Examination of the envelope glycoprotein of yellow fever vaccine viruses with monoclonal antibodies

Two panels of envelope glycoprotein reactive monoclonal antibodies (mAbs) were prepared against yellow fever (YF) 17D vaccine viruses. Five mAbs were prepared against the World Health Organization 17D-204 avian leukosis virus-free secondary seed virus and eight mAbs against 17DD vaccine manufactured in Brazil. The majority of these mAbs were type-specific and displayed differing reactions in neutralization tests. One, B14, would only neutralize YF vaccine virus grown in invertebrate cells. Others would differentiate 17D-204 and 17DD vaccines, from different manufacturers, in neutralization tests when the viruses were grown in vertebrate cells. The data indicate that heterogeneity exists between the epitopes that elicit neutralizing antibody on YF vaccine from different manufacturers.     

Development of a recombinant,chimeric tetravalent dengue vaccine candidate

Dengue is a significant threat to public health worldwide. Currently, there are no licensed vaccines available for dengue. Takeda Vaccines Inc. is developing a live, attenuated tetravalent dengue vaccine candidate (TDV) that consists of an attenuated DENV-2 strain (TDV-2) and three chimeric viruses containing the prM and E protein genes of DENV-1, -3 and -4 expressed in the context of the attenuated TDV-2 genome backbone (TDV-1, TDV-3, and TDV-4, respectively). TDV has been shown to be immunogenic and efficacious in nonclinical animal models. In interferon-receptor deficient mice, the vaccine induces humoral neutralizing antibody responses and cellular immune responses that are sufficient to protect from lethal challenge with DENV-1, DENV-2 or DENV-4. In non-human primates, administration of TDV induces innate immune responses as well as long lasting antibody and cellular immunity. In Phase 1 clinical trials, the safety and immunogenicity of two different formulations were assessed after intradermal or subcutaneous administration to healthy, flavivirus-naïve adults. TDV administration was generally well-tolerated independent of dose and route. The vaccine induced neutralizing antibody responses to all four DENV serotypes: after a single administration of the higher formulation, 24–67%% of the subjects seroconverted to all four DENV and >80% seroconverted to three or more viruses. In addition, TDV induced CD8⁺ T cell responses to the non-structural NS1, NS3 and NS5 proteins of DENV. TDV has been also shown to be generally well tolerated and immunogenic in a Phase 2 clinical trial in dengue endemic countries in adults and children as young as 18 months. Additional clinical studies are ongoing in preparation for a Phase 3 safety and efficacy study.     

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

Dengue. virus infection is the leading arboviral cause of disease worldwide. A vaccine is being developed based on the attenuated DEN-2 virus, DEN-2 PDK-53. In this review, we summarize the characteristics of the parent DEN-2 PDK-53 strain as well as the chimeric viruses containing the prM and E genes of DEN-1, DEN-3 or DEN-4 virus in the genetic backbone of the DEN-2 PDK-53 virus (termed DENVax). Tetravalent DENVax formulations containing cloned, fully sequenced isolates of the DEN-2 PDK-53 virus and the three chimeras have been evaluated for safety and efficacy in preclinical animal models. Based on the safety, immunogenicity and efficacy in preclinical studies, Phase 1 clinical testing of DENVax has been initiated.     

Aedes aegypti in Brazil: genetically differentiated populations with high susceptibility to dengue and yellow fever viruses

Aedes aegypti was eliminated from Brazil in 1955, but re-infested the country in the 1970s. Dengue outbreaks have occurred since 1981 and became endemic in several cities in Brazil after 1986. Urban yellow fever has not occurred since 1942, and only jungle yellow fever cases have been reported. A population genetic analysis using isoenzyme variation combined with an evaluation of susceptibility to both yellow fever and dengue 2 viruses was conducted among 23 A. aegypti samples from 13 Brazilian states. We demonstrated that experimental infection rates of A. aegypti for both dengue and yellow fever viruses (YFV) are high and heterogeneous, and samples collected in the endemic and transition areas of sylvatic yellow fever were highly susceptible to yellow fever virus. Boa Vista, a border city between Brazil and Venezuela, and Rio de Janeiro in the Southeast region are considered as the most important entry points for dengue dissemination. Considering the high densities of A. aegypti, and its high susceptibility to dengue and yellow fever viruses, the risk of dengue epidemics and yellow fever urbanization in Brazil is more real than ever.     

Persistence of yellow fever vaccine RNA in urine

To evaluate possible persistence of 17D yellow fever vaccine, we tested urine samples from 44 healthy recipients of yellow fever vaccine at varying times up to one year after vaccination. Urine samples from two vaccine recipients had detectable yellow fever virus RNA. The time since vaccination was reported as 21 days for one sample and 198 days for the other sample. These results suggest that yellow fever vaccine virus might persist for at least 6 months after vaccination in some people.     

Live virus vaccines based on a yellow fever vaccine backbone: Standardized template with key considerations for a risk/benefit assessment

The Brighton Collaboration Viral Vector Vaccines Safety Working Group (V3SWG) was formed to evaluate the safety of live, recombinant viral vaccines incorporating genes from heterologous viruses inserted into the backbone of another virus (so-called “chimeric virus vaccines”). Many viral vector vaccines are in advanced clinical trials. The first such vaccine to be approved for marketing (to date in Australia, Thailand, Malaysia, and the Philippines) is a vaccine against the flavivirus, Japanese encephalitis (JE), which employs a licensed vaccine (yellow fever 17D) as a vector. In this vaccine, two envelope proteins (prM-E) of YF 17D virus were exchanged for the corresponding genes of JE virus, with additional attenuating mutations incorporated into the JE gene inserts. Similar vaccines have been constructed by inserting prM-E genes of dengue and West Nile into YF 17D virus and are in late stage clinical studies. The dengue vaccine is, however, more complex in that it requires a mixture of four live vectors each expressing one of the four dengue serotypes. This vaccine has been evaluated in multiple clinical trials. No significant safety concerns have been found. The Phase 3 trials met their endpoints in terms of overall reduction of confirmed dengue fever, and, most importantly a significant reduction in severe dengue and hospitalization due to dengue. However, based on results that have been published so far, efficacy in preventing serotype 2 infection is less than that for the other three serotypes. In the development of these chimeric vaccines, an important series of comparative studies of safety and efficacy were made using the parental YF 17D vaccine virus as a benchmark. In this paper, we use a standardized template describing the key characteristics of the novel flavivirus vaccine vectors, in comparison to the parental YF 17D vaccine. The template facilitates scientific discourse among key stakeholders by increasing the transparency and comparability of information. The Brighton Collaboration V3SWG template may also be useful as a guide to the evaluation of other recombinant viral vector vaccines.     

High stability of yellow fever 17D-204 vaccine: a 12-year restrospective analysis of large-scale production

We have retrospectively analyzed 12 bulk lots of yellow fever vaccine Stamaril, produced between 1990 and 2002 and prepared from the same seed lot that has been in continuous use since 1990. All vaccine batches displayed identical genome sequence. Only four nucleotide substitutions were observed, compared to previously published sequence, with no incidence at amino-acid level. Fine analysis of viral plaque size distribution was used as an additional marker for genetic stability and demonstrated a remarkable homogeneity of the viral population. The total virus load, measured by qRT-PCR, was also homogeneous pointing out reproducibility of the vaccine production process. Mice inoculated intracerebrally with the different bulks exhibited a similar average survival time, and ratio between in vitro potency and mouse LD(50) titers remained constant from batch-to-batch. Taken together, these data demonstrate the genetic stability of the strain at mass production level over a period of 12 years and reinforce the generally admitted idea of the safety of YF17D-based vaccines.     

Pediatric measles vaccine expressing a dengue tetravalent antigen elicits neutralizing antibodies against all four dengue viruses

Dengue disease is an increasing global health problem that threatens one-third of the world's population. To control this emerging arbovirus, an efficient preventive vaccine is still needed. Because four serotypes of dengue virus (DV) coexist and antibody-dependent enhanced infection may occur, most strategies developed so far rely on the administration of tetravalent formulations of four live attenuated or chimeric viruses. Here, we evaluated a new strategy based on the expression of a single minimal tetravalent DV antigen by a single replicating viral vector derived from pediatric live-attenuated measles vaccine (MV). We generated a recombinant MV vector expressing a DV construct composed of the four envelope domain III (EDIII) from the four DV serotypes fused with the ectodomain of the membrane protein (ectoM). After two injections in mice susceptible to MV infection, the recombinant vector induced neutralizing antibodies against the four serotypes of dengue virus. When immunized mice were further inoculated with live DV from each serotype, a strong memory neutralizing response was raised against all four serotypes. A combined measles-dengue vaccine might be attractive to immunize infants against both diseases where they co-exist.     

Construction and biological properties of yellow fever 17D/dengue type 1 recombinant virus

Dengue virus, a mosquito-borne flavivirus, is one of the most formidable public health threats in tropical and subtropical regions. As yet, there is no licensed vaccine to protect against the disease. A chimeric yellow fever (YF) 17D/dengue (DEN) type 1 virus was constructed by replacing the pre-membrane and envelope genes of YF 17D virus with those from DEN 1 VeMir95 virus, a Venezuelan isolate. The chimeric YF 17D/DEN 1 VeMir95 virus was regenerated from full-length infectious clones stably propagated in Escherichia coli by transfection of Vero cells with in vitro transcribed RNA. The chimeric virus proliferated efficiently in Vero cells ( approximately 6.6 log(10) plaque-forming units/ml). The chimeric virus was not neurovirulent to 3-week-old Swiss Webster mice inoculated by the intracerebral route, in contrast to the YF 17DD vaccine strain that was lethal for 90% of the mice. The YF 17D/DEN 1 virus at Passage 6 was more attenuated for rhesus monkeys than the YF 17DD commercial vaccine after intracerebral inoculation according to the standard neurovirulence test. This virus is a potential candidate to be included in a tetravalent DEN vaccine formulation. The availability of the cloned cDNA allows further structure/function studies on the viral envelope.     

Characterization of a viscerotropic yellow fever vaccine variant from a patient in Brazil

Although the live attenuated yellow fever (YF) 17D vaccine is considered to be one of the safest vaccines in the world today, several cases of disease associated with administration of the vaccine have been reported, including YF vaccine-associated viscerotropic disease (YF-VAVD), which was first described in 1996. All YF-VAVD isolates sequenced to date have shown very little genomic change when compared to their parental vaccine strains. In this study, we report the characterization of an isolate, BeH291597 (Brazil75), from a 1975 fatal case of YF-VAVD in Brazil. Comparison of Brazil75 with the genomic sequence of the parental 17DD vaccine strain revealed two amino acid substitutions (at positions M-49 and NS4B-240) that were unique to Brazil75. Although still a rare occurrence, this isolate suggests that YF-VAVD has been present much longer than previously recognized.     

Comparative clinical study of a new 17D thermostable yellow fever vaccine

The immunogenicity and the safety of a new heat-stable 17D yellow fever vaccine have been assessed in a randomized comparative study by reference to a non-stabilized vaccine preparation. Seronegative adults were used and 115 and 143 were given the heat-stable and the non-stabilized vaccine, respectively. Fifty two days after the immunization, haemagglutination inhibiting antibodies were found in 77.6 and 73.9% of the vaccinees, neutralizing antibodies in 99.3 and 100% of them. The percentage of seroconversion and the geometric mean of antibody titres were not significantly different. No complaints or adverse reactions in association with the vaccines were recorded. This study demonstrates the high immunogenicity of this new stabilized vaccine whose stability has already been proven.