The Evaluation Of Ebola Virus Glycoprotein Subunit Vaccine Immunized By Microneedles

Ebola virus(EBOV)disease has become a great threat to humans across the world in recent years.The 2014 Ebola epidemic in West Africa caused numerous deaths and attracted worldwide attentions.Since no specific drugs and treatments against Ebola virus disease was available,vaccination was considered as the most promising and effective method of controlling this epidemic.Development of a safe and efficacious Ebola virus vaccine is of high importance to public health.The glycoprotein(GP)is the single surface virion structural component of Ebola virus that mediates attachment to and fusion with host cells,so it is the main target of Ebola vaccine study.GP genes encode several forms of GP as producs with unedited and edited transcripts,especially the soluble GP(sGP)which takes up 70% of all products.Currently,vaccinations are commonly administered via intramuscular(IM)injection of vaccines,which are prepared in solution and need to be stored under frozen or refrigerated conditions to maintain their stability.Over the last decade,microneedle(MN)have been under development as a new vaccine delivery technology for intradermal vaccination.The immunization taget skin is the largest immunological organ of the human body.In its dermis layer,it is largely populated by dendritic cells(DCs)ang Langerhans cells(LCs).Additionally,the expression of Toll-like receptors of DCs and LCs,combination with other immunologically active cells residing in the skin including monocytes,and macrophages make skin an ideal target for vaccine delivery.In this study,we focused on Ebola GP as the antigen of subunit vaccine.Predicted the 3-dimensional structure of GP and sGP and compared the same amino acide sequences that shared between these two proteins,the conformation structures are extramly similar which provide the theoretical base that sGP may a promising alternative antigen of GP.Then we constructed the recombinant vaccinia virus system which express EBOV GP and sGP successfully.As subunit vaccine,GP or sGP were immunized and further compared the immune responses induced by MN or IM.Comparison of immune responses induced by MN immunized GP subunit vaccine and the IM administered GP in mice showed that MN delivery of GP induced higher levels and longer lasting antibody responses against GP than IM injection of the same vaccine at the same dose.The antibody level of MN immunized GP is 2-fold higher than IM injection,and the neutralizing antibody is 900 by MN delivery compared to 300 by IM administration;After 16 weeks of second immunization,the antibody level only dropped 30% of MN delivery method which IM adminisered GP dropped 60%.In addition,as the alternative antigen of GP,sGP subunit vaccine also introduced higher and longer lasting antibody induced by MN than IM,but the neutralizing activites are in the same level.Further,we found that EBOV GP in formulation with an adjuvant,Matrix-M,can be efficiently loaded onto MNs for the first time.Co-delivery of the Matrix-M adjuvant with the GP subunit vaccines significantly enhanced induction of antibody responses and neutralizing activity induced by MN delivery,as also observed for IM injection.The IgG level is 5-fold higher than subunit vaccine without adjuvant,and the IgG2 a can be induced only in adjuvant groups which diversified the subtypes of antibody and balanced the ratio of IgG1/IgG2 a.Results from challenge studies showed that all mice that received the adjuvant with GP or sGP formulation by MN or IM immunizations were protected from lethal EBOV challenge,without significant body weight loss and clinical symptoms.Further,4 out of 5 mice that received unadjuvanted GP by MN immunization also survived the challenge,whereas only 1 out of 5 mice that received the unadjuvanted GP by IM immunization survived the challenge.These results demonstrate that MN delivery of EBOV GP subunit vaccines can offer effective protection against EBOV infection,and sGP is a promising alternative antigen of GP with the presence of adjuvant Matrix-M.The study provides new experimental and theoretical basis for development of Ebola vaccine.