Nano-graphene Oxide For Gene And Drug Delivery

Currently, cancer has become one of the major diseases to threat human health. Commonly used cancer therapeutics, especially chemotherapy, which could restrict the growth of tumors to some degree, often fail to significantly prolong patient survial due to their serious side effects to normal tissues and the development of multidrug resistance(MDR) of cancerous cells. Therefore, a great deal of attention has been paid to the development of novel cancer therapeutics which are potentially effective in targeted drug delivery and overcoming MDR. In this doctoral dissertation, by using nano-graphene oxide(NGO) as a core nanomaterial, we have fabricated an effective gene delivery carrier with low cytotoxicity and high transfection efficiency without the interference of serum after screening a series of surface functionalization strategies. Afterwards, utilizing the strong photothermal conversion ability of NGO in near infrared(NIR) region, we have realized a photothermally enhanced gene transfection. By choosing polo-like kinase 1(Plk1) as the therapeutic target, we have found that our fabricated NGO based gene delivery carrier could effectively deliver siRNA(siPlk1) into cells and down-regulate the expression of Plk1. On the other hand, we have designed and prepared an NGO based drug delivery carrier with dual pH responsiveness to the acidic environments of tumor extracellular matrix(ECM) and intracellular endosomes & lysosomes. Utilizing such pH responsive NGO based drug delivery carrier, we have successfully overcome the MDR by using the combined chemo- and photothermal therapy(PTT). This doctoral study presents the fabrication of various novel gene and drug delivery carriers, aiming at overcoming of MDR and improving the cancer therapeutic effect.The main contents and conclusions of this dissertation are summarized as follows:1. In the first study, we non-covalently modified the negatively charged NGO with positively charged polyethylenimine(PEI) via the electrostatic interaction, obtaining a positively charged NGO/PEI complex and explored its capacity for gene delivery. Our results indicated that PEI could be effectively adsorbed onto the surface of NGO via the electrostatic interaction in a layer-by-layer(LBL) manner. Besides, the PEI modification could greatly enhance the physiological stability of NGO and reduce its cytotoxicity in comparison with bare PEI polymers. Gel electrophoresis retardation assay showed that as-prepared NGO/PEI complexes could efficiently bind with plasmid DNA. The ultimate gene transfection assay demonstrated that as-prepared NGO/PEI complexes with lower cytotoxicity to cells, showed a comparable transfection efficiency to bare PEI molecules if not better.2. As a follow-up study, we covalently conjugated NGO with polyethylene glycol(PEG) and PEI to improve its stability and gene transfection efficiency and reduce its cytotoxicity in serum containing conditions. It was found that the obtained NGO-PEG-PEI exhibited a greatly improved physiological stability and gene transfection efficiency in serum-containing conditions compared with bare PEI molecule and GO-PEI without PEGylation. By utilizing the high photothermal conversion ability of NGO, we found that a 20-min NIR laser irradiation at a low power density could effectively induce mild photothermal heating and promote the cellular uptake of NGO-PEG-PEI, realizing a dramatically increased plasmid DNA expression and RNA interfering effect. With such superior photothermally ehanced gene delivery ability, our fabricated NGO-PEG-PEI demonstrated to be a promising candidate for combined gene and photothermal therapies.3. Considering the different pH values of normal and tumoral tissues, we designed an NGO-based drug delivery carrier with dual pH responsiveness to the different acidic conditions of tumoral ECMs and intracellular endosome & lysosome to overcome MDR. Firstly, we conjugated NGO with PEG and cationic poly(allylamine hydrochloride)(PAH), obtaining a positively charged NGO-PEG-PAH conjugate of great physiological stability. Then, we modified the NGO-PEG-PAH with 2,3-dimethylmaleic anhydride(DA) to prepare a negatively charged NGO-PEG-DA, which was stable in normal pH value(e.g. pH 7.4) while could be rapidly reversed to be positively charged when incubated in a mild acidic condition(e.g. pH 6.8). Utilizing the presence of hydrophobic region on the surface of NGO, doxorubicin(DOX), a widely used anticancer drug, could be effectively loaded onto the surface of NGO-PEG-DA, obtaining NGO-PEG-DA/DOX complexes. As-prepared NGO-PEG-DA/DOX exhibited a pH-dependent drug release profile, cellular uptake and cell killing ability, and was found to be efficient in overcoming MDR by combining the superior photothermal conversion ability of NGO.In this doctoral dissertation, we systemically studied the applicable potentials of GO for controllable drug & gene delivery. We found that the excellent photothermal conversion ability of GO could be exploited to enhance the gene transfection efficiency of GO based gene delivery carrier and to improve its ability in overcoming the MDR, comfirming its promising advantages in cancer treatment via combined PTT and gene therapy or chemotherapy. This doctoral dissertation presents a promising strategy for the development of novel cancer treatment methods, and some references for the design of various smart photothermal or tumor microenvironmental responsive gene & drug delivery carriers.