A novel method for enhancement of peptide vaccination utilizing T-cell epitopes from conventional vaccines.

Peptide vaccines have two fundamental weak points, namely low antigenicity and MHC-restriction. In our previous study, we proposed the design of vaccine peptide to overcome these weakpoints. The vaccine was constructed in the following order, N-terminal, Arg-Gly-Asp (RGD), T-cell epitope peptide, di-lysine linker (KK) to B-cell epitope peptide. Although the vaccine peptide can basically induce B-cell epitope peptide specific antibodies to the host without immune adjuvants via intraperitoneal, subcutaneous and intranasal administration, some peptide antigens require adjuvants for antibody induction. In this study, we propose a novel protocol to enhance the immunogenicity of the peptide utilizing the host immune response to a conventional toxoid vaccine, which are lymphocyte activities to the T-cell epitope peptide. We selected multiagretope-type T-cell epitopes from diphtheria toxoid, a conventional vaccine antigen, and a part of amyloid-beta peptide (Aß) as a B-cell epitope. The conventional toxoid vaccine was immunized before the peptide immunization. Using this protocol, we succeeded in the enhancement of the anti-Aß antibodies induction by intranasal immunization without any immune adjuvants in C57BL/6 and Balb/c mice. Furthermore, the vaccine peptide induced the transformation of peripheral blood lymphocytes collected from healthy volunteers carrying immunities to diphtheria toxoid. These results suggested that our peptide vaccines with the novel protocol would provide an effective method for antibody induction.

Production of Fab fragment corresponding to surface protein antigen of Streptococcus mutans serotype c-derived peptide by Escherichia coli and cultured tobacco cells.

The cDNA of a mouse Fab fragment was cloned from a hybridoma cell line that produces a mouse monoclonal antibody, KH5, that reacts with the peptide fragment of the surface protein antigen of Streptococcus mutans serotype c (PAc). After transfection with cDNA, recombinant Fab fragments were produced by Escherichia coli (T15 Fab) and cultured tobacco cells (X253 and X262 Fabs). The antipeptide activities of T15 and X253 were similar to that of KH5. X253 was secreted into the culture media, which had a specific affinity for the PAc peptide.

An ingenious design for peptide vaccines.

For humoral immunization, it may be possible to make effective and safe peptide vaccines for various diseases by selection of proper B-cell epitopes. However, a lack of T-cell epitopes on short peptides, such as those associated with major histocompatibility complex (MHC)-restriction, is a major problem for peptide vaccine development. We propose a solution for the design of peptide vaccines that involves induction of broadly reactive T-cell epitopes via agretopes. The strategy involves positioning multi-agretope type peptides on the N-terminal side of a di-lysine linker and B-cell epitopes on the C-terminal side. The addition of the arginine-glysine-aspartate (RGD)-motif to the N terminus of the peptide enhances its immunogenicity, and enables nasal immunization without adjuvants.

Oral streptococci exhibit diverse susceptibility to human beta-defensin-2: antimicrobial effects of hBD-2 on oral streptococci.

We examined the antimicrobial effects of human beta-defensin-2 (hBD-2) on 17 species of oral streptococci to investigate the involvement of antimicrobial peptide activity in oral microflora development and the clinical use of the antimicrobial peptide for oral microflora control. Oral streptococci exhibit diverse levels of susceptibility to human beta-defensin-2 (hBD-2). Two major cariogenic bacterial species, Streptococcus mutans ( S. mutans) and S. sobrinus, were found to be susceptible to the peptide, indicating that it is a potential therapeutic agent for preventing dental caries. S. mitis exhibited the lowest susceptibility to the peptide. S. mitis is a major indigenous bacterium in the oral microflora, and our results suggest that it might possess a certain resistance mechanism against hBD-2.

RGD motif enhances immunogenicity and adjuvanicity of peptide antigens following intranasal immunization.

The use of peptides for various aspects of medical science has been a significant advance. Peptide-based vaccines are promising, but weak immunogenic potency is impeding the clinical application. We have remarkably enhanced the immunogenicity of peptide antigens by addition of motifs that bind to cell attachment proteins, such as arginine-glysine-aspartate (RGD), to the amino acid sequence. The modified peptides induced antigen-specific serum antibodies by intranasal immunization without adjuvants. RGD, an integrin-binding motif was the strongest, among several molecules tested in this experiment, giving an average of 10 times enhancement of antibody titers when incorporated into several peptide antigens. The peptides also acted as an efficient adjuvant following the intranasal immunization with protein antigens. Our data support the feasibility of developing peptide vaccines and peptide adjuvants for intranasal vaccination.