Cell Penetrating Peptides-conjugated Insulin-loaded PLGA Nanoparticles:Preparation, Brain Delivery Efficiency Evaluation Following Intranasal Administration

Objective To prepare Insulin-loaded PLGA copolymer nanoparticles(Insulin-NPs), modify cell-penetrating peptides(Tat) on their surface and study the physical and chemical properties and release behavior in vitro. By cell uptake and nasal administration, uptake and brain targeting efficiency can be evaluated.Methods The Insulin-NPs were prepared with Poly lactic acid-glycolic acid(PLGA) by double emulsion solvent evaporation method in this study. The orthogonal design was used to optimize the preparation procedure and Tat was modified on their surface. The transmission electron microscope(TEM) and laser particle size analyzer/Zeta potential were utilized to investigate the physicochemical properties of Insulin-NPs, and the drug encapsulation efficiency, drug loading and release behavior in vitro were studied by BCA and HPLC, respectively. Amino-fluorescein was applied to study uptake of Caco-2cells and brain targeting performance.Results The results showed that Insulin-NPs/Tat were formed of (237.6±4.07)nm in average particle size with polydispersity index of (0.269+0.103), Zeta potential of (+11.34±0.852)mV. The entrapment efficiency and drug loading of Insulin-NPs were (70.56±3.32)%and (4.24±1.10)%, respectively. The in vitro release profiles in PBS buffer at PH7.4was expressed well by Weibull equation, that is lnln(1/1-Q)=0.34051nt-2.2548, R=0.9879. Caco-2cell uptake showed that the amino-fluorescein as a fluorescence probe of Insulin-NPs can be uptaken, the results were affected by drug concentration and the time of experiment. Compared to Insulin-NPs, Insulin-NPs/Tat showed high efficiency of cell-penetrating ability. The efficiency was:Insulin-NPs/Tat> Insulin-NPs(p<0.01). Insulin-NPs also showed brain targeting performance, while the amount of Insulin-NPs/Tat into brain increased. The olfactory bulb and brain targeting efficiency of Insulin-NPs and Insulin-NPs/Tat were:(0.405±0.509),(0.950±0.896) and (2.637±0.615),(3.363±0.239), respectively. The brain targeting efficiency was:Insulin-NPs/Tat>Insulin-NPs(p<0.01).Conclusion PLGA is suitable for preparation of Insulin-loaded nanoparticles with high entrapment efficiency, stable properties and excellent drug sustained release behavior and Tat is an excellent penetrating material with high performance. Insulin-NPs/Tat might be a potential new carrier for intranasal administration. It provides a feasible path for the administration of giant molecule peptide and protein drugs.