Preparation And Immunological Studies Of MUC1 Gold Nanoparticle Anti-cancer Vaccine Based On Host-guest Assembling

Cancer seriously threatens human health,and imposes a huge economic burden on individuals and society.In recent years,tumor vaccines,as a safer immunotherapy,have attracted wide attentions.Tumor-associated carbohydrate antigens(TACAs)abundently expressed on the surface of tumor cells,with conserved chemical structure,have been attractive targets for the development of cancer vaccine.Among them,MUC1,which is widely expressed on a series of tumor cells and has truncated glycosylation,is an important tumor marker and common-used in clinical diagnosis and monitoring of treatment response.However,the MUC1 peptide has low immunogenicity and causes immune tolerance that results in difficulties in provoking robust immune responses.In order to enhance the immunogenicity of MUC1,the traditional method is to conjugate MUC1 peptide to protein carrier,immunostimulants,or novel nanoparticle carriers.With the helps from vaccine carriers,the MUC1 peptide can be efficiently taken and treated by antigen-presenting cells and the immune responses can be increased.Gold nanoparticle,as a nano-scale carrier,has been applied in vaccine construct due to its strong immune stimulating ability,controllable size and excellent biocompatibilities.The covalent coupling of antigens on gold nanoparticles generally needs complicated route of chemical synthesis and characterization,which induces the problems in quanlity and safety control.Aim to improve the loading of antigen on gold nanoparticles and simplify vaccine preparation route,this thesis developed novel non-covalent MUC1-gold nanoparticle vaccines based on host-guest assembleing between adamantane and cyclodextrin.The immunological propertises of these vaccines are evaluated as well.The main results are as follows:(1)Three MUC1 peptides with adamantane modification at the N-terminus were successfully synthesized and demonstrated by mass spectrometry.The purity of compounds was higher than 95% according to the analysis by high performance liquid chromatography.The gold nanopariticles(Au NP)with beta-cyclodextrin(β-CD)modification was successfully prepared from chloroauric acid reduced by sodium borohydride.Then,MUC1 peptide with adamantine modification was assembled to Au NP by the host-guest interaction to be MUC1 nanovaccines.(2)The Au NP and the assembled MUC1 nanovaccine were characterized by transmission electron microscopy and thermogravimetric analysis.The particle size of gold nanoparticles is between 3-5 nm,and the loading of β-CD is about 37% of the total mass of nanoparticle.After inclusion,the ratio of β-CD assembled with MUC1 peptides is about70～80% of the total β-CD on the nanoparticles,which is relatively stable.(3)The biocompatibility of Au NP was quite well according to the result of in vitro studies.Comparing with the peptide itself,the nanoparticle formulation can siginificantly promote the uptake efficiency by macrophage.Finally,the MUC1 nanovaccines were demonstrated that can generate robust immune responses represented by the high antibody titers.The elicited antibodies in antisera can successfully recognize tumor cells with MUC1 expression and induce complement-dependent cytotoxicity against tumor cells.