Studies On Preparation And Effect Of Self-assembled Nanovaccine

Vaccines can activate the body’s immune system to induce antigen-specific immune responses,and have been widely applied to the resistance of virus,bacteria and tumor.The development of vaccines has gone through several stages such as pathogen,protein and synthetic peptide.Compared with inactivated or attenuated vaccines,the birth of protein and peptide vaccines extremely enhances vaccine safety;however,it is difficult to effectively provoke the body’s specific immune response for their low immunogenicity.To improve the immune responses of vaccines is an important scientific issue for enhancing the effectiveness of disease treatment.With the rapid development of novel vaccines,adjuvant has become an vital component of vaccine,and is injected simultaneously or in advance with antigen to enhance or change the body’s immune responses to antigen.According to different mechanisms of action,adjuvants can be classified into two categories:immunomodulatory molecules and delivery systems,which can enhance antigen immunogenicity and immune response by means of enhancing the in vivo residence time of antigens,promoting antigen presentation and activating lymphocyte.Adjuvants are used to enhance the immune responses of immunocompromised vaccine systems,accelerate the generation of long-lasting immune responses,cut down cost,reduce antigen dosage,promote antigen-specific antibody responses and cellular immune responses,and induce different types of immune responses.In this study,appropriate adjuvants were selected to realize the self-assembly of antigens and adjuvants into nanovaccines,and to improve vaccine immune responses.(1)Porcine Deltacoronavirus(PDCoV)was introduced into motherland China in2014,and there is still no effective vaccine so far.In the first work,two adjuvants with different mechanisms of action such as CpG and Nanoemulsion(0.5%w/v Span 85,0.5%w/v Tween 80 and 5%v/v squalene),were used in the research and development of PDCoV inactivated vaccines.The results indicated that Nanoemulsion can load inactivated PDCoV antigen components and form nanoparticle(157.6±0.4 nm)with uniform size,compared with the irregularly mixing of CpG and inactivated PDCoV.The results of bone marrow-derived dendritic cell(BMDC)in vitro activation experiment indicated that both of CpG/PDCoV and Nanoemulsion/PDCoV inactivated vaccines could further promote the maturation and activation of BMDCs,compared with the sole inactivated PDCoV.The mouse subcutaneous immunization experiments had shown that both of CpG/PDCoV and Nanoemulsion/PDCoV inactivated vaccines evoked effective antibody responses and cellular immune responses,but the latter was able to induce stronger humoral and cellular immunity.Therefore,Nanoemulsion adjuvant with low cost and good effects is a good choice for the preparation of efficient PDCoV inactivated vaccines.However,the prophylactic and therapeutic effects of Nanoemulsion/PDCoV inactivated vaccine need to be further validated in pigs.(2)Epitopes are the minimal immunogenic units of protein antigens,and can directly combine with major histocompatibility complex(MHC)and then present epitope information to T lymphocytes.Compared with whole protein vaccine,epitope vaccine can cut down or even leave out the protein processing,extremely enhancing the efficiency of antigen epitope presentation.However,the results of many clinical trials indicated that compared with protein vaccines,attenuated vaccines and inactivated vaccines,the antitumor immune responses induced by peptide vaccines are weaker for poor immunogencity and easy degradation of peptides.In this study,three CD8 T cell antigen epitopes such as SIINFEKL,SVYDFFVWL and LCPGNKYEM,were modified by Arginine octapeptide(denoted by R8)into cationic epitopes(epitope-R8s),endowing peptides good water solubility and promoting the uptake and cross-presentation of antigenic epitopes.Epitope-R8s could assemble with TLR9 agonist CpG into nanoparticles through electrostatic interaction,which could overcome the disadvantages of epitopes,including low immunogenicity and esay degradation in vivo.In this experiment,the SII-R8/CpG nanovaccine that was self-assembled by SIINFEKL-R8(SII-R8)and CpG,was taken as an example to study the body’s immune responses and the application prospects in tumor immune prevention and treatment.The experimental results indicated that SII-R8/CpG nanovaccine effectively facilitated the migration of SII-R8 to the draining lymph nodes and induced SII-specific memory immune response,generating the effective immune preventive effect of melanoma.In tumor therapeutic model,compared with the direct mixed dosage form of SII and CpG,SII-R8/CpG nanovaccine significantly promoted tumor filtration of CD8~+T cells,and facilitated cytotoxic T lymphocytes to kill tumor cells,thereby inhibiting tumor growth.In addition,the combination of SII-R8/CpG nanovaccine and anti-PD1 antibody produced the“1+1>2”anti-tumor immune effect.Epitope peptides cationization and its assembly with anionic adjuvant CpG is an universal,convenient,efficient and low-cost preparation method of tumor nanovaccine,holding good prospects in cancer immunotherapy.In summary,the construction of self-assembled nanovaccines was an effective method to improve antigen-specific immune response,having potential application prospects in veterinary virus vaccines and cancer immunotherapy.