Rational design of an EDⅢ mRNA vaccine candidate against Zika virus
Zika virus(ZIKV)is an arbovirus belonging to the Flavivirus genus within the Flaviviridae family,which includes other important mosquito-borne human pathogens such as dengue(DENV),West Nile(WNV),Japanese encephalitis(JEV),and yellow fever viruses(YFV).ZIKV was largely overlooked until the first major human epidemic in the Pacific Ocean in 2007,followed by a notable outbreak in French Polynesia in 2013.Due to its association with Guillain-Barré syndrome(GBS)in adults and congenital Zika syndrome(CZS)in infants born to ZIKV-infected mothers,the World Health Organization declared the largest ZIKV outbreak that began in Brazil in 2015 a Public Health Emergency of International Concern on February 1,2016.Currently,there is no approved vaccine for ZIKV.mRNA vaccines,which offer numerous advantages over traditional vaccines in terms of safety,protection,and production efficiency,represent a new and promising platform for vaccine development.In this study,we selected domain Ⅲ(EDⅢ)of the envelope protein(E)of ZIKV FSS13025 strain as the immunogen.A recombinant plasmid containing the EDⅢ-coding region,untranslated regions,and a 120-nt long poly(A)tail was designed and constructed.EDⅢ-mRNA transcripts were produced using the linearized plasmid by in vitro transcription.Abundant EDⅢ antigens were detected in the supernatant of HEK293TN cells transfected with the EDⅢ-mRNA transcript by western blotting and ELISA.The EDⅢ-mRNA transcript was then packaged into lipid nanoparticles(LNPs).Two doses of EDⅢ-mRNA LNPs were administered to BALB/c mice via intramuscular injection using an enhanced immunization schedule at a 3-week interval.ELISA and PRNT analyses revealed that the vaccine induced high levels of dose-dependent E-protein-specific IgG antibodies and neutralizing antibodies against ZIKV,as well as the secretion of Th 1-biased cytokines,such as IFN-γ,TNF-α,and IL-2.Furthermore,the challenge study revealed a significant decrease in viremia and viral RNA loads in the major organs of the mice in the immune groups compared to those in the PBS control group,indicating a substantial immune protective effect in adult mice.Moreover,to further tested the ability of EDⅢ-mRNA immunization to prevent vertical ZIKV transmission during pregnancy,female mice were immunized with EDⅢ-mRNA using the same immunization schedule as described above,followed by mating with male mice.Following the ZIKV challenge,the viremia was significantly reduced in EDⅢ-mRNA-immunized dams compared with PBS-immunized mice.Consistent with this observation,high levels of viral RNA were detected in maternal spleens and brains of PBS-immunized mice,whereas no detectable viral RNA was observed in EDⅢ-mRNA-immunized mice.More importantly,viral RNA was not detectable in maternal brains and fetal heads from EDⅢ-mRNA-immunized dams.These findings suggested that female mice immunized with EDⅢ-mRNA were protected against ZIKV challenge during pregnancy.The mRNA-LNP vaccine encoding EDⅢ of ZIKV designed in this study holds great potential as a new candidate for a ZIKV vaccine.
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