| Glioma is the most common malignant tumor of the central nervous system(CNS)with an annual average incidence rate of 6 per 100 thousand individuals.The median survival of glioma patients is not more than 14.6 months,and only 3-5%of patients can survive for 5 years.Surgery combined with post-operative radiotherapy and chemotherapy still is currently the preferred treatment method for glioma.However the operation cannot completely remove the tumor;the side effect of radiotherapy is large,and the survival quality of the patients is greatly reduced;about 98 percent of small molecular drugs and almost 100 percent of macromolecular drugs can’t be delivered through the BBB,leading to poor results and easy relapsing,due to exists of blood brain barrier,immune escape capacity of glioma and characteristics of tumor infiltration growth.Therefore,it is an urgent problem to explore new drugs and new therapies to treat glioma.Docetaxel(DTX)is a potent anticancer drug that appears to be 2 to 4-fold more potent than paclitaxel.However,similar to many chemotherapeutic agents,DTX has a fairly low solubility in water and a poor ability to penetrate the blood brain barrier(BBB).Moreover,even if transported across BBB,it is difficult to penetrate through the deeper tumor tissue and into the core of the tumor suppressing tumor growth.In recent years,studies have shown that the ultrasound combined with microbubbles can effectively,reversibly and safely open BBB.This is of great significance for the treatment of glioma.At present,more and more nanomaterials are applied to the targeted delivery of anti-tumor drugs.Drugs coated with nanoparticles not only can keep drug properties stabilization,but also can target tumor due to enhanced permeability and retention effect(EPR).On the basis of this,the surface of nanoparticles can be combined with a specific ligand that can combine with receptor expressed on the surface of tumor cells,so as to realize the active targeted transport of drugs to the tumor site.Active targeting can further improve the drug concentration of tumor sites base on EPR and enhance anti-tumor effect.In recent years,it has become a research hotspot in drug targeting delivery.In this paper,selecting polyethylene glycol-polylactic acid(PEG-PLA)as nanoparticles.Based on the characteristics of tumor cell microenvironment,and targeting low density lipoprotein receptor(LDLR)related proteins expressing highly on glioma cells by peptide named Angiopep-2.Designing and synthesis targeted-reduction response micelles loaded with docetaxel.Secondly,the physical and chemical properties of the polymer micelle,the ability of reducing responsivewas ness and the antitumor characteristics were studied.In situ model of brain glioma established and the regulation of BBB opening using ultrasound combined with microbubbles was studied.On the basis of this,a study on the treatment of tumor-bearing mice was carried out.The results obtained were as follows:(1)Through the addition reaction,Angiopep-2 was successfully connected to PEG-PLA,and the connection efficiency is 51.25%.Pyrene fluorescence probe measured the critical micelle concentration(CMC)of PEG-PLA and PEG-SS-PLA was 5.66 mg/L and 9.33 mg/L,reflected the good stability against dilution.Finally,SANP micelle was obtained by dialysis.The uniform size of micelle was observed by transmission electron microscope.The dynamic light scattering method measured a diameter of 112.7 nm,which can keep the particle size stable within a week.(2)The capacity of DTX was investigated by high performance liquid chromatography(HPLC).The micelles displayed high drug loading capacity of DTX with an encapsulation efficiency(EE)of 92%and a drug loading capacity(DLC)of 4%.The drug release rate was determined by dialysis method.The in vitro release behavior showed that the release of DTX from micelles was 70.06%under physiological conditions over a period of 48 h.Nevertheless,when added 10 mM dithiothreitol(DTT),the release rate of micelles obviously increased(83.06%,12 h).The release profile of DTX from micelles exhibited a typical redox-responsive property.(3)On the above basis,C6 cells of rat brain glioma were studied.By MTT assay,the cytotoxicity effect of the loaded micelles on C6 cells was determined,and the results showed that the SANP group had obvious cytotoxicity effect,and the killing rate was 88.2%.Using coumarin-6 instead of DTX,micelle have fluorescent properties.Then by laser confocal microscope and flow cytometry analysis the micelle target ability in vitro,proves the SANP can improve cells on drug intake,with active targeting and the characteristics of intracellular microenvironment to stimulate drug release.The study of Annexin V/PI double dye combined with flow cytometry found that the SANP group could mediate the apoptosis of C6 cells,and the apoptosis rate was 62.92%.(4)In situ model of brain glioma of ICR mice was established.Study had proved US combined with microbubbles could be reversibly open BBB,and after 2 h US treatment BBB open extremely.After the treatment of mice with DTX micelle,the main organs of the mice were observed:the main organs were not significantly damaged by this therapy,and this therapy had safety.In order to study the targeting of micelle in mice,using near infrared dye DIR780 instead of DTX was used in vivo imaging.The imaging results show that:ultrasound combined with microbubbles can significantly improve the concentration of micelles in glioma,and the SANP joint ultrasonic group has better targeting effect.In the experimental study on the treatment of tumor-bearing mice with micelle loaded with DTX,ex vivo images showed that SANP combined with the US group significantly inhibited the proliferation of tumor,and effectively prolonged the survival cycle of the tumor-bearing mice,the median survival rate is 45 days. |
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