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Development, characterization and application of monoclonal antibodies against Brazilian Dengue virus isolates.

ABSTRACT: Dengue is the most prevalent human arboviral disease. The morbidity related to dengue infection supports the need for an early, quick and effective diagnostic test. Brazil is a hotspot for dengue, but no serological diagnostic test has been produced using Brazilian dengue virus isolates. This study aims to improve the development of immunodiagnostic methods for dengue virus (DENV) detection through the production and characterization of 22 monoclonal antibodies (mAbs) against Brazilian isolates of DENV-1, -2 and -3. The mAbs include IgG2b?, IgG2a? and IgG1? isotypes, and most were raised against the envelope or the pre-membrane proteins of DENV. When the antibodies were tested against the four DENV serotypes, different reactivity patterns were identified: group-specific, subcomplex specific (DENV-1, -3 and -4 and DENV-2 and -3) and dengue serotype-specific (DENV-2 or -3). Additionally, some mAbs cross-reacted with yellow fever virus (YFV), West Nile virus (WNV) and Saint Louis encephalitis virus (SLEV). None of the mAbs recognized the alphavirus Venezuelan equine encephalitis virus (VEEV). Furthermore, mAbs D3 424/8G, D1 606/A12/B9 and D1 695/12C/2H were used to develop a capture enzyme-linked immunosorbent assay (ELISA) for anti-dengue IgM detection in sera from patients with acute dengue. To our knowledge, these are the first monoclonal antibodies raised against Brazilian DENV isolates, and they may be of special interest in the development of diagnostic assays, as well as for basic research.

SUBMITTER: Zanluca C

PROVIDER: S-EPMC4239016 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

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Development, characterization and application of monoclonal antibodies against Brazilian Dengue virus isolates.

Zanluca Camila C Mazzarotto Giovanny Augusto Camacho Antevere GA Bordignon Juliano J Duarte Dos Santos Claudia Nunes CN

PloS one 20141120 11

Dengue is the most prevalent human arboviral disease. The morbidity related to dengue infection supports the need for an early, quick and effective diagnostic test. Brazil is a hotspot for dengue, but no serological diagnostic test has been produced using Brazilian dengue virus isolates. This study aims to improve the development of immunodiagnostic methods for dengue virus (DENV) detection through the production and characterization of 22 monoclonal antibodies (mAbs) against Brazilian isolates ...[more]

PMID: 25412181

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Project description:Zika virus (ZIKV) is a mosquito-borne flavivirus responsible for thousands of cases of severe fetal malformations and neurological disease since its introduction to Brazil in 2013. Antibodies to flaviviruses can be protective, resulting in lifelong immunity to reinfection by homologous virus. However, cross-reactive antibodies can complicate flavivirus diagnostics and promote more severe disease, as noted after serial dengue virus (DENV) infections. The endemic circulation of DENV in South America and elsewhere raises concerns that preexisting flavivirus immunity may modulate ZIKV disease and transmission potential. Here, we report on the ability of human monoclonal antibodies and immune sera derived from dengue patients to neutralize contemporary epidemic ZIKV strains. We demonstrate that a class of human monoclonal antibodies isolated from DENV patients neutralizes ZIKV in cell culture and is protective in a lethal murine model. We also tested a large panel of convalescent-phase immune sera from humans exposed to primary and repeat DENV infection. Although ZIKV is most closely related to DENV compared to other human-pathogenic flaviviruses, most DENV immune sera (73%) failed to neutralize ZIKV, while others had low (50% effective concentration [EC50], <1:100 serum dilution; 18%) or moderate to high (EC50, >1:100 serum dilution; 9%) levels of cross-neutralizing antibodies. Our results establish that ZIKV and DENV share epitopes that are targeted by neutralizing, protective human antibodies. The availability of potently neutralizing human monoclonal antibodies provides an immunotherapeutic approach to control life-threatening ZIKV infection and also points to the possibility of repurposing DENV vaccines to induce cross-protective immunity to ZIKV. ZIKV is an emerging arbovirus that has been associated with severe neurological birth defects and fetal loss in pregnant women and Guillain-Barré syndrome in adults. Currently, there is no vaccine or therapeutic for ZIKV. The identification of a class of antibodies (envelope dimer epitope 1 [EDE1]) that potently neutralizes ZIKV in addition to all four DENV serotypes points to a potential immunotherapeutic to combat ZIKV. This is especially salient given the precedent of antibody therapy to treat pregnant women infected with other viruses associated with microcephaly, such as cytomegalovirus and rubella virus. Furthermore, the identification of a functionally conserved epitope between ZIKV and DENV raises the possibility that a vaccine may be able to elicit neutralizing antibodies against both viruses.

Project description:Dengue virus (DV) infection causes a spectrum of clinical diseases ranging from dengue fever to a life-threatening dengue hemorrhagic fever. Four distinct serotypes (DV1-4), which have similar genome sequences and envelope protein (E protein) antigenic properties, were divided. Among these 4 serotypes, DV1 usually causes predominant infections and fast diagnosis and effective treatments are urgently required to prevent further hospitalization and casualties.To develop antibodies specifically targeting and neutralizing DV1, we immunized mice with UV-inactivated DV1 viral particles and recombinant DV1 E protein from residue 1 to 395 (E395), and then generated 12 anti-E monoclonal antibodies (mAbs) as the candidates for a series of characterized assays such as ELISA, dot blot, immunofluorescence assay, western blot, and foci forming analyses.Among the mAbs, 10 out of 12 showed cross-reactivity to four DV serotypes as well as Japanese encephalitis virus (JEV) in different cross-reactivity patterns. Two particular mAbs, DV1-E1 and DV1-E2, exhibited strong binding specificity and neutralizing activity against DV1 and showed no cross-reactivity to DV2, DV3, DV4 or JEV-infected cells as characterized by ELISA, dot blot, immunofluorescence assay, western blot, and foci forming analyses. Using peptide coated indirect ELISA, we localized the neutralizing determinants of the strongly inhibitory mAbs to a sequence-unique epitope on the later-ridge of domain III of the DV1 E protein, centered near residues T346 and D360 (346TQNGRLITANPIVTD360). Interestingly, the amino acid sequence of the epitope region is highly conserved among different genotypes of DV1 but diverse from DV2, DV3, DV4 serotypes and other flaviviruses.Our results showed two selected mAbs DV1-E1 and DV1-E2 can specifically target and significantly neutralize DV1. With further research these two mAbs might be applied in the development of DV1 specific serologic diagnosis and used as a feasible treatment option for DV1 infection. The identification of DV1 mAbs epitope with key residues can also provide vital information for vaccine design.

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Development, characterization and application of monoclonal antibodies against Brazilian Dengue virus isolates.

Publications

Development, characterization and application of monoclonal antibodies against Brazilian Dengue virus isolates.

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