Expression of enhancing-activity-free neutralizing antibody against dengue type 1 virus in plasmid-inoculated mice

Abstract

BackgroundMost candidate dengue vaccines currently under development induce neutralizing antibodies, which are considered important for immunoprotection. However, the concomitant induction of infection-enhancing antibodies is an unavoidable concern. In contrast, a neutralizing antibody developed for passive immunotherapy has been engineered to eliminate its enhancing activity. Therefore, a strategy for the long-term expression of enhancing-activity-free neutralizing antibodies may resolve this concern. MethodsA mouse monoclonal antibody, 7F4, of the IgG3 subclass and with no detectable enhancing activity, was selected as the model neutralizing antibody to evaluate the potential of this strategy. Equal amounts of commercial vector (pFUSE)-based plasmids containing 7F4 heavy (H)- or light (L)-chain variable region genes were mixed and used for the cotransfection of 293T cells and co-delivery into ICR and BALB/c mice. The recombinant plasmids were designed to express IgG2b or IgG3 subclass antibodies (p7F4G2b or p7F4G3, respectively). Results293T cells transfected with 2μg of p7F4G2b or p7F4G3 produced approximately 15,000 or 800ng/ml IgG in the culture fluids, respectively. The dose is expressed as the total amount of H- and L-chain plasmids. Neutralizing antibody was detected dose-dependently in ICR mice inoculated with 50–200μg of p7F4G2b. A 1:2 dilution of sera from ICR and BALB/c mice inoculated with 100μg of p7F4G3 showed average plaque reduction levels of >70% on day 3 and >90% on days 5–9. BALB/c mice maintained detectable neutralizing antibody for at least 3 months. The neutralizing antibody expressed by p7F4G3 in mice showed no enhancing activity. DiscussionAlthough the expression of neutralizing antibodies from immunoglobulin genes is a type of passive immunization, its durability can be utilized as a dengue vaccine strategy. This “proof-of-concept” study using a mouse model demonstrates that the enhancing-activity-free characteristic of this strategy augurs well for dengue vaccine development, although further improvement is required.