| Malignant tumor with greatly high death rate remains to be one of the most devastating diseases of the world. Currently, chemotherapy has often been used in cancer treatments. However, the clinical application of most chemotherapeutic drugs is limited by their low selectivity to tissues and cells, in-ignorable toxicity and side effect and the poor solubility. Therefore, there is urgent need to develop a tumor-targeting drug delivery system. In our present work, the folate-conjugated polymer micelles were developed to explore a new tumor targeted delivery way of poorly soluble antitumor drugs.In this study, folate-poly (ethylene glycol)-distearoylphosphatidylethanolamine (FA-PEG3500-DSPE) as the carrier materials of folate-conjugated polymer micelles was synthesized and the 9-nitro-camptothecin(9-NC)-loaded folate-conjugated polymer micelles were prepared and the physico-chemical properties of them were determined. Furthermore, the investigation of tumor-targeting ability and pharmacodynamic evaluation in vitro and in vivo of the folate-conjugated micelles were performed, respectively.Folate-conjugated polymer was synthesized mainly via acylation reaction of carboxy group and amino group catalysized by dicyclohexylcarbodiimide (DCC) and the structure was confirmed by 1H-NMR. The result of RP-HPLC analysis revealed that the purity of FA-PEG3500-DSPE was above 95%. The 9-NC-loaded folate-conjugated polymer micelles were prepared by mixing folate-poly(ethylene glycol)-distearoylphosphatidylethanolamine (FA-PEG3500-DSPE) and methoxy-poly(ethylene glycol)-distearoylphosphatidylethanolamine (mPEG2000-DSPE) to encapsulate 9-NC via film-hydration method and the analysis of particle size, encapsulation efficiencies and drug loading capacity were performed by dynamic light scattering measurer and HPLC, respectively. The mean diameters of optimized micelles was about 24-27 nm and the size distribution were all rather narrow; the encapsulation efficiencies and loading capacity of 9-loaded micelles were 95.7% and 3.24%, respectively. Besides, the study of release in vitro of drug-loaded polymer micelles by dialysis method revealed that the 9-NC-loaded folate-conjugated polymer micelles showed sustained release profiles.The pharmacodynamic study in vitro were carried out by cellular uptake and cytotoxicity assay, and the results revealed micelles with folate modification had higher binding affinity to KB cells, and more effectively enhanced uptake of drug by some tumor cells with over-expressed folate receptor. Compared with free 9-NC and micelles group without folate modification, the antitumor activity in vitro of 9-NC-loaded folate-conjugated micelles (molar ratio, FA-PEG35oo-DSPE/MPEG2ooo-DSPE=1:100) group increased by about 3.7 and 2.0 times, respectively. And the in vivo and ex vivo fluorescence imaging tests showed that folate-conjugated micelles significantly enhanced accumulation in tumor and exhibited more effective tumor-targeting ability in comparison with micelles without folate modification. Finally, the pharmacodynamic evaluation in vivo of folate-conjugated micelles was performed by the tests of growth inhibition of KB tumor subcutaneously implanted into nude mice, and the result revealed that all the micelles groups with folate modification enhanced antitumor activity in vivo to great extent. Among all the test groups including folate-conjugated micelles with different molar ratio of FA-PEG3500-DSPE and mPEG-DSPE2000 (molar ratio, FA-PEG35oo-DSPE/mPEG2000-DSPE=0:100,0.1:100,1:100,10:100), the micelles group with the suitable molar ratio of FA-PEG3500-DSPE and mPEG2000-DSPE (1:100) showed the strongest antitumor efficacy.Obviously, folate-conjugated micelles in current thesis provided a promising active tumor-targeting nanocarrier for lipophilic drugs. And the folate-conjugated polymer ratio in FA-PEG35oo-DSPE/mPEG2000-DSPE mixed micelles had significant influence on the biological behavior and antitumor activity in vivo of micelles, to which great attention should be attached for the study concerning folate-mediated tumor-targeting drug deliver system. |
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