Dengue 2 PDK-53 Virus as a Chimeric Carrier for Tetravalent Dengue Vaccine Development

Abstract

Dengue (DEN) viruses, which comprise four serotypes (D1 to D4), are members of the Flavivirus genus and contain a single-stranded positive-sense genomic RNA of approximately 11 kb. The RNA genome organization is 5ЈNCR-C-prM-E-NS1-NS2A-NS2B-NS3-NS4A-NS4B-NS5-3ЈNCR (where NCR is noncoding region, C is capsid, prM is premembrane, E is envelope, and NS is nonstructural protein). The structural proteins (C, prM, and E) and the NS proteins are translated as a single polyprotein precursor which is processed by cellular and viral proteases (37). DEN viruses are the leading cause of mosquito-transmitted viral disease in humans. DEN virus transmission results in an estimated 100 million cases of dengue fever and up to several hundred thousand cases of its more severe form, dengue hemorrhagic fever/dengue shock syndrome (DHF/DSS), every year (16). Both viral virulence and host immune responses have been considered responsible for the pathogenesis of DHF/DSS (38). Immunopathological mechanisms, including antibody-dependent enhancement of virus replication and cell-mediated factors, have been regarded as important factors that contribute to DHF/DSS (31,35,39). It is a general concern that sequential infection with different serotypes of DEN virus will increase the risk of DHF/DSS through such immunopathological mechanisms (19,31). Therefore, an efficacious tetravalent vaccine is needed to provide solid and long-term immunity against all four serotypes of DEN virus.One of the most promising tetravalent DEN virus vaccine candidates consists of live attenuated DEN viruses that were derived by serial passage of wild-type viruses in primary dog kidney (PDK) cells or primary African green monkey kidney cells at Mahidol University, Bangkok, Thailand (45). Human phase I and II clinical trials have been conducted in Thailand and the United States, and the results indicate that these vaccine candidates are safe and immunogenic in humans (3,5,6,14,25,40,41,44). However, some tetravalent formulations failed to induce neutralizing antibodies and/or equivalent Tcell responses against all four serotypes (3, 25, 40, 41). The Mahidol D2 vaccine virus, the PDK-53 strain, was derived by passage of the wild-type D2 16681 virus 53 times in PDK cells. It had the lowest 50% minimum infectious dose (5 PFU) among the four vaccine serotypes for humans (4). When tested alone, PDK-53 virus produced no untoward clinical symptoms, elicited neutralizing antibodies that lasted for at least 2 years, and induced significant memory T-cell responses in humans (4,6,14,44).The uncloned PDK-53 virus vaccine contains a mixture of two genotypic variants (29), designated PDK53-E and PDK53-V in this report. The PDK53-V variant contains all nine PDK-53 virus vaccine-specific nucleotide mutations, including the NS3-250 Glu-to-Val mutation. The PDK53-E variant contains eight of the nine mutations and the NS3-250 Glu of the wild-type 16681 virus. The phenotypic markers associated with the attenuation of PDK-53 virus, including small plaque size and temperature sensitivit...

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

“…Since the first reports of the generation of intratypic (DEN2/DEN4) and intertypic (TBE/ DEN4) antigenic chimeric flaviviruses using reverse genetics [15,22], numerous chimeric viruses have been created using genes from tick-borne and mosquito-borne flaviviruses, and many of these viruses are currently being evaluated as vaccines [10,16,19,20,[23][24][25][26]. Recent clinical studies have indicated that antigenic chimeric flaviviruses are attenuated and immunogenic in human volunteers and may serve as live attenuated virus vaccines for protection against disease caused by DEN, JE, WN, and TBE viruses [27][28][29][30][31].…”

Development of DENVax: A chimeric dengue-2 PDK-53-based tetravalent vaccine for protection against dengue fever

“…It is possible that the PDK-53 virus-specific mutations in prM (DEN-2 PDK-53-V virus only), NS2A, and/or NS4A might further modulate attenuation to an undefined extent. Additionally, the juxtaposition of heterologous genes in the chimeric DEN viruses, particularly in ChiDEN-3-V and -4-V, may also contribute to an attenuated phenotype, relative to wild-type virus [41].…”

“…2, modified from fig. 3 in [41]). No mouse mortality, or evidence of illness requiring euthanasia, occurred in mice injected with these three viruses.…”

“…Wild-type DEN-2, DEN-3, and DEN-4 viruses fail to induce robust anti-viral neutralizing antibody responses after administration to ICR mice [41,44]. Therefore, the immunogenicity of each chimeric virus was tested in transgenic, AG129 knockout mice.…”

“…Attenuated vaccine strains for the DEN-1, -3 and -4 were engineered by replacing the DEN-2 PDK-53 structural genes, premembrane (prM) and envelope (E), with the prM and E genes of wild-type DEN-1, DEN-3 or DEN-4 virus, as shown in Fig. 1 [41]. The origins of the four wild-type DEN virus strains on which the DENVax vaccine is based are shown in Table 2.…”