Dengue virus (DENV) infects an estimated 400 million people every year, causing prolonged morbidity and sometimes mortality. Development of an effective vaccine has been hampered by the lack of appropriate small animal models; mice are naturally not susceptible to DENV and only become infected if highly immunocompromised. -cell response to viral infection, compared to a weak response in IFNAR؊/؊ mice. Furthermore, mice lacking IFNAR on either CD11c ؉ or LysM ؉ cells were also sufficiently immunocompetent to raise a protective immune response to a candidate subunit vaccine against DENV-2. These data demonstrate that mice with conditional deficiencies in expression of the IFNAR represent improved models for the study of DENV immunology and screening of vaccine candidates. IMPORTANCEDengue virus infects 400 million people every year worldwide, causing 100 million clinically apparent infections, which can be fatal if untreated. Despite many years of research, there are no effective vaccine and no antiviral treatment available for dengue. Development of vaccines has been hampered in particular by the lack of a suitable small animal model. Mouse models used to test dengue vaccine are deficient in interferon (IFN) type I signaling and severely immunocompromised and therefore likely not ideal for the testing of vaccines. In this study, we explored alternative models lacking the IFN receptor only on certain cell types. We show that mice lacking the IFN receptor on either CD11c-or LysM-expressing cells (conditional IFNAR mice) are susceptible to dengue virus infection. Importantly, we demonstrate that conditional IFN receptor knockout mice generate a better immune response to live virus and a candidate dengue vaccine compared to IFNAR mice and are resistant to subsequent challenge. D engue virus (DENV, a member of the Flaviviridae family, is a mosquito-borne pathogen that infects approximately 400 million people every year (1, 2). Each of the four DENV serotypes causes a spectrum of clinical symptoms ranging from mild fever to potentially fatal manifestations of dengue shock syndrome. DENV causes an acute infection with high fever, which usually resolves after 5 to 7 days. At this time, most patients have cleared the high virus load. Intriguingly, however, this is also the time point when some patients start to develop vascular leakage, which, if untreated, can lead to a collapse of the metabolism and organ failure. The frequency, severity, and geographical spread of cases has increased over the past decades (3, 4), and DENV infection is now considered a leading cause of morbidity in the tropics.There are no effective treatments for dengue fever, and the development of a vaccine has been hampered by the lack of suitable small animal models. Wild-type (wt) mice are not susceptible to infection with field strains of DENV, and while viral replication in these animals can be forced by intracranial injections of hightiter mouse-adapted DENV strains, the resulting clinical disease bears little resemblance to dengue fev...

A dengue vaccine should induce long-lasting, simultaneous protection to the four dengue viruses while avoiding the immune enhancement of viral infection. Domain III of the dengue envelope protein has been implicated in receptor binding, and is also the target of specific neutralizing antibodies. Domain III has emerged as a promising region for a subunit vaccine candidate. Here, we review the current state of knowledge on vaccine candidates based on domain III. Due to the results obtained concerning the immune response and protection in mice and monkeys, particular attention is paid to the chimeric protein domain III fused to p64k of Neisseria meningitidis.

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The capsid protein is one of the three structural proteins of flaviviruses and is the building block of the nucleocapsid. It has also a predominant role in the replication of dengue virus. To obtain nucleocapsid-like particles from recombinant dengue-2 capsid protein produced in E. coli, a purification process using cation exchange chromatography was established. The purified protein exhibited a molecular mass corresponding to a dimer; therefore, similar to that reported for alphaviruses, an in vitro assembly reaction using single-stranded DNA was performed. In all cases, particles were obtained independently of the specificity and the length of the oligonucleotides used. The present work is the first report of in vitro assembly of the recombinant dengue capsid protein, which could constitute a powerful tool in the development of vaccine candidates.

Based on the immunogenicity of domain III from the Envelope protein of dengue virus as well as the proven protective capacity of the capsid antigen, we have designed a novel domain III-capsid chimeric protein with the goal of obtaining a molecule potentially able to induce both humoral and cell-mediated immunity (CMI). After expression of the recombinant gene in Escherichia coli, the domain III moiety retained its antigenicity as evaluated with anti-dengue sera. In order to explore alternatives for modulating the immunogenicity of the protein, it was mixed with oligodeoxynucleotides in order to obtain particulated aggregates and then immunologically evaluated in mice in comparison with non-aggregated controls. Although the humoral immune response induced by both forms of the protein was equivalent, the aggregated variant resulted in a much stronger CMI as measured by in vitro IFN-gamma secretion and protection experiments, mediated by CD4(+) and CD8(+) cells. The present work provides additional evidence in support for a crucial role of CMI in protection against dengue virus and describes a novel vaccine candidate against the disease based on a recombinant protein that can stimulate both arms of the acquired immune system.

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