Analysis of antibody reactivities in lysozyme-immunized mice with peptide ligands: A model for epitope-targeted vaccine design

Many viruses evade antibody (Ab)-mediated clearance through the presentation of immunodominant, highly variable epitopes, and the masking of conserved ones. It has been proposed that vaccines incorporating peptides that cross-react with conserved epitopes could be used to target the production of Abs that protect against a range of viral isolates. For this thesis the concept of epitope-targeted vaccine design is explored with the model protein hen egg lysozyme (HEL). The goals of the work are to develop peptide markers for murine anti-HEL Abs that can also be used as immunogen to amplify Ab production against target HEL epitopes. Three types of peptides are analyzed: (i) a peptide selected from a phagedisplayed random peptide library (RPL) with the anti-HEL monoclonal Ab D1.3, (ii) peptides identified from RPLs with polyclonal (pc) Abs from the sera of HEL-immunized mice, and, (iii) peptides derived from the linear sequence of HEL. A peptide selected with Dl .3 Ab shares critical binding residues in common with its cognate discontinuous epitope but showed no reactivity with anti-HEL sera. A prime-boost immunization study, in which mice were primed with HEL to produce anti-HEL pcAbs, and boosted with the peptide to amplify the production of Dl .3 or Dl .3-like Abs, did not yield crossreactive Abs. Thus, Dl .3 Ab may not have been produced in the priming immunization because it is a rare specificity in the immune response to HEL. A peptide marker for an anti-HEL Ab commonly produced amongst mice was identified in a RPL screening with anti-HEL pcAbs. The commonly reactive peptide was also used in ELISPOT analysis to characterize epitope-specific B cell responses to HEL; the peptide-reactive cells represented less than 1% of IgG-producing B cells. The peptide was not recognized by anti-HEL IgG from rabbit, however, emphasizing the limitations of using animal models for developing epitope-targeted vaccines. Lastly, a set of 10- mer overlapping HEL peptides showed common patterns of reactivity with different murine anti- HEL sera. These findings indicate that common HEL epitopes are targeted amongst mice in the humoral immune response to HEL and that a prime-boost immunization strategy may succeed with a commonly reactive peptide marker.