Paclitaxel Long-Circulating Nanomicelles Combined With NT21MP To Reverse The Drug Resistance Of Breast Cancer

Objective: Through drug molecular design,modification,and modification technology,the construction of long circulating nanoparticles drug taxol micelle PEG2000-DSPE-TPGS-PTX,and through the cell in vitro experiment verify paclitaxel drug-loading micellar system for breast cancer drug resistance cell line proliferation,migration,invasion and the influence of apoptosis and cell cycle,further explore paclitaxel drug-loading micellar system and its joint whether peptide NT21 MP have combined effects,and related molecular biology mechanism is discussed in this paper.Methods: 1.Thin-film hydration-dispersion method was used to prepare paclitaxel nano-loaded micelles and freeze-dried;2.The morphology of paclitaxel micelles was observed by transmission electron microscopy,and the micelle size distribution and Zeta potential were measured by dynamic light scattering particle size analyzer;3.Determination of encapsulation efficiency and drug loading of paclitaxel nanometer micelles by low-speed centrifugation and UV spectrophotometer,and investigation of storage stability of paclitaxel nanometer micelles,and the storage stability of paclitaxel nanometer drug loading micelles was investigated.4.The uptake of nano-micelle cells by drug-resistant breast cancer cells was observed with the fluorescent probe coumarin-6.5.The effect of Sulforhodamine B(SRB)colorimetric method on the survival rate of paclitaxel nanometer drug loading micelles and paclitaxel nanometer micelles combined with NT21 MP polypeptide on MCF-7 /PR and SKBR3/PR cell lines of breast cancer;6.Cell scratch and Transwell assay were used to detect the effects of PEG-DSPE2000-TPGS-PTXand PEG-DSPE2000-TPGS-PTX with NT21 MP on the migration ability of breast cancer resistant cells drug-resistant breast cancer cells.7.Flow cytometry analysis of the effects of PEG-DSPE2000-TPGS-PTX and PEG-DSPE2000-TPGS-PTX combined with polypeptide NT21 MP on the cell cycle and apoptosis of drug-resistant breast cancer cell lines;8.Western blot assay was used to detect the effects of PEG-DSPE2000-TPGS-PTX and PEG-DSPE2000-TPGS-PTX combined with NT21 MP on P-gp,BCRP,Bcl-2,Bax,Caspase 3,and expression of EMT-related proteins.Results: 1.Paclitaxel nano-loaded micelles were successfully prepared by thin-film-hydration-dispersion method,and the water solubility of the drug carrier system was significantly improved.2.Transmission electron microscopy results showed that paclitaxel long-circulating nano-micelles were spherical in shape,with particle size of about 30-40 nm,Zeta potential of-5.4mV,3.The drug load was detected and calculated by ultraviolet spectrophotometer at 25.37%,and it had certain slow-release effect in PH7.4 environment.3.The cell phagocytic assay showed that the PEG-DSPE2000-TPGS-PTXhad better cellular uptake than the free paclitaxelgroup.4.Compared with the control group and the single-drug paclitaxel group,the paclitaxel nano-micelle group can effectively inhibit the proliferation of drug-resistant cells.5.Compared with the control group and the free paclitaxel group,the paclitaxel nano-micelle group can effectively inhibit the proliferation,migration and invasion of drug-resistant cells.6.Compared with the control group and free paclitaxelgroup,nano micelle taxol group can induce drug-resistant breast cancer cells apoptosis,cell cycle arrest in G2 / M phase of the highest ratio of 7.Compared with the control group and free paclitaxelgroup,PEG-DSPE2000-TPGS-PTX group treatment after breast cancer drug resistant cell lines,cell apoptosis proteins Bax,Caspase 3 expression,resistance to apoptosis protein expression of Bcl-2;8.Compared with the control group and the free paclitaxel group,the expression levels of Emt-related proteins Vimentin and Slug were down-regulated and E-cadherin was up-regulated in the paclitaxel group after the treatment of drug-resistant breast cancer cell lines.9.The biological effects of paclitaxel nano-micelle combined with polypeptide NT21 MP on cell proliferation,migration,invasion and apoptosis were more obvious.Conclusion: 1.We successfully constructedPEG-DSPE2000-TPGS-PTX with a spherical shape,a particle size of about 40 nm,a potential of-5.4mV,and good storage stability,and a certain sustained release effect 2.PEG-DSPE2000-TPGS-PTX can effectively reverse the biological effect of drug resistance of tumor cells in vitro;3.PEG-DSPE2000-TPGS-PTXtcombined with polypeptide NT21 MP can reverse the biological effects of breast cancer resistance and EMT in vitro,showing a better synergistic effect.