Single-dose vaccine against tick-borne encephalitis.

Tick-borne encephalitis (TBE) virus is the most important human pathogen transmitted by ticks in Eurasia. Inactivated vaccines are available but require multiple doses and frequent boosters to induce and maintain immunity. Thus far, the goal of developing a safe, live attenuated vaccine effective after a single dose has remained elusive. Here we used a replication-defective (single-cycle) flavivirus platform, RepliVax, to generate a safe, single-dose TBE vaccine. Several RepliVax-TBE candidates attenuated by a deletion in the capsid gene were constructed using different flavivirus backbones containing the envelope genes of TBE virus. RepliVax-TBE based on a West Nile virus backbone (RV-WN/TBE) grew more efficiently in helper cells than candidates based on Langat E5, TBE, and yellow fever 17D backbones, and was found to be highly immunogenic and efficacious in mice. Live chimeric yellow fever 17D/TBE, Dengue 2/TBE, and Langat E5/TBE candidates were also constructed but were found to be underattenuated. RV-WN/TBE was demonstrated to be highly immunogenic in Rhesus macaques after a single dose, inducing a significantly more durable humoral immune response compared with three doses of a licensed, adjuvanted human inactivated vaccine. Its immunogenicity was not significantly affected by preexisting immunity against WN. Immunized monkeys were protected from a stringent surrogate challenge. These results support the identification of a single-cycle TBE vaccine with a superior product profile to existing inactivated vaccines, which could lead to improved vaccine coverage and control of the disease.

Characterization of the RepliVax platform for replication-defective flavivirus vaccines.

RepliVax, a novel replication-defective vaccine platform has recently been described as a suitable means of generating potent vaccines targeting flaviviruses. In this study, we directly compared attenuation, immunogenicity and efficacy of several prototype RepliVax constructs to available, well characterized live attenuated (LAV) and inactivated (INV) flavivirus vaccine controls in mice and hamsters. Other important aspects of general mechanisms and properties of RepliVax vaccines were also studied. The prototypes were found to be nonpathogenic in sensitive suckling mouse neurovirulence tests, and highly immunogenic and efficacious in mice and hamsters, with evidence that immunogenicity can be comparable to LAV controls in terms of both magnitude and durability of response. Our data also suggest that choice of inoculation route can be beneficial for maximizing RepliVax immunogenicity. Additionally, different vaccine constructs can be administered as cocktail formulations without compromising immunogenicity of individual components. RepliVax constructs were determined to induce a Th1 biased immune response, similar to LAVs, and different from INV inducing a Th2 type response. The results presented validate the utility of the RepliVax platform for development of novel flavivirus vaccines.

Direct random insertion of an influenza virus immunologic determinant into the NS1 glycoprotein of a vaccine flavivirus.

A live chimeric vaccine virus against Japanese encephalitis (JE), ChimeriVax-JE, was used to define methods for optimal, random insertion of foreign immunologic determinants into flavivirus glycoproteins. The conserved M2e peptide of influenza A virus was randomly inserted into the yellow fever-specific NS1 glycoprotein of ChimeriVax-JE. A technique combining plaque purification with immunostaining yielded a recombinant virus that stably expressed M2e at NS1-236 site. The site was found permissive for other inserts. The insertion inhibited NS1 dimerization in vitro, which had no significant effect on virus replication in vitro and immunogenicity in vivo. Two different NS1-specific monoclonal antibodies and a polyclonal antibody efficiently recognized only the NS1 protein dimer, but not monomer. Adaptation of the virus to Vero cells resulted in two amino acid changes upstream from the insert which restored NS1 dimerization. Immunized mice developed high-titer M2e-specific antibodies predominantly of the IgG2A isotype indicative of a Th1-biased response.

Tau exon 10, whose missplicing causes frontotemporal dementia, is regulated by an intricate interplay of cis elements and trans factors.

Tau is a microtubule-associated protein whose transcript undergoes complex regulated splicing in the mammalian nervous system. In humans, exon 10 of the gene is an alternatively spliced cassette which is adult-specific and which codes for a microtubule binding domain. Mutations that affect splicing of exon 10 have been shown to cause inherited frontotemporal dementia (FTDP). In this study, we reconstituted naturally occurring exon 10 FTDP mutants and classified their effects on its splicing. We also carried out a comprehensive survey of the influence of splicing regulators on exon 10 inclusion and tentatively identified the site of action for several of these factors. Lastly, we identified the domains of regulators SWAP and hnRNPG, which are required for regulation of exon 10 splicing.