Graphene Oxide Derivatives As Carriers For Anti-tumor Drug Or PDNA

Graphene, a two-dimensional nanomaterial with a thickness of 0.35 nm, is composed of sp2-bonded carbons and has attracted considerable interests over the past few years. The solubility of graphene in aqueous solution is limited by the exposed carbon atoms on its surface. Graphene oxide (GO), an oxidative derivative of graphene, could be well dispersed in water owning to the numerous amount of oxygen-containing groups such as carboxyl, carbonyl, hydroxyl and ether. However, GO trends to aggregate in physiological environment due to the electrostatic charges screening and nonspecific proteins adsorption, which requires further surface modification. Owning to the improved biocompatibility and stability, GO derivatives with surface modification have been widely investigated in biomedicine fields including drug delivery. Besides, target ligands have played an important role in GO modification with the regard of facilitating cellular uptake. Trimethyl chitosan (TMC), partially quaternized from biocompatible and biodegradable chitosan, is positively charged and exhibits higher solubility than chitosan in water. Herein, folate (FA) modified TMC (FTMC) was synthesized and utilized to conjugate with GO. Insoluble anticancer drug doxorubicin (DOX) and plasmid DNA (pDNA) were then loaded onto the obtained GO-FTMC, respectively. Cytotoxicity and cellular uptake of GO-FTMC was also investigated.GO was prepared by oxidation of pristine graphite and subsequent sonication. After removal of the large segments by centrifugation, GO suspension was obtained with desired solubility. Its thickness was determined to be 0.838 nm by atomic force microscope (AFM). Dynamic light scattering (DLS) results showed that the particle size and Zeta potential of GO were 87.7 nm and -32.9 mV, respectively. FTMC with good aqueous solubility and positive charge was synthesized by covalent linkage of TMC and FA. The peaks at 7.14,7.76 and 8.82 ppm in 1H nuclear magnetic resonance (’H NMR) spectrum of FTMC were ascribed to FA moiety, confirming the successful linkage of TMC and FA. GO-FTMC was prepared via a facile electrostatic self-assembly process between negatively-charged GO and positively-charged FTMC. The particle size and Zeta potential of GO-FTMC were 112.0 nm and 30.9 mV, respectively. AFM image of GO-FTMC visualized a thickness of 3.038 nm.DOX was loaded onto GO-FTMC and formed GO-FTMC-DOX nanocomplex via π-π staking, hydrophobic interaction and hydrogen bonding. The particle size and Zeta potential of GO-FTMC-DOX nanocomplex were 116.1 nm and 27.0 mV, respectively. The loading capacity and encapsulation efficiency of DOX on GO-FTMC were 30.9% and 44.7%, respectively. In vitro release profile of DOX from DOX-GO-FTMC revealed an initial burst in the first 2 h and a following slower and sustained release later with a 24-h accumulative release percentage of 33.3% at pH 7.4. pDNA was also encapsulated into GO-FTMC at different weight ratios of GO and FTMC. With the weight ratios of FTMC and GO increase from 9:5 to 14:5, GO-FTMC-pDNA nanocomplex possessed similar particle sizes ranging from 120 nm to 150 nm and Zeta potentials increasing from 19 mV to 24 mV. Agarose gel electrophoresis of GO-FTMC-pDNA revealed GO and FTMC could effectively retard the migration of pDNA.Water-soluble tetrazolium salt-8 (WST-8) assay was employed to evaluate the cytotoxicity of GO-FTMC. GO-FTMC with a concentration of 0.01-80 μg/mL showed no toxicity in A549 and Hela cells after 48-h incubation. Rhodamine B (RhB) labled GO-FTMC was used to study the uptake behavior in A549 and Hela cells. Confocal laser scanning microscope (CLSM) showed superior uptake of GO-FTMC in Hela cells as compared with A549 cells which might resulting from the higher amount of FA receptor expressed on Hela cells membrane than A549 cells. The results might be ascribed to the recognition between FA ligands in GO-FTMC and their corresponding receptors on the plasma membrane of Hela cells, indicating that GO-FTMC could be actively internalized through receptor mediated endocytosis.