Study Of RGD-mediated Chitosan-modified Carbon Nanotubes For Drug Delivery System

Objective: In this paper, RGD-decorated chitosan-modified single walled carbonnanotubes (SWCNTs) were constructed for targeted delivery of docetaxel. Thephysicochemical properties of carrier complexs were characterized and the drug release,cytotoxicity, cellular uptake, intracellular localization and cellular uptake mechanismswere evaluated in vitro. In addition, the anti-tumor effects of carrier complexs were alsostudied. This study can provide experimental evidences for the construction offunctionalized SWCNTs as a novel drug delivery system.Methods: SWCNTs were purified by acid oxidation, then Chitosan (CS) modifiedO-SWCNTs (CS-SWCNTs) were synthesized and characterized. The O-SWCNTs wereconjugated with docetaxel via π–π stacking interaction (SWCNT-DTX), and thennoncovalently functionalized with RGD-conjugated CS to prepareRGD-CS-SWCNT-DTX complex. Functionalized SWCNTs were characterized byultraviolet-visible spectrophotometry (UV-Vis), fourier transform infrared resonancespectroscopy (FT-IR), raman spectroscopy, transmission electron microscopy (TEM) andatomic force microscope (AFM). The drug loading efficiency was determined by HPLC.The drug release from RGD-CS-SWCNT-DTX was studied at pH5.0and pH7.4conditions. In vitro antitumor activities of RGD-CS-SWCNT-DTX were studied in A549and MCF-7cells. The cell viability of carriers was determined in tumor cells using theMTT assay. The cellular uptake of RGD-CS-SWCNT-DTX was analyzed usingfluorescence microscope and flow cytometry. The intracellular localization ofRGD-CS-SWCNT-DTX in A549cell was observed by confocal microscopy.The effects ofcell inhibitors on tumor cells uptake of RGD-CS-SWCNT-DTX were quantitative analyzedby flow cytometry.The anti-tumor effects of RGD-CS-SWCNT-DTX were evaluated inA549tumor-bearing nude mice. Results: According to a series of oxidative modification and characterization,O-SWCNT and CS-SWCNT were successfully prepared and easily dispersed in water. Theresults of MTT assay showed that CS-SWNTs and O-SWNTs have little toxicity to A549and MCF-7cells. The studies of phycochemical properties demonstrated thatRGD-CS-SWCNT-DTX was successfully synthesized. The drug loading efficiency ofSWCNT-DTX was up to50%and the amount of chitosan in the complex was about40%.The AFM and TEM images show that the diameter of RGD-CS-SWCNT-DTX was10-20nm and the length were200-500nm. The drug release of RGD-CS-SWCNT-DTX at pH5.0was68%, while it was49%at pH7.4, which indicating that it had a pH responsive drugrelease. RGD-CS-SWCNT-DTX could significantly increase the cytotoxicity of A549cells,but failed to increase the toxicity of MCF-7cells. Cellular uptake of carriers wassignificantly higher in A549than in MCF-7cells, and the extent of cellular uptake ofRGD-CS-SWCNT-DTX was higher than that of CS-SWCNT-DTX in A549cells.Confocal microscopy images showed that RGD-CS-SWCNT-DTX can enter the lysosomeby endocytosis or penetrate the cellular membrane into the nucleus.RGD-CS-SWCNT-DTX was internalization into A549cells through clathrin-mediatedendocytosis and caveolae-mediated endocytosis, while it was mainly throughcaveolae-mediated pathway in MCF-7cells. RGD-CS-SWCNT-DTX significantly reducedtumor volume in A549lung cancer cells-bearing nude mice.Conclusion: RGD-CS-SWCNT-DTX shows higher drug loading efficiency,significant anti tumor effects, effective tumor targeting and controlled drug releaseproperties. This study provides a theoretical basis for a novel SWCNT-based targeted drugdelivery.