Flaviviruses
Program Leader: Aravinda deSilva
SE-RP-004: Antibody in protective and pathogenic immunity to Dengue type 3
Aravinda deSilva and Ralph Baric
University of North Carolina
A large body of work has demonstrated that people infected with DENV develop long-term protective immunity to the infecting serotype but not to other serotypes. During a second infection with a different serotype, the risk of severe disease is greater because cross-reactive immunity can exacerbate the disease. Efforts to develop DEN vaccines have been hampered by the dual role of immunity in protection and pathogenesis. The main goal of this project is to use DENV3 as a model for defining the viral epitopes engaged by human antibody and to determine the functional outcome of these interactions. Our overall hypothesis is that distinct epitopes on the DENV envelope protein are responsible for virus neutralization and enhancement. The goal of aim 1 is to produce human monoclonal antibodies (hMAbs) from people who have recovered from DENV infections. In collaboration with Jim Crowe’s laboratory at Vanderbilt we have already produced 38 new hMAbs and completed the initial characterization of these antibodies. Most hMAbs were dengue serotype cross reactive and weakly neutralizing, indicating that strongly neutralizing antibodies are minor fraction of the total response. Many hMAbs recognized preM protein, which has hitherto not been considered a significant target of antibody. Under aim 2 we are performing a more detailed mapping of the epitopes recognized by hMAbs. Studies are also underway to characterize the polyclonal human antibody response to dengue. To date our studies have demonstrated that serum antibodies consist of distinct populations of dengue serotype specific and dengue cross reactive antibodies. These antibodies were also functionally distinct- the type specific antibodies were responsible for virus neutralization, whereas the cross reactive antibodies were responsible for enhanced infection of Fc receptor bearing cells. Under aim 3 studies have been conducted in collaboration with Ralph Baric’s group to determine if natural strain variation in DENV3 influences antibody interactions. Using an infectious clone of DENV3 generated at UNC, we have created recombinant viruses with the E glycoprotein from each of the major genotypes of DENV3. Studies with this panel of viruses have demonstrated that natural variation in DENV3 leads to loss of antibody binding and neutralization. This new information challenges the dogma that all strains within a dengue serotype are equally susceptible to antibody neutralization. We are also producing and purifying large quantities of dengue virions and recombinant protein antigens to support our project and other DENV projects within SERCEB.
SE-RP-005: T cell response to Dengue virus serotype 3 in mice and human
LaJolla Institute of Allergy and Infectious Disease
The four serotypes of Dengue virus (DENV) cause dengue fever (DF) and dengue hemorrhagic fever/dengue shock syndrome (DHF/DSS) in humans. Studies suggest that DHF/DSS is an immunopathogenic disease in which T cell responses may be dysregulated. Understanding the mechanisms that regulate the balance between T cell mediated pathology versus protection is crucial for developing therapies and, importantly, a safe dengue vaccine. A major goal of this project is to discover and validate cytotoxic T cell epitopes presented by HLA class I molecules that are derived from DENV serotype 3 (DENV3). Epitopes will be identified by the use of epitope predictions method, and then validated by determining the reactivity of the epitopes with peripheral blood mononuclear cells (PBMC) from DENV-exposed human donors. For selected epitopes, the phenotype and pattern of cytokines observed in response to the epitope itself and its crossreactivity pattern with homologous sequences derived from the other serotypes will be examined. For a subset of the epitopes recognized in humans, the protective capacity of the epitopes will be assessed in HLA transgenic mice. This project focuses on DENV3 because epidemiologic studies suggest that certain DENV serotypes, genotypes, and the sequence of DENV infections may be associated with DHF/DSS, and little is known about DENV3 infection as compared with DENV2, although DENV3 co-circulates with DENV2 in endemic countries, and DENV3 has been responsible for recent epidemics in South Asia and Latin America.
Relevance
The current vaccine candidates that exploit antibody responses alone may be ineffective against new DENV variants and may enhance disease. This research may aid in generating DENV-specific vaccines that are broadly cross-reactive against all four serotypes, effectively clear virus, and increase protective immunity while avoiding immunopathology.
