This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. There currently exists a need for a vaccine that is protective against Epstein-Barr virus (EBV) infection. Such a vaccine would have a potential global impact on the incidence of infectious mononucleosis, Hodgkin's and non-Hodgkin's lymphoma, nasopharyngeal carcinoma, and lymphoproliferative syndrome. The EBV protein, gp350 has the potential to meet this need. Thus, 1) gp350 binds to the complement receptor type 2 (CD21) on B cells and follicular dendritic cells (FDCs) and thus may act as a potent adjuvant, 2) gp350 is the main target for antibody-mediated EBV neutralization, 3) multimerization of gp350 on a PS backbone will itself enhance anti-gp350 responses, and 4) gp350 is an immunogenic foreign protein that will recruit CD4+ T cell help for induction of anti-gp350 antibody. In the developing world infection with EBV, occurs in the infant population. Since, in contrast, peak EBV infection in the developed world occurs in adolescents, gp350 would also be of use as an anti-EBV vaccine in this latter population. In preliminary studies we have: 1) expressed and purified a recombinant glycosylated N-terminal 72kDa fragment of the gp350 molecule, 2) conjugated multiple copies of gp350 to pneumococcal capsular polysaccharide, serotype 14 (PPS14) [PPS14-gp350], 3) demonstrated the ability of PPS14-gp350 to specifically bind to CD21 expressed on rhesus and human, but not murine, B cells, and 4) induced boosted plasma IgG anti-PPS14 and IgG anti-gp350 antibodies in young adult rhesus monkeys following i.m. immunization with as little as 0.05 mg of PPS14-gp350 adsorbed on alum. However, careful analysis of the data, and EBV neutralizing ability of the plasma samples led us to hypothesize that a new, genetically engineered multimeric gp350 containing strong T cell epitopes would be a superior vaccine. The goal of this proposal is to extend the above studies to determine, using rhesus monkeys, the potential ability of a new genetically engineered multimeric gp350 to serve as a highly potent EBV vaccine . These pre-clinical studies will form the basis for progressing directly to human clinical trials. In this proposal we will utilize young adult rhesus monkeys to: Directly compare the ability of two distinct doses of alum-adsorbed monomeric or genetically engineered multimeric gp350 to a. elicit primary, secondary, and tertiary plasma IgG anti-gp350 titers and induce affinity maturation as determined by ELISA. b. induce EBV-neutralizing antibody as determined by both an in vitro B cell transformation and ELISA neutralization assay. c. prime specific CD4+ T cells as determined by an in vitro re-stimulation assay. Collectively, these studies are designed to establish a proof-of-principle in non-human primates, for using gp350 clinically as a protective, antibody-based vaccine for EBV relative to monomeric gp350.
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