An effective HIV vaccine capable of inducing broadly neutralizing antibodies remains a significant goal in combating the HIV/AIDS epidemic. Recent discovery of a new class of glycan-reactive broadly neutralizing antibodies (bNAbs), represented by PG9, PG16, PGT121, and PGT128, has shed light on design of novel epitope-based vaccine. These bNAbs neutralize primary HIV-1 strains with remarkable breadth and potency and appear to target glycopeptide epitopes located in the variable domains (V1/V2 or V3) of HIV-1 gp120. Mutational, biochemical and structural studies suggest that the epitopes of PG9 and PG16 are glycopeptides consist of two N-glycans (at the N160 and N156/N173 glycosylation sites) and a patch of peptide in the V1/V2 domain, while the epitopes of PGT128 appear to involve the N-glycans at the N332 and N301 sites. However, further characterization of the glycan specificity was complicated by the heterogeneity in glycosylation of gp120. We used a synthetic approach to constructing homogeneous V1V2 and V3 glycopeptides carrying defined glycans for further defining the glycan specificity, which led to the identification of a cyclic glycopeptide epitope carrying a Man5GlcNAc2 glycan at N160 and a sialylated N-glycan at N156/N173 site as the minimal epitopes for PG9/PG16. We also designed and synthesized cyclic V3 glycopeptides that confirm the essence of a Man9/Man8GlcNAc2 glycan at the N332 site for PGT128 recognition. With the ability to reconstitute these glycopeptide epitopes via synthesis, we propose to design novel immunogens and hypothesize that displaying the glycopeptide epitopes on bacteriophage Qb (a well-characterized virus-like particle) in a highly ordered repetitive pattern will lead to immunogens capable of eliciting glycopeptide-specific neutralizing antibodies. We will test this hypothesis through pursuing three specific aims.
Aim 1 is to design and synthesize Qb-based immunogens carrying polyvalent V1V2 and V3 glycopeptide epitopes. Construction of the immunogens includes chemoenzymatic synthesis of alkyne-glycopeptides and their subsequent conjugation with azide-tagged Qb via the copper-catalyzed azide-alkyne cycloaddition (CuAAC) click reaction;
Aim 2 is to evaluate the antigenicity of the synthetic immunogens by antibody binding studies;
and Aim 3 is to evaluate the immunogenicity of the synthetic vaccines in rabbits. The anti-sera from immunized rabbits are analyzed by ELISAs and viral neutralization assays. These experiments will assess the magnitude, breadth and duration of neutralizing antibody responses from immunizations with the glycopeptide epitope-based immunogens. This study is likely to yield important new insights in the design of a truly effective HIV-1 vaccine.
A truly effective vaccine is the best hope to combat the global HIV/AIDS epidemic that threatens human health and social stability. The proposed research aims to design and test novel immunogens based on the newly identified glycopeptide epitopes of broadly neutralizing antibodies, which will provide important new information to speed up HIV vaccine development.
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