Towards a solution for hepatitis C virus hypervariability: mimotopes of the hypervariable region 1can induce antibodies cross-reacting with a large number of viral variants

Abstract

The hypervariable region 1 (HVR1) of the putative envelope protein E2 of hepatitis C virus (HCV) is the most variable antigenic fragment in the whole viral genome and is mainly responsible for the large interand intra-individual heterogeneity of the infecting virus. It contains a principal neutralization epitope and has been proposed as the major player in the mechanism of escape from host immune response. Since anti-HVR1 antibodies are the only species shown to possess protective activity up to date, developing an effective prevention therapy is a very difficult task. We have approached the problem of HVR1 variability by deriving a consensus profile from Ͼ200 HVR1 sequences from different viral isolates and used it as a template to generate a vast repertoire of synthetic HVR1 surrogates displayed on M13 bacteriophage. This library was affinity selected using many different sera from infected patients. Phages were identified which react very frequently with patients' sera and bind serum antibodies that cross-react with a large panel of HVR1 peptides derived from natural HCV variants. When injected into experimental animals, the 'mimotopes' with the highest cross-reactivity induced antibodies which recognized the same panel of natural HVR1 variants. In these mimotopes we identified a sequence pattern responsible for the observed crossreactivity. These data may hold the key for future development of a prophylactic vaccine against HCV.

Monoclonal Antibodies with Broad Specificity for Hepatitis C Virus Hypervariable Region 1 Variants Can Recognize Viral Particles

“…HVR1 surrogate peptides (mimotopes) were obtained by constructing an HVR1 library by back-translation of an HVR1 consensus profile, accounting for ϳ80% of the total sequence variability, into the corresponding nucleotide sequence (15). A degenerate synthetic oligonucleotide was cloned as a fusion to the gene encoding for the major coat protein (pVIII) in a phagemid vector for display on M13 bacteriophage.…”

“…Phage were subjected to two rounds of affinity selection using sera from anti-HCV-positive patients. A number of clones reacting exclusively with HCV sera were identified, with the best clones reacting with ϳ80% of the sera (15). Nine HVR1 mimotopes were synthesized as multiple antigenic peptides (MAPs) (16) and used for immunization as described below (Table I).…”

“…To examine binding to naturally occurring HVR1 isolates, two panels of previously validated biotinylated peptides of 16-and 27-aa residues (9,15) were used on streptavidin-coated ELISA plates. Similarly, to determine the fine specificity of murine hybridomas, 14 overlapping biotinylated 14-mers scaled by one residue and covering the entire HVR1 region (amino acids 384 -410) were synthesized based on the sequence of the HVR1 consensus profile reported previously (15).…”

“…Similarly, to determine the fine specificity of murine hybridomas, 14 overlapping biotinylated 14-mers scaled by one residue and covering the entire HVR1 region (amino acids 384 -410) were synthesized based on the sequence of the HVR1 consensus profile reported previously (15).…”

“…Several studies identified binding sites for allegedly neutralizing Ab in this region (13,14), and it is therefore conceivable that HVR1 expressed on integral HCV particles could represent a major target for immune responses that may play an important role in the outcome of HCV infection. In this study, we have generated mAbs by immunizing mice with peptides derived from an HVR1 consensus profile, accounting for ϳ80% of the total sequence variability (15), and we characterized such mAbs in terms of fine specificity and ability to bind to HVR1 sequences expressed in the context of correctly folded chimeric E2-HVR1 proteins and on recombinant and natural bona fide viral particles. The long-term objective was to analyze in detail the features of the humoral immune responses elicited by peptides that are thought to represent efficient antigenic and immunogenic mimics of natural HVR1 sequences (15) and that may be considered as potential components of a synthetic vaccine inducing broad specificity for several HCV variants.…”