Glucan Functionalized Glycogen As Drug Carrier And Immune Adjuvant

Glycogen is a kind of natural polymer material with nano structure,good biocompatibility,and easy to functionalize.Deoxyoligonucleotides containing unmethylated cytosine-guanine dinucleotide motifs（Cp G ODNs）can activate innate and adaptive immune responses by interacting with TLR9.However,Cp G ODNs have a skeleton structure composed of phosphodiester bonds which are easily degraded by nucleases in the blood,so they have a short half-life and low stability.Because of electrostatic repulsion,Cp G ODNs,a kind of nucleic acid drug,are difficult to pass through the negatively charged cell membrane.These defects severely limit the clinical application of Cp G ODNs.Therefore,it is key to develop a safe and efficient delivery strategy,in which Cp G ODNs can be targeted to the immune cells and effectively released following interacting with TLR9 in the lysosome.In this paper,a new macrophage targeting Cp G ODNs delivery system（Gly-Glu/Cp G ODNs）was built based on glycogen.The macrophage-targetedβ-glucan was grafted on it through the Schiff base reaction.The Gly-Glu/Cp G ODNs was finally formed by stronger electrostatic interaction between cationic vehicles and electronegative Cp G ODNs.The structure of Gly-Glu was confirmed by infrared spectroscopy,nuclear magnetic resonance spectrum and other methods.Agarose gel electrophoresis showed that Gly-Glu had a strong Cp G ODNs condensation ability and its maximum drug loading achieved the degree of 73μg/mg.Gly-Glu/Cp G ODNs has positive potential（5.1±0.5 m V）and its size is reached to about 180 nm which is conducive to cell uptake and endosomal retention.The results of MTT experiments showed that Gly-Glu/Cp G ODNs has good biocompatibility and no cytotoxic on RAW264.7 cells.The cell uptake behaviors were evaluated by the laser confocal microscope and flow cytometer,which revealed that macrophages showed an efficient and specific uptake capacity for Gly-Glu/Cp G ODNs.Therefore,Gly-Glu/Cp G ODNs showed a potential application prospect in the field of immunotherapy.In this paper,a new macrophage targeting Gly-Glu system was built based on glycogen.OVA is selected as the antigen.The Gly-Glu/OVA was finally formed by stronger electrostatic interaction between cationic vehicles and electronegative OVA.Gly-Glu/OVA has a particle size of about 280 nm.Gly-Glu’s drug loading for OVA is about 165μg·mg^-1.The results of MTT experiments showed that Gly-Glu/OVA has good biocompatibility and no cytotoxic on RAW264.7 cells.The cell uptake behaviors were evaluated by the laser confocal microscope and flow cytometer,which revealed that macrophages showed an efficient and specific uptake capacity for Gly-Glu/OVA.Gly-Glu/OVA had a higher degree of cell surface antibody activation,which promotes The expression of TNF-αand IL-6.Therefore,Gly-Glu showed a potential application prospect in the field of immune adjuvants.