Study On A Targeting And Controlled-release Drug Delivery System Based On Mesoporous Silica And The Treatment For Liver Cancer

Mesoporous silica nanoparticles (MSNs) are inorganic solid materials. Owing to their good biocompatibility, good chemical and thermal stability, as well as their ability to be effectively endocytosed by the cells, the application in pharmaceutics received more and more attentions. In this study, we reported the synthesis and multiple functionalization of MSN for bioimaging and controlled drug release. Firstly, MSN with the functionalization of carboxyl (MSN/COOH) was synthesized by co-condensation and was used as the drug carrier for anticancer drugs doxorubicin hydrochloride (DOX). The functionalization with carboxyl caused the pore surface of the nanocarrier more negative than native MSN, which makes the drug-loaded delivery system pH-dependent. The drug delivery system has been applied for intracellular responsive drug delivery, reversal of multi-drug resistance of cancer cells and the reducing of the cardiotoxicity of DOX. Meanwhile, Cy5-hydrazide was reacted with the carboxyl on the surface of MSN/COOH to fabricate fluorescent nanoparticles (MSN/COOH-Cy5) for cell and tumor imaging.In order to enhance the in vivo efficacy of DOX-MSN/COOH further, folate was conjugated to MSN/COOH via functional polyethyleneglycol (PEG), constructing the vector MSN/COOH-PEG-FA. The prepared DOX-MSN/COOH-PEG-FA was able to achieve pH dependent release of DOX and reduce the toxic side effects of DOX. What’s more, compared with DOX-MSN/COOH, DOX-MSN/COOH-PEG-FA could promote tumor targeting delivery of DOX, increase tumor cell uptake, and enhance the potent effects to the in situ liver cancer, thus has a good application prospect.The main content and conclusions are summarized below:Firstly, based on the inorganic material mesoporous silica nanoparticles, carboxy-modified mesoporous silica nanoparticles (MSN/COOH) were synthesized by co-condensation and hydrolysis reaction. Then, Cy5-hydrazide was reacted with the carboxyl group on the surface of MSN/COOH to fabricate fluorescent nanoparticles (MSN/COOH-Cy5) for bioimaging. MSN/COOH samples were regular and spherical, and particle size distribution was narrow, with the average particle size of about50nm. Internal channels arranged uniform, with the pore size of2-3nm. Compared with unmodified MSN, carboxyl modification reduced the the zeta potentials of the nanoparticles, thereby significantly enhanced the dispersibility of the nanoparticles in an aqueous medium. In addition, the connection of Cy5makes the nanocarrier fluorescent, which was suitable for live cell and in vivo bioimaging.The utility of MSN/COOH as an intracellular carrier for anticancer drug delivery system was evaluated using weak base type anti-cancer drug doxorubicin hydrochloride (DOX) as a model anticancer drug. The drug loading and release of DOX under different pH conditions were evaluated. Owing to the charge interactions, DOX loading capacity and encapsulation rate were18.76%and49.48%, respectively. The release of DOX from DOX-MSN/COOH was pH dependent, with a slow release under physiological conditions, and release acceleration under acidic conditions. The pH-sensitive release pattern is conducive for the nanoparticles to play a cytotoxic effect in the tumor.The cellular uptake of blank nanoparticles and drug-loaded nanoparticles in human breast cancer cells (MCF-7) were measured by flow cytometry and the intracellular DOX distribution by DOX-MSN/COOH were observed by confocal laser scanning microscopy. The study showed that the MSN/COOH were able to be uptaken into lysosomes via endocytosis by MCF-7cells, and located in the cytoplasm. The loaded DOX molecules were capable of a quick release from the lysosome and quickly into the nucleus to play a therapeutic effect. The results from MTT assay shows MSN/COOH nanoparticles had no significant toxic effects to MCF-7cells in the range of0-500μg mL"1, while the toxicity of DOX-MSN/COOH to MCF-7cell was equal to that of free DOX, indicating that contained DOX in DOX-MSN/COOH had a rapid intracellular release behaviour and and drug loading process does not affect the efficacy of DOX.The multidrug resistance of cancer cells against chemotherapeutic drugs is a major factor in the failure of chemotherapy. Furthermore, MTT assay was used to compare the free DOX and DOX-MSN/COOH in the cytotoxicity to human breast cancer resistant cells (MCF-7/ADR). The results showed that the drug-loaded nanoparticles could overcome the resistance of MCF-7/ADR to free DOX. According to the results of the cellular uptake study of blank MSN/COOH, we can see the nanoparticles could enter the MCF-7/ADR-cells through the endocytie way, thus-avoided the contact-of the drug with the P-gp in the cell membrane, thereby reducing the efflux and overcome the resistance of MCF-7/ADR cells to free DOX. According to the results of the intracellular distribution study of DOX-MSN/COOH, it can be seen that after DOX-MSN/COOH entering the cells, due to the acidic environment of the cells (especially in the lysosome), the loaded DOX molecules released rapidly in the cytoplasm and quickly went into the nucleus to play the therapeutic effect, so as to further improve the efficacy, slow or overcome the generation of multidrug resistance.Via the use of targeted drug delivery systems, the drugs can be positioned in the diseased tissue, organs or cells, thus improves the in vivo efficacy. On the basis of MSN/COOH, a targeted long-circulating folic acid modified mesoporous silica nanoparticles (MSN/COOH-PEG-FA) was synthesized. MSN/COOH was reacted with NH2-PEG-FA to form the folate modified nanoparticles. After PEG-FA’s modification, there were no obvious change of the nanoparticles with particle sizes, surface charges, pore structures, etc.The cellular uptake behavior of blank MSN/COOH-PEG-FA was observed by confocal laser scanning microscopy (CLSM). Folate receptor highly expressed KB cells were chosen as the model cells. The results confirmed that the cellular uptake of MSN/COOH-PEG-FA was partly due to the FA receptor-mediated endocytosis. Compared with DOX-MSN/COOH-PEG, DOX-MSN/COOH-PEG-FA can promote cellular uptake, and increase the toxic effects of the drug-loaded nanoparticles on KB cells.Furthermore, mice bearing hepatic H22tumor mice bearing liver tumors in situ were used as the model animals to evaluate the antitumor activity of DOX-MSN/COOH-PEG-FA on liver cancer. Compared with free DOX, DOX-MSN/COOH, DOX-MSN/COOH-PEG and DOX-MSN/COOH-PEG-FA all prolonged the survival time of mice bearing liver tumors in situ significantly, and reduced the cardiac side effects of free DOX. This is mainly because, compared with free DOX, DOX loaded nanoparticles increased the uptake by the liver, and maintained a long efficacy time, so that the treatment is better. Compared with DOX-MSN/COOH, PEG-FA’s modification makes the nanoparticles entered the tumor cells by EPR effect and FA receptor-mediated uptake, thus further extended the survival time of tumor-bearing mice and enhanced the anti-tumor efficacy.In summary, this study constructed a new tumor delivery system based on mesoporous silica, and the modified drug delivery system with of PEG and FA, in order to improve the water dispersibility of the carrier and tumor targeting ability. The results showed that the drug carrier was capable of specifically gathering in the liver area, reducing the distribution of DOX in the heart to reduce the side effects of DOX, and extending the survival time of mice bearing liver cancer in situ. The results of this study provided a new way of thinking for the treatment of tumors, and also provided new theoretical basis and experimental evidence for inorganic nanomaterials as targeted drug delivery systems.