrin(hepta-hydrochloride) (HaβCD) and azido-poly(ethylene glycol)-block-poly(L-glutamic acid sodium salt) (N 3 -PEG-PLE). The process involved ternary (hydrophobic/ionic/supramolecular) interactions in three steps: 1) CFZ was entrapped in the cavity of HaβCD by hydrophobic interaction, 2) the drug-cyclodextrin inclusion complexes were mixed with N 3 -PEG-PLE to form polyion complex nanoparticles, and 3) the nanoparticles were modified with fluorescent dyes (AFDye 647) for imaging and/or epithelial cell adhesion molecule (EpCAM) antibodies for cancer cell targeting. CFZ/tPNPs were characterized for particle size, surface charge, drug release, stability, intracellular uptake, proteasome inhibition, and in vitro cytotoxicity. Results tPNPs maintained an average particle size of 50 nm after CFZ entrapment, EpCAM conjugation, and freeze drying. tPNPs achieved high aqueous solubility of CFZ (>1 mg/mL), sustained drug release (t 1/2 = 6.46 h), and EpCAM-mediated cell targeting, which resulted in increased intracellular drug accumulation, prolonged proteasome inhibition, and enhanced cytotoxicity of CFZ in drug-resistant DLD-1 colorectal cancer cells. Conclusions tPNPs improved stability and efficacy of CFZ in vitro , and these results potentiate effective cancer treatment using CFZ/tPNPs in future vivo studies.