Study On A Novel Osthol-loaded PH-sensitive Nanoparticles

Objective:In this paper, a novel pH-sensitive nanparticle, Osthol-loaded S100nanoparticle (Ost-S100-NP) and its freeze-drying formulation (Fd-Ost-S100-NP), were prepared with enteric dissolved material Eudragit S100-The Preparation technology,physicochemical characteristics,pharmacokinetics as well as anti-tumor activity were also investigated in order to obtain a safe and effective formulation of Ost.Methods:(1) Ost-S100-NPs and Fd-Ost-S100-NPs were prepared with quasi-emulsion solvent diffusion technique.The orthogonal experiment design was applied to optimize the formulation and preparation procedure with particle distribution, entrapment rate and drug-loading rate as integrated indexes. The particle size, size distribution, encapsulation efficiency and the morphological characteristic were studied individually.(2) DSC, X-ray diffraction, and TEM were applied to evaluate nanoparticle representable structure,and the release in vitro was investigated by dialysis membrane.(3)A random two-way crossover study in mice was adopted. Pharmacokinetic parameters were calculated by3P97program.The tissues distribution and targeting eficiency were evaluated by pharmacokinetic parameters (AUC, AUQ, Cmax, Tmax) and targeting parameters (Te, RTe,TI).(4) The anti-tumor activities in vitro were evaluated by their cytotoxic effects on Hela,A549and SMMC-7721cells with MTT and the flow cytometric analysis was applied to confirm the probable mechanism of antitumor effects.(5) The H22,U14and S180cell line bearing mice were establishmented to evaluate the in vivo anti-tumor activity of Ost, Ost-S100-NPs and Fd-Ost-S100-NPs, including neoplasm volume, inhibition on body weight, inhibition rate on tumor weight, immune organ coefficient; Pathology characteristic of stripped tumor was observed by biological microscope.Results:(1) The average particle size of the prepared nanoparticles was (55.46±2.19) nm, entrapment rate was (98.99±0.16)%, and drug-loading rate was (23.85±0.29)%.(2) In DSC and X-ray diffraction graphs, the diagnostic absorption peak of Fd-Ost-S100-NPs were significantly different from the physicalmixture. TEM graph showed that nanoparticles were sphere and distinct. The curve of the release in vitro showed evidently pH-sensitive and was suitable for Weibull equation.(3) The concentration-time data of Ost and Ost-S100-NPs were fitted as a two-compartment open model in mice. The AUC of Ost-S100-NPs was higher than that of Ost (P<0.05), while the CL(s) significantly decreased (P<0.05).The relative bioavailability of Ost-S100-NPs was163.9%compared.The distribution experiment showed that Ost-S100-NPs changed the distribution of drug in vivo, the drug was dispersed more in liver and kidney, less in spleen.(4)The results of anti-tumor activities in vitro showed that the IC50values of Ost-S100-NP cytotoxicity against Hela,A549and SMMC-7721cells were lower compared to that of Ost. The results of flow cytometric analysis showed that Ost-S100-NPs can stimulate the apoptosis of Hela cell, suggesting probable mechanism of antitumor effects.(5) The antitumor activity suggested that tumor growth was effectively inhibited to all treatment groups with Ost-S100-NP being the most effective,followed by Fd-Ost-S100-NPs and next was Ost. Comparison with Ost, Ost-S100-NPs and Fd-Ost-S100-NPs improved in vivo tumor inhibitory activity, reduced the side effect, and increased therapeutical ratio of Ost in mice bearing tumor.Conclusions:Optimized preparation technique of Ost-S100-NPs was satisfied with the simple process, low cost, and stable reproducibility. The pH sensitivity and sustained drug release characteristic of Ost-S100-NPs were showed in vitro, and were distributed and metabolized extensively in vivo.The absorption and bioavailability of Ost were significantly improved.The anti-tumor activity of Ost was improved both in vitro and in vivo in due to the distinct enhanced permeability and retention effect (EPR) of Ost-S100-NPs. This novel pH-sensitive nanoparticles delivery system had wide perspective and practicability on the clinical application of Ost.