Globular Protein-Coated Paclitaxel Nanosuspensions: Interaction Mechanism, Direct Cytosolic Delivery, and Significant Improvement in Pharmacokinetics

About 40% of the marketed drugs and 70-90% of new drug candidates are insoluble in water and therefore poorly bioavailable, which significantly compromises their therapeutic effects. A formulation of nanosuspensions achieved by reducing the pure drug particle size down to seb-micron range is one of the most promising approaches to overcome the insolubility. However, the nanosuspension formulations are subject to instability because of nucleation and particle growth. Therefore, a stabilizer is needed to be incorporated into the nanosuspension formulation during the preparation process to suppress the aggregation of drug particles. β-LG, a globular protein, is broken by heat-induced denaturation, and its hydrophobic area is exposed, which allows it to associate with organic particles. PTX, an insoluble drug, is widely used for the clinical treatment of human cancer. However, this drug's clinical application is greatly limited by intrinsic defects including poor solubility, adverse side effects, and poor tumor penetration. In this study, we prepared β-LG-stabilized PTX nanosuspensions (PTX-NS) by coating the protein onto nanoscaled drug particles, investigating the stabilization effect of β-LG on PTX-NS, and evaluating its in vitro and in vivo performance. PTX-NS with a diameter of approximately 200 nm was easily prepared. β-LG produced significantly stabilized effect on PTX-NS via the interaction between the hydrophobic area of the protein and the hydrophobic surface of the drug particles, which resulted in a conformational change of the protein, the loss of both secondary and tertiary structures, and the transition of Trp residues to a less hydrophobic condition. Importantly, unlike other conventional nanoparticles, PTX-NS could directly translocated across the membrane into the cytosol in an energy-independent manner, without entrapment within the endosomal-lysosomal system. Moreover, compared with Taxol, PTX-NS increased AUC and Cmax by 26- and 16-fold, respectively, and prolonged T1/2 by 314-fold. As expected, PTX-NS had better in vitro and in vivo antitumor activity compared to PTX alone. Additionally, β-LG is cyto- and bio-compatible, and PTX-NS is not toxic to healthy tissues. In conclusion, the present study has suggested the high potency of globular proteins, such as β-LG, as novel biomaterials for nanosuspension platform to improve the drug delivery for disease treatment.