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Dengue virus infection-enhancing activity in serum samples with neutralizing activity as determined by using Fc?R-expressing cells.

ABSTRACT: BACKGROUND: Progress in dengue vaccine development has been hampered by limited understanding of protective immunity against dengue virus infection. Conventional neutralizing antibody titration assays that use Fc?R-negative cells do not consider possible infection-enhancement activity. We reasoned that as Fc?R-expressing cells are the major target cells of dengue virus, neutralizing antibody titration assays using Fc?R-expressing cells that determine the sum of neutralizing and infection-enhancing activity, may better reflect the biological properties of antibodies in vivo. METHODS AND FINDINGS: We evaluated serum samples from 80 residents of a dengue endemic country, Malaysia, for neutralizing activity, and infection-enhancing activity at 1?10 serum dilution by using Fc?R-negative BHK cells and Fc?R-expressing BHK cells. The serum samples consisted of a panel of patients with acute DENV infection (31%, 25/80) and a panel of donors without acute DENV infection (69%, 55/80). A high proportion of the tested serum samples (75%, 60/80) demonstrated DENV neutralizing activity (PRNT(50)?10) and infection-enhancing activity. Eleven of 18 serum samples from patients with acute secondary DENV infection demonstrated neutralizing activity to the infecting serotype determined by using Fc?R-negative BHK cells (PRNT(50)?10), but not when determined by using Fc?R-expressing cells. CONCLUSION: Human serum samples with low neutralizing activity determined by using Fc?R-negative cells showed DENV infection-enhancing activity using Fc?R-expressing cells, whereas those with high neutralizing activity determined by using Fc?R-negative cells demonstrate low or no infection-enhancing activity using Fc?R-expressing cells. The results suggest an inverse relationship between neutralizing antibody titer and infection-enhancing activity, and that neutralizing activity determined by using Fc?R-expressing cells, and not the activity determined by using Fc?R-negative cells, may better reflect protection to DENV infection in vivo.

SUBMITTER: Moi ML

PROVIDER: S-EPMC3289619 | biostudies-literature | 2012

REPOSITORIES: biostudies-literature

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Dengue virus infection-enhancing activity in serum samples with neutralizing activity as determined by using FcγR-expressing cells.

Moi Meng Ling ML Lim Chang-Kweng CK Chua Kaw Bing KB Takasaki Tomohiko T Kurane Ichiro I

PLoS neglected tropical diseases 20120228 2

<h4>Background</h4>Progress in dengue vaccine development has been hampered by limited understanding of protective immunity against dengue virus infection. Conventional neutralizing antibody titration assays that use FcγR-negative cells do not consider possible infection-enhancement activity. We reasoned that as FcγR-expressing cells are the major target cells of dengue virus, neutralizing antibody titration assays using FcγR-expressing cells that determine the sum of neutralizing and infection- ...[more]

PMID: 22389741

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Project description:Antibodies are critical responses to protect the host from dengue virus(DENV) infection. Antibodies target DENV by two pathologic mechanisms: virus neutralization and infection enhancement. In dengue patients, the absence of neutralizing activity in the presence of Fc?R implies that infection-enhancing activity hampers the neutralizing activity of antibodies, which could potentially lead to symptomatic presentations and severe clinical outcomes.A total of 100 pair serum samples from adult healthy volunteers were obtained during the dengue season in Ha Noi in 2015 for evaluation of neutralizing and infection-enhancing activity. Additionally, 20 serum samples from acute secondary DENV infection patients were also used as the patient group in this study. PRNT was performed on BHK cells and Fc?R-expressing BHK cell lines for all serum samples.Out of 100 residents, positive neutralizing antibodies (N.A) were found in 44.23 and 76.92% for DENV-1; 38.46 and 75% for DENV-2; 19.23 and 15.38% for DENV-3; and 1.92 and 9.62% for DENV-4 for pre and post-dengue season respectively. The percentage of post-exposure residents having positive responses against single, two, or more than three DENV serotypes were 38.46, 44.23 and 15.38%, respectively. A total of 34 residents were DENV seropositive before the dengue season and these individuals demonstrated further elevation of IgG antibodies after the dengue season. At the end of the season, 18 residents were confirmed to be new asymptomatic DENV infection cases. In both groups, N.A titers determined on BHK cells were higher than that on Fc?R-expressing BHK cells. In heterotypic N.A responses, N.A titers to the infecting serotype from the samples obtained from pre-exposure group were significantly higher than those of the patient group. However, fold enhancement to the infecting serotypes from the samples in the pre-exposure group was substantially lower as compared to that of the patient group.Before and after the dengue season, serum samples from healthy volunteers demonstrated high levels of neutralizing antibodies and low or absence of infection-enhancement activity. The results suggest that while infection-enhancement activity hampers neutralizing activity of antibodies, high levels of DENV neutralizing antibodies set a critical threshold in facilitating the prevention of disease progression.

Project description:BackgroundDengue disease is a leading cause of illness and death in the tropics and subtropics. Most severe cases occur among patients secondarily infected with a different dengue virus (DENV) serotype compared with that from the first infection, resulting in antibody-dependent enhancement activity (ADE). Our previous study generated the neutralizing human monoclonal antibody, D23-1B3B9 (B3B9), targeting the first domain II of E protein, which showed strong neutralizing activity (NT) against all four DENV serotypes. However, at sub-neutralizing concentrations, it showed ADE activity in vitro.MethodsIn this study, we constructed a new expression plasmid using the existing IgG heavy chain plasmid as a template for Fc modification at position N297Q by site-directed mutagenesis. The resulting plasmid was then co-transfected with a light chain plasmid to produce full recombinant IgG (rIgG) in mammalian cells (N297Q-B3B9). This rIgG was characterized for neutralizing and enhancing activity by using different Fc?R bearing cells. To produce sufficient quantities of B3B9 rIgG for further characterization, CHO-K1 cells stably secreting N297Q-B3B9 rIgG were then established.ResultsThe generated N297Q-B3B9 rIgG which targets the conserved N-terminal fusion loop of DENV envelope protein showed the same cross-neutralizing activity to all four DENV serotypes as those of wild type rIgG. In both Fc?RI- and RII-bearing THP-1 cells and Fc?RII-bearing K562 cells, N297Q-B3B9 rIgG lacked ADE activity against all DENV serotypes at sub-neutralizing concentrations. Fortunately, the N297Q-B3B9 rIgG secreted from stable cells showed the same patterns of NT and ADE activities as those of the N297Q-B3B9 rIgG obtained from transient expression against DENV2. Thus, the CHO-K1 stably expressing N297Q-B3B9 HuMAb can be developed as high producer stable cells and used to produce sufficient amounts of antibody for further characterization as a promising dengue therapeutic candidate.DiscussionHuman monoclonal antibody, targeted to fusion loop of envelope domainII (EDII), was generated and showed cross-neutralizing activity to 4 serotypes of DENV, but did not cause any viral enhancement activity in vitro. This HuMAb could be further developed as therapeutic candidates.

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Dengue virus infection-enhancing activity in serum samples with neutralizing activity as determined by using Fc?R-expressing cells.

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Dengue virus infection-enhancing activity in serum samples with neutralizing activity as determined by using FcγR-expressing cells.

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