Zwitterionic Polymers-based Nanoparticles For The Delivery Of Nucleic Acid Drug And Their Immulogical Properties Investigation

Nucleic acid drugs show promising application for the prevention and the treatment of various diseases.However,defects due to the intrinsic difficiencies of themselves hinder its clinical applications greatly.Nanoparticles attract much attention in biomedical field in recent years due to their excellent physicochemical properties.Although numerous research have reported the usage of nanosystem for nucleic acid drugs delivery(such as plasmid DNA,small interfering RNA(siRNA))for the prevention/treatment of HIV,cancer and so on,great deficiencies remain,such as low loading efficiency,lack of targeted delivery,low intracellular uptake and low endosomal/lysosome escape efficiency.Hence,new nanosystem with specific characters should be established:(I)high loading efficiency of nuclear acid drug;(II)selectively targeting ability;(III)efficient intracellular uptake and followed endosomal/lysosomal escape avoiding being pumped out of cells;(IV)adjunvant effect once nuclear acid drugs function as antigens.Zwitterionic polymer PCB based nanosystem shown excellent nuclear acid loading efficiency and target delivery capability and promoted efficiency of treatment.Hence,PCB based liposomal nanosystem were designed and developed in this dissertation,which were applied in two major fileds(a)HIV prevention and(b)cancer treatment.The proposed nanosystem could accumulate in target site efficiently and delivery cargoes into target cells,and finally enhance efficiency of drugs.In detail,these thesis mainly included the following issues:(1)Human immunodeficiency virus(HIV)DNA vaccine can induce cellular and humoral immunity.A safe and effective HIV DNA vaccine is urgent need to prevent the spread of acquired immune deficiency syndrome(AIDS).The major drawback of DNA vaccines is the low immunogenicity,which is caused by the poor delivery to antigen presenting cells and insufficient antigen expression.Sparked by the capability of endosomal/lysosomal escape of the zwitterionic lipid distearoyl phosphoethanol-aminepolycarboxybetaine(DSPE-PCB),we attempted to develop a zwitterionic-based cationic liposome with enhanced immunogenicity of DNA vaccines.The mannosylated zwitterionic-based cationic liposome(man-ZCL)was constructed as a DNA vaccine adjuvant for HIV vaccination.Man-ZCL could complex with DNA antigens to forma tight structure and protect them from nuclei enzyme degradation.Benefited from the capability of the specific mannose receptor mediated antigen processing cells targeting and enhanced endosomal/lysosomal escape,the man-ZCL lipoplexes were supposed to promote antigen presentation and the immunogenicity of DNA vaccines.In vitro and in vivo results revealed that man-ZCL lipoplexes showed enhanced anti-HIV immune responses and lower toxicity compared with CpG/DNA and Lipo2k/DNA,and triggered a Th1/Th2 mixed immunity.An antigen-depot effect was observed in the administration site,and this resulted in enhanced retention of DNA antigens in draining lymph nodes.Most importantly,the man-ZCL could assist to activate T cells through a non-inflammasome pathway.These findings suggested that the man-ZCL could be potentially applied as a safe and efficient DNA adjuvant for HIV vaccines.(2)Understanding the effects of magnetic iron nanoparticles(MINPs)on the immune response is vitally important for biomedical applications such as cancer therapy,disease diagnosis and novel cancer imaging.In this study,lipid modified MINPs were designed and prepared by introducing the neutral lipid DSPE-PEG or the zwitterionic lipid DSPE-PCB into hydrophobic MINPs through hydrophobic interaction(L-MINPs and ZL-MINPs,respectively).The effect of L-MINPs and ZL-MINPs on the intracellular accumulation and immune responses of three kinds of antigen processing cells was examined.The results indicated that the high cellular uptake efficiency of surface coated MINPs was strongly related to the nature of the coating lipid,with the zwitterionic lipid being more effective than PEGylated ones.Besides,the results from flow cytometry(FCM),confocal laser scanning microscopy(CLSM)and Prussian blue staining demonstrated a time-and concentration-dependent MINP internalization.The uptake of zwitterionic lipid modified MINPs(ZL-MINPs)induced very low cytotoxicity and a strong mixed Th1/Th2 type immune response.L-MINPs could induce a strong increase in pro-inflammatory cytokines with a slight secretion of Th2 cytokines.Besides,no IL-10 was observed in both groups,indicating that MINPs with lipid modification were absence of immunosuppression.In conclusion,this study addresses an important implication of the lipid type and Fe concentration on the immune stimulation of cells and supports the potential for further development of biomedical applications.(3)The chemotherapy of glioblastoma is severely hindered by the immunosuppressive tumor microenvironment,especially the tumor growth factor β(TGF-β),an immunesuppressive cytokine.In this study,we proposed to employ RNAi-based immunomodulation to modify the tumor immune microenvironment and improve the effect of chemotherapy.Herein,a nanotheranostic system(Angiopep-LipoPCB(Temozolomide+BA-PDMAEA/SPIONs@siTGF-β),ALBTA)with dual-targeting and ROS response was established for intracranial glioblastoma treatment.The traceable nanoparticles exhibited strong siRNA condensation,high drug loading efficiency,good serum stability and magnetic property.They could efficiently cross blood brain barrier(BBB)and target to glioblastoma cells via receptor-mediated transcytosis(RMT).The zwitterionic lipid(distearoyl phosphoethanol-amine-polycarboxybetaine lipid,DSPE-PCB)in ALBTA promoted endosomal/lysosomal escape,and thus enhanced the cytotoxicity of temozolomide(TMZ)and improved gene silencing efficiency of siTGF-β.ALBTA could significantly improve the immunosuppressive microenvironment and prolong the survival time of glioma-bearing mice.Moreover,ALBTA could be accurately traced by MRI in brain tumors.The study indicated that this immunochemotherapeutic platform could serve as a flexible and powerful synergistic system for treatment with brain tumors as well as other brain diseases in central nervous system(CNS).