scholarly journals Functional Transplant of a Dengue Virus Serotype 3 (DENV3)-Specific Human Monoclonal Antibody Epitope into DENV1

2016 ◽  
Vol 90 (10) ◽  
pp. 5090-5097 ◽  
Author(s):  
William B. Messer ◽  
Boyd L. Yount ◽  
Scott R. Royal ◽  
Ruklanthi de Alwis ◽  
Douglas G. Widman ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Dengue Virus ◽  
Neutralizing Antibodies ◽  
Human Monoclonal Antibody ◽  
Viral Envelope ◽  
Severe Dengue ◽  
Denv Serotypes ◽  
The Core ◽  
Antibody Epitope ◽  
Virus Serotype

ABSTRACTThe four dengue virus (DENV) serotypes, DENV1 through 4, are endemic throughout tropical and subtropical regions of the world. While first infection confers long-term protective immunity against viruses of the infecting serotype, a second infection with virus of a different serotype carries a greater risk of severe dengue disease, including dengue hemorrhagic fever and dengue shock syndrome. Recent studies demonstrate that humans exposed to DENV infections develop neutralizing antibodies that bind to quaternary epitopes formed by the viral envelope (E) protein dimers or higher-order assemblies required for the formation of the icosahedral viral envelope. Here we show that the quaternary epitope target of the human DENV3-specific neutralizing monoclonal antibody (MAb) 5J7 can be partially transplanted into a DENV1 strain by changing the core residues of the epitope contained within a single monomeric E molecule. MAb 5J7 neutralized the recombinant DENV1/3 strain in cell culture and was protective in a mouse model of infection with the DENV1/3 strain. However, the 5J7 epitope was only partially recreated by transplantation of the core residues because MAb 5J7 bound and neutralized wild-type (WT) DENV3 better than the DENV1/3 recombinant. Our studies demonstrate that it is possible to transplant a large number of discontinuous residues between DENV serotypes and partially recreate a complex antibody epitope, while retaining virus viability. Further refinement of this approach may lead to new tools for measuring epitope-specific antibody responses and new vaccine platforms.IMPORTANCEDengue virus is the most important mosquito-borne pathogen of humans worldwide, with approximately one-half the world's population living in regions where dengue is endemic. Dengue immunity following infection is robust and thought to be conferred by antibodies raised against the infecting virus. However, the specific viral components that these antibodies recognize and how they neutralize the virus have been incompletely described. Here we map a region on dengue virus serotype 3 recognized by the human neutralizing antibody 5J7 and then test the functional significance of this region by transplanting it into a serotype 1 virus. Our studies demonstrate a region on dengue virus necessary for 5J7 binding and neutralization. Our work also demonstrates the technical feasibility of engineering dengue viruses to display targets of protective antibodies. This technology can be used to develop new dengue vaccines and diagnostic assays.

scholarly journals A New Quaternary Structure Epitope on Dengue Virus Serotype 2 Is the Target of Durable Type-Specific Neutralizing Antibodies

mBio ◽  
2015 ◽  
Vol 6 (5) ◽  
Author(s):  
E. N. Gallichotte ◽  
D. G. Widman ◽  
B. L. Yount ◽  
W. M. Wahala ◽  
A. Durbin ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Dengue Virus ◽  
Neutralizing Antibodies ◽  
Quaternary Structure ◽  
Natural Infection ◽  
Viral Envelope ◽  
Protective Response ◽  
Denv Serotypes ◽  
Virus Serotype ◽  
Dengue Virus Serotype 2

ABSTRACT Dengue virus serotype 2 (DENV2) is widespread and responsible for severe epidemics. While primary DENV2 infections stimulate serotype-specific protective responses, a leading vaccine failed to induce a similar protective response. Using human monoclonal antibodies (hMAbs) isolated from dengue cases and structure-guided design of a chimeric DENV, here we describe the major site on the DENV2 envelope (E) protein targeted by neutralizing antibodies. DENV2-specific neutralizing hMAb 2D22 binds to a quaternary structure epitope. We engineered and recovered a recombinant DENV4 that displayed the 2D22 epitope. DENV2 neutralizing antibodies in people exposed to infection or a live vaccine tracked with the 2D22 epitope on the DENV4/2 chimera. The chimera remained sensitive to DENV4 antibodies, indicating that the major neutralizing epitopes on DENV2 and -4 are at different sites. The ability to transplant a complex epitope between DENV serotypes demonstrates a hitherto underappreciated structural flexibility in flaviviruses, which could be harnessed to develop new vaccines and diagnostics. IMPORTANCE Dengue virus causes fever and dengue hemorrhagic fever. Dengue serotype 2 (DENV2) is widespread and frequently responsible for severe epidemics. Natural DENV2 infections stimulate serotype-specific neutralizing antibodies, but a leading DENV vaccine did not induce a similar protective response. While groups have identified epitopes of single monoclonal antibodies (MAbs), the molecular basis of DENV2 neutralization by polyclonal human immune sera is unknown. Using a recombinant DENV displaying serotype 2 epitopes, here we map the main target of DENV2 polyclonal neutralizing antibodies induced by natural infection and a live DENV2 vaccine candidate. Proper display of the epitope required the assembly of viral envelope proteins into higher-order structures present on intact virions. Despite the complexity of the epitope, it was possible to transplant the epitope between DENV serotypes. Our findings have immediate implications for evaluating dengue vaccines in the pipeline as well as designing next-generation vaccines.

scholarly journals Cryo-EM structure of an antibody that neutralizes dengue virus type 2 by locking E protein dimers

Science ◽  
2015 ◽  
Vol 349 (6243) ◽  
pp. 88-91 ◽  
Author(s):  
Guntur Fibriansah ◽  
Kristie D. Ibarra ◽  
Thiam-Seng Ng ◽  
Scott A. Smith ◽  
Joanne L. Tan ◽  
...  
Keyword(s):  
Dengue Virus ◽  
Secondary Infection ◽  
Human Monoclonal Antibody ◽  
Viral Envelope ◽  
Severe Dengue ◽  
Virus Surface ◽  
Dengue Disease ◽  
Denv Serotypes ◽  
E Protein ◽  
Protein Dimers

There are four closely-related dengue virus (DENV) serotypes. Infection with one serotype generates antibodies that may cross-react and enhance infection with other serotypes in a secondary infection. We demonstrated that DENV serotype 2 (DENV2)–specific human monoclonal antibody (HMAb) 2D22 is therapeutic in a mouse model of antibody-enhanced severe dengue disease. We determined the cryo–electron microscopy (cryo-EM) structures of HMAb 2D22 complexed with two different DENV2 strains. HMAb 2D22 binds across viral envelope (E) proteins in the dimeric structure, which probably blocks the E protein reorganization required for virus fusion. HMAb 2D22 “locks” two-thirds of or all dimers on the virus surface, depending on the strain, but neutralizes these DENV2 strains with equal potency. The epitope defined by HMAb 2D22 is a potential target for vaccines and therapeutics.

scholarly journals Dengue Virus Serotype 4 Is Responsible for the Outbreak of Dengue in East Java City of Jember, Indonesia

Viruses ◽  
2020 ◽  
Vol 12 (9) ◽  
pp. 913
Author(s):  
Aryati Aryati ◽  
Billy J. Wrahatnala ◽  
Benediktus Yohan ◽  
May Fanny ◽  
Faradila K. N. Hakim ◽  
...  
Keyword(s):  
Dengue Virus ◽  
Clinical Symptoms ◽  
Secondary Infection ◽  
Herd Immunity ◽  
Antibody Detection ◽  
Severe Dengue ◽  
Envelope Gene ◽  
Genotype I ◽  
Denv Serotypes ◽  
Virus Serotype

Outbreaks of dengue virus (DENV) in Indonesia have been mainly caused by the DENV serotype-1; -2; or -3. The DENV-4 was the least-reported serotype in Indonesia during the last five decades. We recently conducted a molecular epidemiology study of dengue in the Jember regency, East Java province, Indonesia. Dengue is endemic in the region and outbreaks occur annually. We investigated the clinical characteristics and etiology of dengue-like febrile illness in this regency to understand the disease dynamics. A total of 191 patients with clinical symptoms similar to dengue were recruited during an 11-month study in 2019–2020. Children accounted for the majority of cases and dengue burden was estimated in 41.4% of the cases based on NS1 antigen, viral RNA, and IgG/IgM antibody detection with the majority (73.4%) being primary infections. Secondary infection was significantly associated with a higher risk of severe dengue manifestation. All four DENV serotypes were detected in Jember. Strikingly, we observed the predominance of DENV-4, followed by DENV-3, DENV-1, and DENV-2. Genotype determination using Envelope gene sequence revealed the classification into Genotype I, Cosmopolitan Genotype, Genotype I, and Genotype II for DENV-1, -2, -3, and -4, respectively. The predominance of DENV-4 in Jember may be associated with a new wave of DENV infections and spread in a non-immune population lacking a herd-immunity to this particular serotype.

scholarly journals Neutralizing antibody titers against dengue virus correlate with protection from symptomatic infection in a longitudinal cohort

2016 ◽  
Vol 113 (3) ◽  
pp. 728-733 ◽  
Author(s):  
Leah C. Katzelnick ◽  
Magelda Montoya ◽  
Lionel Gresh ◽  
Angel Balmaseda ◽  
Eva Harris
Keyword(s):  
Dengue Virus ◽  
Neutralizing Antibodies ◽  
Secondary Infection ◽  
Severe Disease ◽  
Neutralizing Antibody ◽  
Denv Infection ◽  
Severe Dengue ◽  
Symptomatic Infection ◽  
Denv Serotypes ◽  
Antibody Titers

The four dengue virus serotypes (DENV1–4) are mosquito-borne flaviviruses that infect ∼390 million people annually; up to 100 million infections are symptomatic, and 500,000 cases progress to severe disease. Exposure to a heterologous DENV serotype, the specific infecting DENV strains, and the interval of time between infections, as well as age, ethnicity, genetic polymorphisms, and comorbidities of the host, are all risk factors for severe dengue. In contrast, neutralizing antibodies (NAbs) are thought to provide long-lived protection against symptomatic infection and severe dengue. The objective of dengue vaccines is to provide balanced protection against all DENV serotypes simultaneously. However, the association between homotypic and heterotypic NAb titers and protection against symptomatic infection remains poorly understood. Here, we demonstrate that the titer of preinfection cross-reactive NAbs correlates with reduced likelihood of symptomatic secondary infection in a longitudinal pediatric dengue cohort in Nicaragua. The protective effect of NAb titers on infection outcome remained significant when controlled for age, number of years between infections, and epidemic force, as well as with relaxed or more stringent criteria for defining inapparent DENV infections. Further, individuals with higher NAb titers immediately after primary infection had delayed symptomatic infections compared with those with lower titers. However, overall NAb titers increased modestly in magnitude and remained serotype cross-reactive in the years between infections, possibly due to reexposure. These findings establish that anti-DENV NAb titers correlate with reduced probability of symptomatic DENV infection and provide insights into longitudinal characteristics of antibody-mediated immunity to DENV in an endemic setting.

scholarly journals Generation and characterization of cross neutralizing human monoclonal antibody against 4 serotypes of dengue virus without enhancing activity

PeerJ ◽  
2017 ◽  
Vol 5 ◽  
pp. e4021 ◽  
Author(s):  
Subenya Injampa ◽  
Nataya Muenngern ◽  
Chonlatip Pipattanaboon ◽  
Surachet Benjathummarak ◽  
Khwanchit Boonha ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Dengue Virus ◽  
Mammalian Cells ◽  
Human Monoclonal Antibody ◽  
Site Directed Mutagenesis ◽  
Neutralizing Activity ◽  
Dengue Disease ◽  
Denv Serotypes ◽  
Fusion Loop

Background Dengue 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. Methods In 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 FcγR bearing cells. To produce sufficient quantities of B3B9 rIgG for further characterization, CHO-K1 cells stably secreting N297Q-B3B9 rIgG were then established. Results The 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 FcγRI- and RII-bearing THP-1 cells and FcγRII-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. Discussion Human 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.

scholarly journals Dengue Fever and Severe Dengue in Barbados, 2008–2016

2020 ◽  
Vol 5 (2) ◽  
pp. 68
Author(s):  
Kirk Osmond Douglas ◽  
Sudip Kumar Dutta ◽  
Byron Martina ◽  
Fatih Anfasa ◽  
T. Alafia Samuels ◽  
...  
Keyword(s):  
Dengue Virus ◽  
Dengue Fever ◽  
Small Sample Size ◽  
Health Planning ◽  
Case Fatality ◽  
Small Sample ◽  
Severe Dengue ◽  
Medical Attention ◽  
Enzyme Linked Immunosorbent Assay ◽  
Virus Serotype

Analysis of the temporal, seasonal and demographic distribution of dengue virus (DENV) infections in Barbados was conducted using national surveillance data from a total of 3994 confirmed dengue cases. Diagnosis was confirmed either by DENV–specific real time reverse transcriptase polymerase chain reaction (rRT–PCR), or non–structural protein 1 (NS1) antigen or enzyme linked immunosorbent assay (ELISA) tests; a case fatality rate of 0.4% (10/3994) was observed. The dengue fever (DF) prevalence varied from 27.5 to 453.9 cases per 100,000 population among febrile patients who sought medical attention annually. DF cases occurred throughout the year with low level of transmission observed during the dry season (December to June), then increased transmission during rainy season (July to November) peaking in October. Three major dengue epidemics occurred in Barbados during 2010, 2013 and possibly 2016 with an emerging three–year interval. DF prevalence among febrile patients who sought medical attention overall was highest among the 10–19 years old age group. The highest DF hospitalisation prevalence was observed in 2013. Multiple serotypes circulated during the study period and Dengue virus serotype 2 (DENV–2) was the most prevalent serotype during 2010, whilst DENV–1 was the most prevalent serotype in 2013. Two DENV–1 strains from the 2013 DENV epidemic were genetically more closely related to South East Asian strains, than Caribbean or South American strains, and represent the first ever sequencing of DENV strains in Barbados. However, the small sample size (n = 2) limits any meaningful conclusions. DF prevalence was not significantly different between females and males. Public health planning should consider DENV inter–epidemic periodicity, the current COVID–19 pandemic and similar clinical symptomology between DF and COVID–19. The implementation of routine sequencing of DENV strains to obtain critical data can aid in battling DENV epidemics in Barbados.

scholarly journals Tracking the polyclonal neutralizing antibody response to a dengue virus serotype 1 type-specific epitope across two populations in Asia and the Americas

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Daniela V. Andrade ◽  
Colin Warnes ◽  
Ellen Young ◽  
Leah C. Katzelnick ◽  
Angel Balmaseda ◽  
...  
Keyword(s):  
Dengue Virus ◽  
Neutralizing Antibodies ◽  
Neutralizing Antibody ◽  
Sri Lankan ◽  
Human Monoclonal Antibodies ◽  
Virus Serotype ◽  
Serotype 1 ◽  
Dengue Virus Serotypes ◽  
Kinetics Of ◽  
Two Populations

Abstract The four dengue virus serotypes (DENV1-4) cause major public health problems worldwide. Highly neutralizing type-specific human monoclonal antibodies (hmAbs) target conformation-dependent epitopes on the DENV envelope protein, including 1F4, a DENV1 type-specific hmAb. Using a recombinant DENV2 virus displaying the DENV1 1F4 epitope (rDENV2/1), we measured the proportion and kinetics of DENV1 neutralizing antibodies targeting the 1F4 epitope in individuals living in Asia and the Americas where different DENV1 genotypes were circulating. Samples from 20 individuals were analyzed 3 and 18 months post-primary DENV1 infection, alongside samples from 4 individuals collected annually for four years post-primary DENV1 infection, from two studies in Nicaragua. We also analyzed convalescent post-primary DENV1 plasma samples from Sri Lankan individuals. We found that neutralizing antibodies recognizing the 1F4 epitope vary in prevalence across both populations and were detected from 20 days to four years post-infection. Additionally, both populations displayed substantial variability, with a range of high to low proportions of DENV1 type-specific neutralizing antibodies recognizing the 1F4 epitope seen across individuals. Thus, the 1F4 epitope is a major but not exclusive target of type-specific neutralizing antibodies post-primary infection with different DENV1 genotypes in Asia and Latin America, and additional epitopes likely contribute to type-specific neutralization of DENV1.

scholarly journals Isolation of a human monoclonal antibody specific for the receptor binding domain of SARS-CoV-2 using a competitive phage biopanning strategy

2020 ◽  
Vol 3 (2) ◽  
pp. 95-100 ◽  
Author(s):  
Xin Zeng ◽  
Lingfang Li ◽  
Jing Lin ◽  
Xinlei Li ◽  
Bin Liu ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Receptor Binding ◽  
Neutralizing Antibodies ◽  
Magnetic Beads ◽  
Human Monoclonal Antibody ◽  
Binding Domain ◽  
Receptor Binding Domain ◽  
Antibody Library ◽  
Synthetic Antibody ◽  
Blocking Antibodies

Abstract The infection of the novel coronavirus severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused more than 200 000 deaths, but no vaccine or therapeutic monoclonal antibody is currently available. SARS-CoV-2 relies on its spike protein, in particular the receptor-binding domain (RBD), to bind human cell receptor angiotensin-converting enzyme 2 (ACE2) for viral entry, and thus targeting RBD holds the promise for preventing SARS-CoV-2 infection. In this work, a competitive biopanning strategy of a phage display antibody library was applied to screen blocking antibodies against RBD. High-affinity antibodies were enriched after the first round using a standard panning process in which RBD-His was immobilized as a bait. At the next two rounds, immobilized ACE2-Fc and free RBD-His were mixed with the enriched phage antibodies. Antibodies binding to RBD at epitopes different from ACE2-binding site were captured by the immobilized ACE2-Fc, forming a “sandwich” complex. Only antibodies competed with ACE2 can bind to the free RBD-His in the supernatant and be subsequently separated by the nickel-nitrilotriacetic acid magnetic beads. rRBD-15 from the competitive biopanning of our synthetic antibody library, Lib AB1, was produced as the full-length IgG1 format. It was proved to competitively block the binding of RBD to ACE2 and potently inhibit SARS-CoV-2 pseudovirus infection with IC50 values of 12 nM. Nevertheless, rRBD-16 from the standard biopanning can only bind to RBD in vitro, but not have the blocking or neutralization activity. Our strategy can efficiently isolate the blocking antibodies of RBD, and it would speed up the discovery of neutralizing antibodies against SARS-CoV-2.

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