The Study Of The Nano-targeting Agents Of Paclitaxel By The Delivery Of Single Wall Carbon Nanotubes

Paclitaxel have characteristic anti microtubules role, it can promote tubulinpolymerization、inhibit microtubule depolymerized and interfer mitosis. It is mainlyused for treatment of ovarian cancer、breast cancer and non-small cell lung cancer, etc.Paclitaxel is a kind of water insoluble drugs. Now, the paclitaxel preparation saled bydomestic and foreign merchant were almost dissolved by polyoxyethylene castor oilor ethanol.polyoxyethylene castor oil has renal toxicity、allergy and neurotoxicity etc.Paclitaxel lack of tumor targeting,and be easy to cause systemic toxicity, especiallyneutropenia. So the clinical application of paclitaxel is limited.The structure of Single-walled carbon nanotubes(SWCNTs) is very similar tographite. sp2Hybridized carbon is the main component of the carbon in SWCNTs.The surface of SWCNTs is composed of aromatic structure with large surface whichcan be combined with various of organic and inorganic ions, including protein,nucleic acid, drugs and some bioactive molecules, so that the unique ability oftranslocation across mitochondrial membrane can transfer these bioactive moleculesinto the specific site inside the cell. which made SWCNTs a new vector for drugtransfer.The purpose of this project is to prepare NGR-SWCNTs-Paclitaxel deliverysystem and evaluate the antitumor activity in vivo and in vitro. SWCNTs aredispersed in water by amino acid and combined with Paclitaxel through non-covalentbond action, and then connectd with NGR by DSPE-PEG-Maleimide.The study canbe divided into4parts. Chapter1Preformulation studies ofNGR-SWCNTs-PaclitaxelMethods1. Established a HPLC analysis method for Paclitaxel assaying.2.Spectrophotometry for SWCNTs assaying.3.The data were expressed as mean±S.D., and analyzed by statistical softwareSPSS15.0. P-value of <0.05was considered to be statistically significant.Result1. The results show that the linear relation is well between peak area andpaclitaxel concentration in the range of1~40μg/ml.2. The results show that the linear relation is well between absorbance andSWCNTs concentration in the range of2~160μg/ml.Precision and recoveryrate test results are consistent with the methodological requirements.Chapter2Preparation of NGR-SWCNTs-PaclitaxelMethodsThis project makes the final formulation with one-factor experimental designconcerning ratio of charge, surfactant, ultrasound, times and so on, and analysis theeffection on medicine carrying rate and Entrapment rate of these factors. CombinesSWCNTs-Paclitaxel with NGR.ResultPrepare the suspension of NGR-SWCNTs-Paclitaxel and lyophilisate powder ofNGR-SWCNTs-Paclitaxel. The particle size of NGR-SWCNTs-Paclitaxel is about 181nm, zeta potential is about-23mv.Chapter3In vitro activity research ofNGR-SWCNTs-PaclitaxelMethods1. FITC is combined with NGR-SWCNTs-Paclitaxel in order to evaluate theuptake of MCF-7cell in vitro.2. MTT assay and flow cytometry method(FCM) were used to study theeffection on cell proliferation、cell cycle and apoptosis by NGR-SWCNTs-Paclitaxel.Result1. It can be observed that MCF-7cell can devour NGR-SWCNTs-Paclitaxeleffectively, NGR-SWCNTs-Paclitaxel labelled with FITC may transfer into the cellthrough the membrane of MCF-7cell in one hour. and with the extension of timeintake of the complex quantity also increases.2. NGR-SWCNTs-Paclitaxel may inhibit the MCF-7proliferation(P<0.05) witha time-dependent and dose-dependent manner, NGR-SWCNTs-Paclitaxel can inducecell apoptasis and G2/M stage retardant.Chapter4In vivo activity research ofNGR-SWCNTs-PaclitaxelMethods1. The concentration of Paclitaxel in SD rat plasma is measured with HPLC inorder to explore the pharmacokinetics of NGR-SWCNTs-Paclitaxel.2. HPLC is used to test the concentration and distribution of NGR-SWCNTs- Paclitaxel in various tissues of mice with S180sarcoma.Result1. Compared with the control group of Paclitaxel, NGR-SWCNTs-Paclitaxel iscontrolled-released in SD rat plasma,and average retention time extension a times.2. It showing that NGR-SWCNTs-Paclitaxel in vivo distribution changed.NGR-SWCNTs-Paclitaxel is targeting tumor, as well as liver、lung、spleen andkidney.3. The inhibition of NGR-SWCNTs-Paclitaxel on S180sarcoma mice is testedwith Paclitaxel as control, indicating that NGR-SWCNTs-Paclitaxel has bettertherapeutical effect compared with Paclitaxel control(（P<0.05）.Conclution1. Optimized preparation of the NGR-SWCNTs-Paclitaxel.2. NGR-SWCNTs-Paclitaxel can transfer into MCF-7cell through themembrane.3. NGR-SWCNTs-Paclitaxel can inhibit the MCF-7cell proliferation with atime-dependent and dose-dependent manner, and can also induce cell apoptasis andG2/M stage retardant obviously.4. NGR-SWCNTs-Paclitaxel is controlled-released in SD rat plasma.5. NGR-SWCNTs-Paclitaxel have tumor and liver targeting obviously.6. The inhibitory effect of NGR-SWCNTs-Paclitaxel on S180sarcoma mice isobviously.