Study On Reversal Multidrug Resistance Of TPGS-Modified Docetaxel Loaded Liposomes

Objective To effectively improve the anti-tumor effect,reduce severe side effects and reverse multidrug resistance,this work proposed to use liposomes as drug carries and prepared a TPGS-modified docetaxel liposome delivery system.This study evaluated the anti-tumor effect and reversal multidrug resistance of the drug delivery system in vitro and in vivo.The TPGS-modified docetaxel liposome provides a basis for the development of new anti-tumor drug delivery system.Method 1 HPLC method was established to determine the content of docetaxel;the encapsulation efficiency of DTX liposomes was measured by cryogenic ultracentrifugation.2DSPC,cholesterol,DSPE-PEG2000and TPGSwereusedaslipids.DSPC-DTX-liposome,PEG-DTX-liposome,and TPGS-DTX-liposome were prepared by film dispersion method using docetaxel as a model drug.Encapsulation efficiency and drug loading of liposomes were used as indicators to optimize prescriptions.The properties of the resulting doctaxel liposomes were also investigated.3 Using A549,A549/DDP,MCF-7,MCF-7/ADR as model cells,the cytotoxicity,cell uptake,P-gp inhibition assay and apoptosis assay were used to evaluate the reversal multidrug resistant effect of different docetaxel formulations in vitro.4 The A549/DDP xenograft model of BABC/L nude mice was established.After being administered via tail vein injection,the tumor volume and body weight of nude mice were measured to evaluate the multidrug resistant effect of different docetaxel formulations in vivo.Results 1 A HPLC method for the determination of docetaxel was established.The docetaxel had a good linearity in the range of 5~60?g/ml.The precision,reproducibility andmethod recovery rate were in line with determination of content.The recovery of the method were between 98%and 105%,and the RSD values were less than 1.0%,which met the measurement requirements for the determination of content.2 The drug/lipid was 1:15,and the hydrated buffer was 10mmol/ml PBS at pH 7.4,the obtained docetaxel liposomes were prepared by lipid-film hydration method.The particlesize,zetapotential,encapsulationefficiencyanddrugloadingof DSPC-DTX-liposomewere126.2±4.0nm,-4.78±0.14mv,90.8±5.1%and6.0±0.1%,respectively.The particle size,zeta potential,encapsulation efficiency and drug loading of PEG-DTX-liposome were respectively 137.0±0.7nm,-0.165±0.01mv,96.2±0.2%and 5.9±0.3%.The particle size,zeta potential,encapsulation efficiency and drug loading of TPGS-DTX-liposome were 140.9±6.0nm,-0.196±0.08mv 99.0±0.9%and 8.4±0.01%.The three DTX-loaded liposomes showed sustained release without burst release.TEM showed spherical unilamellar nanostructures of liposomes.After 21 days at 4°C,the changes particle size and encapsulation efficiency were not significant,indicating that DTX-loaded liposomes have good stability.3 The cytotoxicity results showed that the cell viability of the TPGS-DTX-liposome group was lower than other groups,and the IC50 value was significantly lower than other groups(p<0.01).From the results of cell uptake experiments,the fluorescence intensity of the Coumarin-6-TPGS liposome group was stronger than that of the other groups,indicating that the cancer cells enhanced the uptake of Coumarin-6-TPGS liposomes.Compared with the other two groups of blank liposomes,the blank TPGS liposome group could significantly increase the uptake of Rhodamine 123 in A549/DDP and MCF-7/ADR cells(p<0.05),resulted inhibition of P-gp,and enhanced accum?lation of drug in cancer cells.The total apoptosis rate of TPGS-DTX-liposome in A549,A549/DDP,MCF-7and MCF-7/ADR cells was 62.85%,44.21%,55.9%,32.6%,respectively,indicating that TPGS-DTX-liposome can promote the apoptosis of resistant and sensitive cells.The apoptosis rate of TPGS-DTX-liposome was significantly different from other docetaxel formulations(p<0.05).4 The tumor volume in the TPGS-DTX-liposome was significantly smaller than that in the other two liposomes(p<0.05).After 14 days,body weight of the nude mice in three docetaxel load liposomes did not significantly change,while the Taxotere~?group was significantly reduced(p<0.05),indicating that Taxotere~?had serious side effects on the nude mice.Conclusion The TPGS-DTX-liposome with higher encapsulation efficiency and stability was prepared by film dispersion method.The results of in vitro showed that the TPGS-DTX-liposome could significantly inhibit cell growth and promote apoptosis.The uptake of Coumarin-6-TPGS liposomes by cancer cells was significantly enhanced.The blank TPGS liposomes inhibited the overexpression of P-gp,resulted the reduction of drug efflux,and increased drug accumulation in cells.The results of in vivo showed that TPGS-DTX-liposome could inhibit tumor growth and have no obvious toxic side effects on nude mice.These results confirmed that TPGS liposomes could reverse multidrug resistance.Therefore,TPGS-modified liposomes have potential applications in clinical treatment of multidrug resistant tumors.