After incubation for 2 (ZIKV) or 3 d (DENV), supernatants were harvested, and viral titers were determined by focus-forming assays

After incubation for 2 (ZIKV) or 3 d (DENV), supernatants were harvested, and viral titers were determined by focus-forming assays. Ethics. This study was carried out in accordance with the recommendations in the Guide for the Care and Use of Laboratory Animals of the National Institutes of Health and the UK Home Office Animals Act Project License. statement, we describe the design and production of covalently stabilized ZIKV E dimers, which lack precursor membrane protein and do not expose the immunodominant fusion loop epitope. Immunization of mice with ZIKV E dimers induces dimer-specific antibodies, which protect against ZIKV challenge during pregnancy. Importantly, the ZIKV E-dimer-induced response does not cross-react with DENV or induce ADE of DENV contamination. Following the Pacific Islands epidemics in 2013 and 2014, Zika computer virus (ZIKV) spread across South America, leading to an explosive outbreak in Brazil in 20156. Although most ZIKV infections lead to asymptomatic or moderate disease, severe neurological Rabbit Polyclonal to BST1 sequelae can ensue, including Guillain-Barr syndrome (which was first recognized after the French Polynesian outbreak7), and microcephaly and other JW-642 congenital anomalies (which were observed in children born to mothers who suffered a ZIKV contamination during pregnancy8). JW-642 There has been much study of ZIKV since the outbreak in Brazil, which has clarified several key features of ZIKV pathogenesis, including the ability of the computer virus to infect and cross the placenta, targeting specific neuroprogenitor cells in the developing fetal brain9. Because of the initial high contamination rate in a naive populace in South America, the ZIKV outbreak produced herd immunity, which has essentially terminated the epidemic, although sporadic cases still occur. However, the catastrophic effects of contamination during pregnancy mandate the development of ZIKV vaccines, for as herd immunity wanes, the likelihood of another epidemic will increase. Several ZIKV vaccine platforms are currently in development, including inactivated viruses, live attenuated viruses, viral-vectored vaccines, and protein subunit- and nucleic acid-based formulations expressing envelope (E) proteins. These have shown protection in small and large animal models of ZIKV contamination, with several in early-phase human clinical trials10. The ability to generate large numbers of human monoclonal antibodies (mAbs) from flavivirus-infected patients has expanded our knowledge of the breadth and specificity of the human response to contamination, and allowed the delineation of antigenic sites that produce poorly or potently neutralizing antibody responses. The fusion loop epitope (FLE) is an immunodominant epitope around the E protein, and antibodies to this highly conserved region frequently show broad flavivirus cross-reactivity2C4,11. FLE-reactive antibodies generated from dengue computer virus (DENV)- or ZIKV-infected patients show low or absent neutralization of ZIKV, yet can promote antibody-dependent enhancement (ADE) of ZIKV contamination12,13. Whether pre-existing DENV immunity can primary ADE to ZIKV remains controversial; in murine models, DENV antibodies can enhance ZIKV contamination and vice versa14C16, whereas small-scale primate studies have not reproduced this17C19. Clinical studies suggest that previous DENV immunity has no detrimental effect on ZIKV contamination but may safeguard them from symptomatic JW-642 JW-642 disease20,21. Some of the best neutralizing mAbs to DENV and ZIKV identify conformational epitopes, such as E domain name III or quaternary JW-642 epitopes made up of two or more copies of E14,22C24. Given their diversity, antibodies realizing epitopes beyond the monomeric conformation of Es can show thin restriction to ZIKV or a single serotype of DENV, or can be broadly reactive. A recently explained class of human mAbs identify the E-dimer epitope (EDE)a quaternary epitope created across the interface of two E monomers arranged in a head-to-tail conformation25. The highly conserved nature of the EDE allows some EDE mAbs to neutralize all four DENV serotypes and ZIKV13,26,27, whereas most other mAbs targeting quaternary epitopes are highly specific. The mAbs HM14c10, 5J7 and ZIKV-117 bind to epitopes defined by the arrangement of two or more E dimers around the viral surface of DENV1, DENV3 and ZIKV, respectively28C30, while the epitopes targeted by the mAbs 2D22 and ZIKV-195 are contained within the E-dimer interfaces of DENV2 and ZIKV31,32. Furthermore, antibodies to complex quaternary epitopes such as the EDE are resistant to ZIKV escape mutation26, whereas computer virus escape from E domain name Ill-specific mAbs can lead to the rapid generation of viral escape mutants33,34. As both precursor membrane protein (prM) and the FLE induce the formation of poorly neutralizing yet enhancing antibodies1C3,22, we sought to develop a subunit ZIKV immunogen that would not induce these responses. We previously explained the production of stable DENV dimers, formed by the introduction of.