| JD27is a small molecule compound and a derivative of active ingredientextracted from Radosia. It is considered as a promising anti-cancer drug candidatebecause of its low toxicity, anti-inflammatory and broad-spectrum anti-cancer activity.However, the clinical application of JD27has been limited by its poor solubility andlow bioavailability.As a carrier material, human serum album (HSA) may be a very promisingnatural carrier of anti-cancer drugs because of its properties of good biocompatible,non-toxic, non-immunogenic, biodegradable. Therefore, in this study human serumalbumin was chosen as a carrier to prepare JD27albumin nanoparticles in order toimprove its water solubility, prolong drug duration and improve the anti-tumor effect.1Preparation, characterization and intracellular kinetics of JD27-loaded human serumalbumin nanoparticles (JD27-NPs)JD27-NPs were obtained using a solvent-chemical crosslinking method. Theformula and technology for preparing JD27-NPs were determined by the uniformdesign method with particle size, encapsulation efficiency and drug encapsulationefficiency as the main evaluation index. The evaluation experiment results showedthat the average particle size, average encapsulation efficiency and average drugloadings of JD27-NPs were (313.6±10.22) nm,(83.5±1.02)%and (7.59±0.09)%,respectively. The preparation method is feasible and reproducible.Lyophilized powder of JD27-NPs was characterized. SEM images suggested thatJD27-NPs were globular with a smooth surface. In vitro releases of JD27andJD27-NPs were evaluated by the method of the dialysis. The experiment resultsindicated that the cumulative release of JD27was95%at20h while cumulativerelease of JD27-NPs was about85%at40h, indicating JD27-NPs had the propertiesof sustained release. A preliminary stability study of JD27-NPs lyophilized powderwas evaluated at4℃. Result of preliminary stability study showed that freeze-driedpowder of JD27-NPs possessed good stability at4℃for6months.MTT assay was performed to measure the effects of JD27and JD27-NPs on inhibition rate of EC-9706cells. The results suggested that24h and48h inhibitionrate of JD27–NPs were significantly higher than JD27(P<0.05). The impact of thedifferent concentrations and treatment times of JD27on cell membranes permeabilitywas detected using fluorescent staining Hoechst33258. The results of Hoechst33258staining experiment showed that JD27could change the permeability of the cellsmembrane, so that the drug could enter into cells more readily and improveanti-tumor effects of JD27. Intracellular kinetics behavior of JD27, JD27-NPs andJD27-β-CD were also evaluated in EC-9706cells using HPLC method. According tothe results of intracellular kinetics, the area under curve (AUC) of JD27-NPs, JD27and JD27-β-CD was (574.88±12.38)(ng/106cells)·h,(366.85±11.95)(ng/106cells)·hand (432.09±15.96)(ng/106cells)·h, respectively; the mean residence time (MRT)was (9.85±0.79) h,(4.41±0.26) h and (6.26±1.30) h, respectively; the eliminationhalf-life (t1/2(Kel)) was (7.37±2.4) h,(3.60±0.23) h and (5.64±0.74) h, respectively.The AUC, MRT and t1/2(Kel) of JD27-NPs were significantly higher than JD27andJD27-β-CD (P<0.05). JD27-NPs could promote JD27intracellular uptake of EC-9706cells, slow the rate of elimination, prolong drug duration and modifypharmacokinetic.2The effect of JD27-NPs on the expression of cell cycle-related proteins of EC-9706cells was detected by Western blotEC-9706cells were treated with JD27-NPs of0,8,16,32μg/ml for24h,respectively. The effect of JD27-NPs on expression of Cdc-2, CyclinB1and CyclinD1was detected by western blot. The results showed that JD27-NPs may induceapoptosis by down-regulation of Cdc-2, CyclinB1and up-regulation of CyclinD1.3The effect of JD27-NPs on enzymic activity of rat CYP1A2, CYP2D6, CYP2E1invitroPhenacetin, dextromethorphan, chlorzoxazone were chosen as probe substrates,The concentrations of probe substrates were determinated using HPLC to investigatethe effect of JD27-NPs on enzymic activity of rat CYP1A2, CYP2D6, CYP2E1invitro. The results showed JD27-NPs could induce the activity of CYP1A2between0and40min, while inhibit its activity between40and80min; low concentrations ofJD27-NPs could induce the activity of CYP2D6, while high concentrations had no effect on its activity; different concentrations of JD27-NPs had no effect on activity ofCYP2E1. The experimental results could predict the interaction of JD27-NPs incombination with other drugs and improve safety and effectiveness of drugcombination. |
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