| Chemotherapy is one of the most common approaches along with surgery andradiation therapy to treat cancers. It has been proved that chemotherapy can be highlyeffective for many forms of cancer such as testicular cancer, breast cancer andcervical cancer. However, chemotherapy is far from perfect because manychemotherapeutic drugs are toxic to normal cells with a lack of water solubility andtumor selectivity and can cause serious side effects at excessive doses. Therefore, anideal drug delivery system that could improve the drug efficiency and minimize sideeffects is necessary in chemotherapy. In our study, chitosan (CS), a polymer of(1,4)-linked2-amino-2-deoxy-β-glucan, was covalently modifed with folate and thesynthetized folate-chitosan (FA–CS) conjugates were characterized with fouriertransform spectroscopy (FTIR) and1H nuclear magnetic resonance (1H NMR). Thenthe FA-CS conjugates were coated on phosphatidylcholine/cholesterol liposomes todevelop a new tumor targeted drug delivery system——folate-modified-chitosancoated liposomes (FCCLs). The characterization, in vitro drug release and physicalstability of FCCLs were studied and the cellular uptake of the nanoparticles was alsoinvestigated with fluorescein as a model drug.Liposomes are closed spherical vesicles composed of a lipid bilayer and havebeen taken as a promising drug delivery system in recent years because they havegood biocompatibility, sustained-releasing potential and can improve the solubility,stability and bioavailability of many medicinal compounds. They have beeninvestigated for the delivery of various chemotherapeutic agents and severalliposomal drugs have been approved or are under evaluation for clinical use in cancertreatments. However, liposomes have some defects such as a lack of tumor selectivityand physicochemical stability, which greatly restricts their applications in chemotherapeutic drug delivery. They are easy to result in aggregation, fusion,phospholipids hydrolysis and oxidation during storage, which may lead to leakage ofencapsulated drugs. To stabilize the liposomes and avoid the rapid leakage ofentrapped compounds, different types of biocompatible polymers, such as chitosan,can be employed to improve the effciency of conventional liposomal systems.Chitosan (CS), a polymer of (1,4)-linked2-amino-2-deoxy-β-glucan, isattracting increasing attention in drug delivery systems due to its excellent propertiessuch as biocompatibility, biodegradability, nontoxicity, immunogenicity andbioadhesivity. It has been reported that coating liposomes with chitosan can not onlymaintain the advantages of liposomes but also add new desirable properties to thesystem. Chitosan coating on liposomes has been found to increase their stability andto provide them with mucoadehsive properties. Moreover, there are abundantfunctional groups such as hydroxyl and amino groups on the backbone of chitosan, sothat some biologically active molecules (eg, ligand, monoclonal antibody andbiosensor) can be introduced to the chitosan coated liposomes by covalent bonds.It has been proved that the chitosan coated liposomes could be formed via ionicinteraction between positively charged chitosan and negatively chargedphosphatidylcholine on the surface of the liposomes. Besides, hydrophobic interactionbetween polysaccharide and lipid also contributes to the formation of chitosan coatedliposomes. Several authors have used chitosan or related polymers as a liposomecoating in order to increase their stability, to provide them with mucoadhesiveproperties, and for targeting purpose.In this study, N-(3-dimethylaminopropyl)-N-ethylcarbodiimide hydrochloride(EDC) and N-hydroxysuccinimide (NHS) were used to produce folate-chitosan(FA-CS). Then the folate-chitosan conjugates were characterized using1H NMR andinfrared spectrum analysis. In the infrared spectrum of FA-CS, there were bothcharacteristic peaks of chitosan at1646cm1and1514cm1and characteristic peak offolate at1606cm1. Meanwhile, in the1H NMR spectrum of FA-CS, the peak at2.02ppm is attributed to three protons of N-acetyl glucosamine (GlcNAc) and the peaks at8.65ppm and7.69ppm are characteristic signals of the pterin ring and benzene ring of folate. These experiments demonstrated the successful introduction of folateligands to the chitosan backbone. It has also been proved that the substituted positionon chitosan is C2-NH2.The preparation and properties of folate-chitosan coated liposomes (FCCLs) werestudied and compared with those of traditional liposomes and chitosan coatedliposomes (CCLs). FCCLs were spherical in shape with a classic core-shell structure.Compared with conventional liposomes, FCCLs had larger size with an averagediameter of182.0nm while that of liposomes is around161.0nm. After coated withFA-CS layer, the zeta potential of nanoparticles changed from-5.3mV to10.1mV.Compared with traditional liposomes, FCCLs had prolonged drug release behaviors(55.76%after24h) and better physical stability when stored at25℃. After storage at25℃for15days, the average particle size of traditional liposomes changed from161.0nm to2852.3nm due to aggregation and fusion. However, the average particlesize of FCCLs had few changes in the same conditions. These characteristics are allsimilar to the properties of CCLs. These changes may attribute to the FA-CS layercoated on liposomes. The rise in zeta potential of nanoparticles can increase therepelling forces between the nanoparticles and in this way, the stability ofnanoparticles could be improved.With fluorescein as a model drug, fluorescein loaded liposomes, CCLs andFCCLs were prepared and their tumor targeting ability was evaluated according to thein vitro cellular uptake of fluorescein loaded nanoparticles by MCF-7and HUVECcells. Results demonstrated that FCCLs had a signifcant higher uptake by folatereceptor positive cells (MCF-7) as compared to traditional liposomes and CCLs andthis internalization process is a kind of folate receptor mediated endocytosis, whichcould be effectively hampered by free folate. As a result, the FCCLs were promisingnano-carriers for tumor targeted drug delivery. |
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