Humanized Antibodies Derived from Chimpanzee Fabs that N

Lai, Ching

Abstract

Chimpanzee monoclonal antibodies that efficiently neutralize dengue viruses, Japanese encephalitis virus, or tick-borne encephalitis virus have been identified from panels of Fab antibody fragments recovered by repertoire cloning. Humanized antibodies derived from selected Fabs proved to be highly efficient for neutralization of these flaviviruses in vitro. The four dengue virus serotypes are most important in terms of human morbidity and geographic distribution. Passive immunization by transfer of monoclonal antibody represents an attractive alternative to a vaccine against dengue or other flaviviruses emerging in human populations. Monoclonal antibodies are also valuable for analysis of the virus structure, replication and pathogenesis. We have identified and characterized a humanized monoclonal antibody cross-reactive to dengue and other major flaviviruses and a highly potent neutralizing antibody specific for dengue type 4 virus (DENV-4). DENV-4 challenge models have been established in mice and in primates for analysis of protection following passive transfer of antibody. Using the primate model, established for passive transfer against dengue for the first time, we have demonstrated proof of principle for use of humanized antibody for protection against dengue infection. We have analyzed the epitope determinants of these selected chimpanzee monoclonal antibodies. The epitope determinants of the cross-reactive antibody have been localized to amino acids within the flavivirus conserved fusion peptide in domain II of the E glycoprotein. The epitope location of the DENV-4-specific monoclonal antibody in domain I has also been determined. To validate our approach to protection by passive immunization, studies were performed to address the antibody safety issue concerning the antibody dependent enhancement (ADE), a phenomenon believed to be the underlying mechanism responsible for severe dengue frequently associated with secondary dengue infections. Taking advantage of the DENV-4 challenge system and the data from in vitro ADE studies, we have demonstrated the ADE activity of dengue replication in primates. Importantly, we have also identified mutations, including those known to be involved in IgG antibody-FcrR binding interactions, for possible use to ablate the ADE activity of promising antibodies. As an extension, we have focused on antibodies against Japanese encephalitis virus and tick-borne encephalitis virus, both of which continue to cause serious illnesses in many parts of the world. Chimpanzee Fabs and their derived humanized antibodies effectively neutralizing Japanese encephalitis virus or tick-borne encephalitis virus have been obtained and characterized. These studies set the stage for further development of the passive immunization strategy for prevention of dengue and other flavivirus diseases.