Plasmablast Expansion Following the Tetravalent, Live-Attenuated Dengue Vaccine Butantan-DV in DENV-Naïve and DENV-Exposed Individuals in a Brazilian Cohort

Cássia G T Silveira, Diogo M Magnani, Priscilla R Costa, Vivian I Avelino-Silva, Michael J Ricciardi, Maria do Carmo S T Timenetsky, Raphaella Goulart, Carolina A Correia, Mariana P Marmorato, Lilian Ferrari, Zelinda B Nakagawa, Claudia Tomiyama, Helena Tomiyama, Jorge Kalil, Ricardo Palacios, Alexander R Precioso, David I Watkins, Esper G Kallás, Cássia G T Silveira, Diogo M Magnani, Priscilla R Costa, Vivian I Avelino-Silva, Michael J Ricciardi, Maria do Carmo S T Timenetsky, Raphaella Goulart, Carolina A Correia, Mariana P Marmorato, Lilian Ferrari, Zelinda B Nakagawa, Claudia Tomiyama, Helena Tomiyama, Jorge Kalil, Ricardo Palacios, Alexander R Precioso, David I Watkins, Esper G Kallás

Abstract

An effective vaccine against the dengue virus (DENV) should induce a balanced, long-lasting antibody (Ab) response against all four viral serotypes. The burst of plasmablasts in the peripheral blood after vaccination may reflect enriched vaccine-specific Ab secreting cells. Here we characterize the acute plasmablast responses from naïve and DENV-exposed individuals following immunization with the live attenuated tetravalent (LAT) Butantan DENV vaccine (Butantan-DV). The frequency of circulating plasmablasts was determined by flow cytometric analysis of fresh whole blood specimens collected from 40 participants enrolled in the Phase II Butantan-DV clinical trial (NCT01696422) before and after (days 6, 12, 15 and 22) vaccination. We observed a peak in the number of circulating plasmablast at day 15 after vaccination in both the DENV naïve and the DENV-exposed vaccinees. DENV-exposed vaccinees experienced a significantly higher plasmablast expansion. In the DENV-naïve vaccinees, plasmablasts persisted for approximately three weeks longer than among DENV-exposed volunteers. Our findings indicate that the Butantan-DV can induce plasmablast responses in both DENV-naïve and DENV-exposed individuals and demonstrate the influence of pre-existing DENV immunity on Butantan DV-induced B-cell responses.

Keywords: dengue; dengue infection; dengue vaccine; humoral response; plasmablast.

Conflict of interest statement

JK and RP are former employees of the Butantan Institute. AP is an employee of the Butantan Institute. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The reviewer (CP) declared a shared affiliation with the author (MT) to the handling editor at the time of review.

Copyright © 2022 Silveira, Magnani, Costa, Avelino-Silva, Ricciardi, Timenetsky, Goulart, Correia, Marmorato, Ferrari, Nakagawa, Tomiyama, Tomiyama, Kalil, Palacios, Precioso, Watkins and Kallás.

Figures

Figure 1
Figure 1
Study design. Naïve and DENV-pre-exposed volunteers received a single subcutaneous dose of the lyophilized vaccine consisting of 1,000 PFU of each attenuated virus or placebo (qualified Leibovitz L-15 medium only). Blood samples were collected pre-vaccination (baseline, day 0) and at days 3, 6, 9, 11-12, 15, 21-22, 28, 56 and 91 post-vaccination, and used for the assessment of vaccine induce-viremia, DENV neutralizing antibodies (nAbs) titers using plaque reduction test (PRNT), and plasmablast responses using flow cytometry.
Figure 2
Figure 2
Logarithm-transformed DENV neutralizing antibody (nAb) titers from naïve (white) (A) and DENV pre-exposed (gray) (B) volunteers at baseline and post-vaccination. Data was generated by 50% plaque-reduction neutralization titer assay (PRNT50). Circles and triangles represent median log titers for each DENV serotype in placebo and vaccinees volunteers, respectively. Horizontal lines span the 25th-75th percentiles (interquartile range). The values on the right of each time point column represents the p-values obtained after the analysis of placebo and vaccinees nAb titers using non-parametric Wilcoxon rank-sum test.
Figure 3
Figure 3
Representative flow cytometric analysis of circulating plasmablast in naïve (DENV (–)) and DENV-pre-exposed (DENV(+)) volunteers enrolled in the Phase II clinical trial evaluated before (baseline) and after vaccination (days 6, 11-12, 15, 21-22). All plots represent the frequency of CD27+ CD38+ cells gated on live CD3- CD14- CD19+ cells.
Figure 4
Figure 4
Plasmablast expansion (as the percentage of CD20- CD27+ CD38+ cells among the CD19+ population) after Butantan-DV vaccination. Results are presented as pooled total plasmablast cells analyzed in a Boolean analysis. (A, B) Frequency of plasmablast in fresh blood from DENV (–) (A) and DENV(+) (B) volunteers at baseline and days 6, 11-12, 15 and 21-22 post-immunization. D, days post-vaccination. P, placebo. V, vaccinee. Horizontal bars represent median and interquartile range. Statistical analysis was performed using a non-paired t test. (C) Difference in the frequency of plasmablast comparing the values obtained at baseline and at the peak of plasmablast expansion (D15 after Butantan-DV vaccination) in naïve [DENV (–)] and DENV pre-exposed [DENV(+)) vaccinees. DENV(+) volunteers were divided into individuals pre-exposed to 1 or 2 or more serotypes, according to PRNT50 results to all four DENV serotypes at baseline. Data were analyzed with the Mann-Whitney U test.

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