Glucosidase inhibition enhances presentation of de-N-glycosylated hepatitis B virus epitopes by MHC class I in vitro and in woodchucks

In this report, the possibility of pharmacologically altering the hepatitis B virus (HBV) epitopes presented by major histocompatibility complex (MHC) class I on infected cells is demonstrated. The HBV middle envelope glycoprotein MHBs maturation appears to require calnexin mediated folding. This interaction is dependent upon glucosidases in the endoplasmic reticulum. Prevention of HBV envelope protein maturation in cultured cells with glucosidase inhibitors, such as 6-O-butanoyl castanospermine and N-nonyl deoxynorjirmycin, resulted in MHBs degradation by proteasomes. The de-N-glycosylation associated with polypeptide degradation was predicted to result in conversion of asparagine residues into aspartic acid residues. This prediction was confirmed by showing that peptides corresponding to the N-glycosylation sequons of MHBs, but with aspartic acid replacing asparagine, (a) can prime human CTLs that recognize HBV producing cells and (b) that the presentation of these envelope motifs by MHC class I is enhanced by incubation with glucosidase inhibitors. Moreover, although peripheral blood mononuclear cells isolated from woodchucks chronically infected with woodchuck hepatitis virus (WHV) and vaccinated with WHV surface antigen could be induced to recognize the natural MHBs asparagine-containing peptides, only cells isolated from glucosidase inhibitor treated animals recognized the aspartic containing peptides. Conclusion: These data suggest that pharmacological intervention with glucosidase inhibitors can alter the MHBs epitopes presented. This editing of the amino acid sequence of the polypeptide results in a new epitope, or “editope”, with possible medical significance.