Study On The Prepartion On And AnTI-Tumor Effect Of Folate-Conjugated Chitosan Nanoparticles

In recent years, there has been a great advancement in the field of chemo therapy and a marked extension on the survival time of suffers from cancer. However, due to the lower specificity of current anti-cancer drugs, normal cells may be damaged while tumor cells are destroyed, resulting in the apparent toxic reaction during the period of treatment. As a result,the all-important challenge for researchers is to improve the specificity and safety of anti-cancer drugs. Among all those researches, targeted drug via the folate receptor, based on the fact that folate receptor can be overexpressed on the surface of some kinds of tumor cells but scarcely expressed on normal cells, has drawn great attention by researchers. As an inexpensive, nontoxic,biocompatiable macromolecule which can be easily modified, chitosan has been extensively used as carrier for drug. What's more, nanotechnology can enhance the stability of drug and achieve sustained-release and controlled-release.In this study, we chose chitosan(CS) as a delivery system material for anticancer drug.The folate-conjugated chitosan was prepared by the conjugation of activated folate and chitosan via amino groups. The folate-conjugated,doxorubicin-loaded chitosan nanoparticles(DOX/FOL CSNp) were prepared by ionically cross-linking method. Several characteristics of nanoparticles such as morpHology, particle size, Zeta potential and drug entrapment efficiency, loading capacity and vitro release kinetics were evaluated.The compound of chitosan or folate-conjugated chitosan grafted on fluorescein isothiocyanate(FITC) by chemical bond was prepared. Then FITC-labelled nanoparticles were prepared by ionically cross-linking method .Using HeLa cell(FR+) as model cell, The interaction of nanoparticles and HeLa cell was investigated. The intracelluar uptake of nanoparticles were observed under inverted fluorescent microscope. The cytotoxicity of DOX-CSNp and DOX/FOL CSNp were estimated and compared by MTT method.Folate-conjugated chitosan with about 11 folate molecules per chitosan molecule could be prepared. The chitosan nanoparticles prepared by ionically cross-linking method showed an spHeric shape and the particle size was about 100nm under the observation of transmission electron microscopy(TEM).The particle size and the Zeta potential of nanoparticles was 271±3.2nm and 18.74±1.85mv respectively. The average entrapment efficiency and loading capacity of doxorubicin was 30.74% and 6.86% respectively. The cumulative release curve showed that the nanoparticles could achieve sustained- release.The investigation of the interaction of chitosan nanoparticles and HeLa cell showed that there was a greater fluorescence intensity when FITC-labelled, folate-conjugated chitosan nanoparticles were incubated in HeLa cell than FITC-labelled chitosan nanoparticles. Due to the low concentrateion of free folate used in this investingation, the intracellular uptake of folate-conjugated chitosan nanoparticles in HeLa cell couldn't be markedly blocked.The cytotoxicity of DOX-CSNp , DOX/FOL CSNp and free doxorubicin to HeLa cells were performed by MTT method. The results showed that DOX/FOL CSNp had higher cytotoxicity than DOX-CSNp and free doxorubicin when incubated in HeLa cells. As a result, we concluded that DOX/FOL CSNp had better intracellular uptake and stronger cytotoxicity than DOX-CSNp and free doxorubicin to folate receptor positive-expressing tumor cells.