The goal of this research proposal is to develop a novel means for obtaining peptides that would serve as vaccine leads against infection by HIV-1. Our strategy begins with a repertoire of antibodies that is specifically designed to mimic the structures of Abs that are elicited by immunization with peptide conjugates. Selected human germline V genes will encode this repertoire of """"""""anti-peptide-like"""""""" Abs, so that it will mimic a subset of the """"""""natural"""""""" repertoire that is typically elicited against peptide-immunogens. The repertoire will be expressed and displayed as Fab-fusions to filamentous phage, and will be screened against recombinant HIV-1 envelope protein to find binding Fabs. These Fabs, in turn, should be predisposed to bind peptides, and, on immunization, they should be elicited by peptide-conjugate vaccines. Thus, when such Fabs are used to screen a peptide library, """"""""immunogenic-mimic"""""""" peptides should be found that will both cross-react with the Fabs and elicit target-antigen-binding Abs when used as immunogens. We propose to apply these concepts in our ongoing search for peptide leads for a vaccine that will produce broadly neutralizing Abs against HIV-1, following the specific aims: 1. Construct a phage-displayed Fab library whose design is based on human germline V-genes that appear in the """"""""anti-peptide"""""""" antibodies. 2. Validate the phage-displayed Fab library for (i) its ability to produce binding Fabs against a panel of """"""""target"""""""" peptide and protein antigens; (ii) the ability of those antigen-binding Fabs to bind to peptides, and (iii) the ability of the Fab-binding peptides to elicit Ab responses that bind to the corresponding target antigen. 3. Screen the Fab library with recombinant HIV- 1 envelope protein and intact HIV-1 particles to obtain specific HIV-1-binding Fabs; and characterize the Fabs for their ability to bind to and neutralize a panel of HIV-1 strains and primary isolates. 4. Screen a panel of 16 phage-displayed peptide libraries with the HIV- 1 binding Fabs to obtain peptide ligands. 5. Determine whether the peptide ligands elicit immune responses that recognize Env proteins and neutralize HIV-1. By this means, we hope to develop a new means of targeting peptide vaccines against neutralizing sites on HIV- l.
