Attenuation and immunogenicity in humans of a live dengue virus type-4 vaccine candidate with a 30 nucleotide deletion in its 3'-untranslated region.

Abstract

Abstract. The recombinant dengue virus type-4 vaccine candidate 2A⌬30 was attenuated in rhesus monkeys due to an engineered 30-nucleotide deletion in the 3Ј-untranslated region of the viral genome. A clinical trial to evaluate the safety and immunogenicity of a single dose of 2A⌬30 was conducted with 20 adult human volunteers. The vaccine candidate was well tolerated and did not cause systemic illness in any of the 20 volunteers. Viremia was detectable in 14 volunteers at a mean level of 1.6 log 10 plaque-forming units/ml of serum, although all 20 volunteers seroconverted with a seven-fold or greater increase in serum neutralizing antibody titer on day 28 post-vaccination (mean titer ϭ 1:580). A mild, asymptomatic, macular rash developed in 10 volunteers, and a transient elevation in the serum level of alanine aminotransferase was noted in five volunteers. The low level of reactogenicity and high degree of immunogenicity of this vaccine candidate warrant its further evaluation and its use to create chimeric vaccine viruses expressing the structural genes of dengue virus types 1, 2, and 3.

Evaluation of St. Louis encephalitis virus/dengue virus type 4 antigenic chimeric viruses in mice and rhesus monkeys

“…Several attenuated flaviviruses have been utilized to generate vaccine candidates including the yellow fever vaccine virus [35], a DEN2 virus that has been attenuated by serial in vitro passage [36], and, in the present study, a DEN4 virus attenuated by a deletion (the Δ30 deletion mutation) in the 3′UTR [37]. The Δ30 deletion mutation in the DEN4 virus has been shown to be highly attenuating and genetically stable in humans and, as such, has been selected for inclusion in antigenic chimeric viruses [38].…”

“…In addition, one or more of these attenuated viruses could be considered for use as a diagnostic reagent at biosafety level 2. (A) The prM/E structural protein region of the DEN4 cDNA plasmid p4 [37] was replaced with the corresponding region from SLE Hubbard to generate the SLE/DEN4 virus. The Δ30 deletion, a 30 nucleotide deletion in the 3′UTR (DEN4 nucleotides 10,478-10,507), was introduced into the SLE/DEN4 cDNA plasmid and used to recover the SLE/DEN4Δ30 virus.…”

Dengue virus type 3 vaccine candidates generated by introduction of deletions in the 3′ untranslated region (3′-UTR) or by exchange of the DENV-3 3′-UTR with that of DENV-4

“…Aliquots of 0.5 ml were removed from flasks daily for seven days. After addition of SPG [22] to a concentration of 1X, samples were frozen on dry ice and stored at −80°C. Virus titer was determined by plaque assay on Vero cells for all samples.…”

“…Viruses were evaluated for replication and immunogenicity in rhesus macaques using established methods [22]. DENV-seronegative monkeys were injected subcutaneously with 10 5 PFU of virus diluted in L-15 medium (Invitrogen) or with a mock inoculum.…”

“…First, reverse genetics has been used to introduce an attenuating 30 nucleotide (nt) deletion (Δ30) mutation into the 3'-UTR of cDNA clones of each DENV serotype [16][17][18][19][20][21]. In initial studies, the rDEN4Δ30 vaccine candidate was found to be attenuated in rhesus monkeys and phase I/II clinical trials in humans have demonstrated that infection with vaccine virus results in low viremia, is strongly immunogenic, and exhibits minimal reactogenicity without serious adverse events [20,22]. Recently, the rDEN1Δ30 vaccine candidate, which was also attenuated in rhesus monkeys, has been found to share a similar set of properties in clinical trials as that observed for rDEN4Δ30: low viremia, strong immunogenicity, and minimal reactogenicity in 20 volunteers [21].…”