Defining levels of dengue virus serotype-specific neutralizing antibodies induced by a live attenuated tetravalent dengue vaccine (TAK-003)

Abstract

The four dengue virus serotypes (DENV1-4) infect several hundred million people each year living in tropical and sub-tropical regions. Clinical development of DENV vaccines is difficult because immunity to a single serotype increases risk of severe disease during a second infection with a new serotype. Leading vaccines are based on tetravalent formulations to induce simultaneous and balanced protective immunity to all 4 serotypes. TAK-003 is a tetravalent live attenuated dengue vaccine candidate developed by Takeda Vaccines Inc, which is currently being evaluated in phase 3 efficacy trials. Here, we use antibody depletion methods and chimeric, epitope transplant DENVs to characterize the specificity of neutralizing antibodies in dengue-naïve adults and non-human primates immunized with TAK-003. Our results demonstrate that TAK-003 induced high levels of DENV2 neutralizing antibodies that recognized unique (type-specific) epitopes on DENV2. In contrast, most vaccinated subjects developed lower levels of DENV1, DENV3 and DENV4 neutralizing antibodies that mainly targeted epitopes that were conserved (cross-reactive) between serotypes. Trial Registration: ClinicalTrials.gov NCT02425098.

Conflict of interest statement

I have read the journal’s policy and the authors of this manuscript have the following competing interests: RB and AdS have served as consultants for dengue vaccine developers and they are inventors on patents filed by the University of North Carolina at Chapel Hill related to dengue vaccines.

Figures

Fig 1

50% Neutralization titers (Neut50) in undepleted (A) and Heterologous virus-depleted (B) baseline seronegative NHP vaccinated with TDV (left) or infected with WT DV1, DV2 or DV3 (right). Non-human primates were vaccinated with one dose (filled circles) or two doses (half filled circles) of TDV, or infected once with wild type DV1, DV2 or DV3 (S2 Table). Animals were bled 180 days after infection or last vaccination. Each sample underwent BSA-depletion (undepleted control) (A), depletion of cross-reactive nAbs by incubation with heterologous serotype viruses (HET depleted) (B) and depletion with the homologous serotype, before determining Neut50 titers by mFRNT assay. Each value represents the titer after subtraction of the background homologous-depletion titer. Each dot represents one animal. Line represents the GMT of the Neut50 titers. The lowest serum dilution tested was 20 (horizontal dotted line). Statistical analysis was done using nonparametric Friedman test with Dunn’s multiple comparison among serotypes after TDV vaccine. * P < 0.05; ** P < 0,005; *** P < 0.0005.

Fig 2

50% Neutralization titers (Neut50) in undepleted (A) and Heterologous virus-depleted (B) baseline seronegative subjects vaccinated with one dose of TDV (left) or after exposure to primary DV1 or DV2 infection (right). The TDV vaccine cohort included sera from 30 subjects collected 180 days after TDV vaccination. The primary infection cohort included 6 primary DENV1 and 8 primary DENV2, collected at baseline in study DEN-205. Each sample was BSA-depleted (Undepleted control) (A) or depleted of cross-reactive nAbs using heterologous serotype viruses (HET Depleted) (B). To measure DV1 and DV3 TS neutralization, HET depletion was done with a mix of DV2+DV4 dynabeads. For DV2 and DV4 TS titers, Het depletion was done with a mix of DV1+DV3 dynabeads. Each value represents the titer after subtraction of background homologous depletion titer. Each dot represents one subjects. Line represents the GMT of the Neut50 titers. The # of subjects with evidence of TS nAbs >20 after HET depletion/#total subjects analyzed is shown at the bottom of panel B. % TS is the percent of subjects with TS nAbs = or > 20. The lowest serum dilution tested was 20 (horizontal dotted line). Statistical analysis was done using nonparametric Friedman test for multiple comparison among serotypes after TDV vaccine. ** P < 0,005; **** P < 0.0001.

Fig 3. Analysis of the levels of TS nAbs in a subset of DEN-205 clinical samples that met enhanced inclusion criteria for TS analysis.

Samples were included in this analysis only if the efficiency of Ab depletion for the serotype used as heterologous was >90%, or if the HET-depleted titer was

Fig 4

Tracking the molecular specificity of DV2 TS nAbs to epitopes on EDIII in sera from TDV vaccinated NHPs (A), WT DV2 infected NHPs (B), TDV vaccinated, baseline seronegative adults (C) or adults who had a DV2 natural infection (D). Neut50 titers to parental viruses DV2 (EDIII epitope donor), DV4 (backbone recipient of epitope transplant) and chimeric virus DV4/2EDIII (DV2 EDIII transplanted into DV4 E protein) were determined in sera collected 180 days after infection or vaccination (A, B, C). Sera used in (D) were collected in subjects pre-TDV vaccination, with evidence of past primary DENV2 infection based on Neut50 titers to DV2 > 4-fold higher than those to the other 3 serotypes. (A) (B) and (D) show titers in control-depleted (BSA-D) and DV4-depleted sera. (C) shows titers in undepleted sera. Horizontal lines and error bars represent GMT +/- 95% CI. Dotted line represents lower limit of detection.