Development of dextran nanoparticles for stabilizing delicate proteins

Fei Wu,Tuo Jin,Jing Su,Liangming Wei,Weien Yuan,Zhihua Zhou

doi:10.1186/1556-276x-8-197

Abstract

One of the most challenging problems in the development of protein pharmaceuticals is to deal with stabilities of proteins due to its complicated structures. This study aims to develop a novel approach to stabilize and encapsulate proteins into dextran nanoparticles without contacting the interface between the aqueous phase and the organic phase. The bovine serum albumin, granulocyte-macrophage colony-stimulating factor (GM-CSF), granulocyte colony-stimulating factor (G-CSF), β-galactosidase, and myoglobin were selected as model proteins. The proteins were added into an aqueous solution containing the dextran and polyethylene glycol, and then encapsulated into dextran nanoparticles by aqueous-aqueous freezing-induced phase separation. The encapsulation efficiency and recovery of dextran nanoparticles were determined. The dextran nanoparticles loaded with proteins were characterized by scanning electron microscopy and particle size analysis. The protein aggregation was determined by size-exclusion chromatography-high-performance chromatography, and the bioactivity of proteins recovered during formulation steps was determined. The bioactivity of GM-CSF, G-CSF, and β-galactosidase were examined by the proliferation of TF-1 cell, NSF-60 cell, and ortho-nitrophenyl-β-galactoside assay, respectively. The results of bioactivity recovered show that this novel dextran nanoparticle can preserve the protein's bioactivity during the preparation process. LysoSensor™ Yellow/Blue dextran, a pH-sensitive indicator with fluorescence excited at two channels, was encapsulated into dextran nanoparticles to investigate the ability of dextran nanoparticles to resist the acidic microenvironment (pH < 2.5). The result shows that the dextran nanoparticles attenuate the acidic microenvironment in the poly (lactic-co-glycolic acid) microsphere by means of the dilution effect. These novel dextran nanoparticles provided an appealing approach to stabilize the delicate proteins for administration.

Highlights

Proteins play crucial roles in virtual pharmaceutical science covering cytokine, antibody, enzyme, supplements, and vaccine [1,2,3,4,5]
Proteins and polyethylene glycol (PEG) were dissolved in dextran solutions and aqueous solution, respectively
After these two solutions were mixed to get a clear solution, the solution was frozen dried under vacuum and washed with dichloromethane to give fine dextran nanoparticles loaded with proteins

Summary

Introduction

Proteins play crucial roles in virtual pharmaceutical science covering cytokine, antibody, enzyme, supplements, and vaccine [1,2,3,4,5]. Considerable progress in the molecular biology and genetic engineering during the past 3 decades has led to a significant increase in the number of approved protein drugs covering nearly 150 diseases [6]. Protein has several advantages over small molecule drugs [7]. The aggregation of the protein can result in an immune response. The protein aggregation either reveals new epitopes recognized as non-self or leads to the spacing of the epitopes known to break self-tolerance [4]. The protein aggregates should be prevented during the nanoparticle preparation steps

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Conclusion