In vitro binding analysis of hepatitis B virus preS-derived putative helper T-cell epitopes to MHC class II molecules using stable HLA-DRB1*0405/-DRA*0101 transfected cells

In designing epitope-based vaccines, the inclusion of a helper T-lymphocyte (HTL) epitope is necessary to elicit both humoral and cellular immune resp onses. Whereas the preS region of the hepatitis B virus (HBV) surface antig en is well-known to raise protective immunity, the epitopes for activating HTLs are poorly characterized. In an attempt to identify such epitopes, the HBV-preS region was screened for peptide sequences with HLA-DR4 binding mo tifs, and putative HTL candidate peptides were synthesized in a biotinylate d form. Using L929 mouse fibroblasts stably transfected with HLA-DRB1*0405 and HLA-DRA*0101 cDNA, specific binding of the peptides was then detected u sing fluorescence-conjugated streptavidin. The cell-surface expression of H LA-DR molecules on transfectants was confirmed by confocal microscopy, and quantitative analysis of candidate peptide binding was performed by fluores cence activated cell sorting. Among eight preS-derived peptides, three cand idate peptides-namely preS1(23-33), preS1(62-72), and preS1(76-86)-showed g ood binding characteristics to HLA-DR4 molecules, among which the preS1(23- 33) epitope was regarded as the most promising HTL epitope. Further studies with these candidate HTL stimulatory peptides will show their ability to a ctivate the human immune system against HBV.