A Rational Strategy to Design Multiepitope Immunogens Based on Multiple Th Lymphocyte Epitopes

Abstract

Four HLA-DR-restricted HIV-derived Th lymphocyte (HTL) epitopes cross-reactive with the murine I-Ab class II molecule were used to evaluate different vaccine design strategies to simultaneously induce multiple HTL responses. All four epitopes were immunogenic in H-2b mice, demonstrating the feasibility of murine models to evaluate epitope-based vaccines destined for human use. Immunization with a pool of peptides induced responses against all four epitopes; illustrating immunodominance does not prevent the induction of balanced multispecific responses. When different delivery systems were evaluated, a multiple Ag peptide construct was found to be less efficient than a linear polypeptide encompassing all four epitopes. Further characterization of linear polypeptide revealed that the sequential arrangement of the epitopes created a junctional epitope with high affinity class II binding. Disruption of this junctional epitope through the introduction of a GPGPG spacer restored the immunogenicity against all four epitopes. Finally, we demonstrate that a GPGPG spacer construct can be used to induce HTL responses by either polypeptide or DNA immunization, highlighting the flexibility of the approach.

Development, Characterization and Immunogenicity of a Multi-Stage, MultivalentPlasmodium falciparumVaccine Antigen (FALVAC-1A) Expressed inEscherichia coli

“…Computer modeling studies suggest that several near natural conformations are potentially available to the FALVAC-1A vaccine antigen. We like to think that the spacers (especially the GPGPG sequences which induce 180°turns in the polypeptide chain), 25 which were not included in FALVAC-1, account for the stability of FALVAC-1A in comparison to FALVAC-1. But without a full 3-dimensional structural elucidation of FALVAC-1A, it is Figure 2.…”

“…Five of these were GPGPG spacer sequences, which tend to induce 180°reverse turns in a polypeptide structure, to aid folding to a more compact form. 25 Five KG spacers were also inserted to allow for flexibility of certain regions of the structure, and 2 KAA spacers immediately after the carboxy termini of the two LSA-1 CTL epitopes to aid in their processing and presentation in the Th-1 pathway. 26 The residues Met-Ala were placed on the N-terminus to provide the start signal and to facilitate cloning; six histidines were placed on the carboxy terminus to facilitate purification by metal ion chromatography.…”

Conformation-sensitive Antibodies against Alzheimer Amyloid-β by Immunization with a Thioredoxin-constrained B-cell Epitope Peptide

“…1B). Instrumental to the production of properly assembled A␤15 multimers were the directionality and in-frame fusion capability of the unique CpoI site present within the Trx active site loop region as well as the incorporation into A␤15 DNA of a terminal sequence coding for an intervening Gly-Gly-Pro linker, thus also preventing the formation of junctional epitopes (52). A fourth construct (TrxA␤42) bearing a single copy of the fulllength A␤42 peptide was prepared in a similar way (see "Experimental Procedures" for details).…”

Frontiers in peptide–MHC class II multimer technology

“…Are there also fundamental differences between MHC class I and II molecules in the strength of their binding interactions with peptides or TCRs? Binding studies of peptides with isolated class II MHC molecules indicate that, at least for certain strong-binding peptides, the IC50 (concentration needed to competitively inhibit 50% of the binding of a standard peptide) can be in the nanomolar range (L. Teyton, La Jolla, CA), similar to other reported pMHC class I and II interactions 10,11 . Dissociation constants of TCR-MHC class II interactions in the range of 10 × 10 -6 -35 × 10 -6 M (L. Stern, Cambridge, MA; E. Ward, Dallas, TX) are also similar to those reported for MHC class I-TCR binding 12 .…”