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Wei S, Lei Y, Yang J, Wang X, Shu F, Wei X, Lin F, Li B, Cui Y, Zhang H, Wei S. Neutralization effects of antibody elicited by chimeric HBV S antigen viral-like particles presenting HCV neutralization epitopes. Vaccine 2018; 36:2273-2281. [PMID: 29576303 DOI: 10.1016/j.vaccine.2018.03.036] [Citation(s) in RCA: 17] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/25/2017] [Revised: 02/23/2018] [Accepted: 03/14/2018] [Indexed: 01/29/2023]
Abstract
Hepatitis C virus (HCV) infection is a major public health problem despite effectual direct-acting antivirals (DAAs) therapy. Development of a prophylactic vaccine is essential to block spread of HCV infection. The HBV small surface antigen (HBsAg-S) can self-assemble into virus-like particles (VLPs), has higher immunogenicity and is used as a vaccine against HBV infections. Chimeric HBsAg-S proteins with foreign epitopes allow VLP formation and induce the specific humoral and cellular immune responses against the foreign proteins. In this study, we investigated the immune responses induced by chimeric VLPs with HCV neutralizing epitopes and HBV S antigen in mice. The chimeric HCV-HBV VLPs expressing neutralizing epitopes were prepared and purified. BALB/c mice were immunized with purified chimeric VLPs and the serum neutralizing antibodies were analyzed. We found that these chimeric VLPs induced neutralizing antibodies against HCV in mice. Additionally, the murine serum neutralized infections with HCV pseudoparticles and cell-cultured viruses derived from different heterologous 1a, 1b and 2a genotypes. We also found that immunization with chimeric VLPs induced anti-HBsAg antibodies. This study provides a novel strategy for development of a HCV prophylactic neutralizing epitope vaccine and a HCV-HBV bivalent prophylactic vaccine.
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Affiliation(s)
- Sanhua Wei
- Department of Clinical Laboratory and Research Center, Tangdu Hospital, The Fourth Military Medical University, No. 569 Xinsi Road, Xi'an, Shaanxi 710038, China
| | - Yingfeng Lei
- Department of Microbiology, The Fourth Military Medical University, No. 17 West Road, Xi'an, Shaanxi 710032, China
| | - Jie Yang
- Department of Nephrology, Tangdu Hospital, The Fourth Military Medical University, No. 569 Xinsi Road, Xi'an, Shaanxi 710038, China
| | - Xiaoyan Wang
- Department of Clinical Laboratory and Research Center, Tangdu Hospital, The Fourth Military Medical University, No. 569 Xinsi Road, Xi'an, Shaanxi 710038, China
| | - Fang Shu
- Department of Clinical Laboratory, Xi'an Third Hospital, No. 10 Eastern Section of The Third FengCheng Rd., WeiYang District, Xi'an, Shaanxi 710018, China
| | - Xin Wei
- Department of Infectious Disease, Tangdu Hospital, The Fourth Military Medical University, No. 569 Xinsi Road, Xi'an, Shaanxi 710038, China
| | - Fang Lin
- Department of Clinical Laboratory and Research Center, Tangdu Hospital, The Fourth Military Medical University, No. 569 Xinsi Road, Xi'an, Shaanxi 710038, China
| | - Bin Li
- Department of Clinical Laboratory and Research Center, Tangdu Hospital, The Fourth Military Medical University, No. 569 Xinsi Road, Xi'an, Shaanxi 710038, China
| | - Ying Cui
- Department of Clinical Laboratory and Research Center, Tangdu Hospital, The Fourth Military Medical University, No. 569 Xinsi Road, Xi'an, Shaanxi 710038, China
| | - Hai Zhang
- Laboratory Animal Research Center, The Fourth Military Medical University, No. 17 West Road, Xi'an, Shaanxi 710032, China.
| | - Sanhua Wei
- Department of Clinical Laboratory and Research Center, Tangdu Hospital, The Fourth Military Medical University, No. 569 Xinsi Road, Xi'an, Shaanxi 710038, China.
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Gupte GM, Arankalle VA. Evaluation of the immunogenicity of liposome encapsulated HVR1 and NS3 regions of genotype 3 HCV, either singly or in combination. Virol J 2012; 9:74. [PMID: 22452828 PMCID: PMC3349533 DOI: 10.1186/1743-422x-9-74] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/12/2011] [Accepted: 03/27/2012] [Indexed: 01/15/2023] Open
Abstract
Background Hepatitis C virus displays a high rate of mutation and exists as a quasispecies in infected patients. In the absence of an effective universal vaccine, genotype-specific vaccine development represents an alternative. We have attempted to develop a genotype 3 based, liposome encapsulated HCV vaccine with hypervariable region-1 (HVR1) and non-structural region-3 (NS3) components. Results HCV RNA extracted from serum samples of 49 chronically infected patients was PCR amplified to obtain HVR1 region. These amplified products were cloned to obtain 20 clones per sample in order to identify the quasispecies pattern. The HVR1 consensus sequence, along with three variants was reverse transcribed to obtain peptides. The peptides were checked for immunoreactivity individually, as a pool or as a single peptide tetramer interspersed with four glycine residues. Anti-HCV positivity varied from 42.6% (tetramer) to 92.2% (variant-4) when 115 anti-HCV positive sera representing genotypes 1, 3, 4 and 6 were screened. All the 95 anti-HCV negatives were scored negative by all antigens. Mice were immunized with different liposome encapsulated or Al(OH)3 adjuvanted formulations of HVR1 variants and recombinant NS3 protein, and monitored for anti-HVR1 and anti-NS3 antibody titres, IgG isotypes and antigen specific cytokine levels. A balanced Th1/Th2 isotyping response with high antibody titres was observed in most of the liposome encapsulated antigen groups. The effect of liposomes and aluminium hydroxide on the expression of immune response genes was studied using Taqman Low Density Array. Both Th1 (IFN-gamma, Il18) and Th2 (Il4) genes were up regulated in the liposome encapsulated HVR1 variant pool-NS3 combination group. In-vitro binding of the virus to anti-HVR1 antibodies was demonstrated. Conclusion The optimum immunogen was identified to be combination of peptides of HVR1 consensus sequence and its variants along with pNS3 encapsulated in liposomes, which could generate both cellular and humoral immune responses in mice deserving further evaluation in a suitable cell culture system/non-human primate model.
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Affiliation(s)
- Gouri M Gupte
- Hepatitis Division, National Institute of Virology, Microbial Containment Complex, Sus Road, Pashan, Pune, India 411021
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Wang Y, Keck ZY, Foung SKH. Neutralizing antibody response to hepatitis C virus. Viruses 2011; 3:2127-45. [PMID: 22163337 PMCID: PMC3230844 DOI: 10.3390/v3112127] [Citation(s) in RCA: 57] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/14/2011] [Revised: 10/18/2011] [Accepted: 10/22/2011] [Indexed: 12/14/2022] Open
Abstract
A critical first step in a "rational vaccine design" approach for hepatitis C virus (HCV) is to identify the most relevant mechanisms of immune protection. Emerging evidence provides support for a protective role of virus neutralizing antibodies, and the ability of the B cell response to modify the course of acute HCV infection. This has been made possible by the development of in vitro cell culture models, based on HCV retroviral pseudotype particles expressing E1E2 and infectious cell culture-derived HCV virions, and small animal models that are robust tools in studies of antibody-mediated virus neutralization. This review is focused on the immunogenic determinants on the E2 glycoprotein mediating virus neutralization and the pathways in which the virus is able to escape from immune containment. Encouraging findings from recent studies provide support for the existence of broadly neutralization antibodies that are not associated with virus escape. The identification of conserved epitopes mediating virus neutralization that are not associated with virus escape will facilitate the design of a vaccine immunogen capable of eliciting broadly neutralizing antibodies against this highly diverse virus.
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Affiliation(s)
- Yong Wang
- Department of Pathology, School of Medicine, Stanford University, Stanford, CA 94305, USA.
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Wei SH, Yin W, An QX, Lei YF, Hu XB, Yang J, Lu X, Zhang H, Xu ZK. A novel hepatitis C virus vaccine approach using recombinant Bacillus Calmette-Guerin expressing multi-epitope antigen. Arch Virol 2008; 153:1021-9. [PMID: 18421415 DOI: 10.1007/s00705-008-0082-1] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/25/2007] [Accepted: 02/27/2008] [Indexed: 10/22/2022]
Abstract
Hepatitis C virus (HCV) is a major cause of liver disease worldwide. HCV infection is associated with high morbidity and has become a major problem in public health. Until now, there has been no effective prophylactic or therapeutic vaccine. BCG, a live vaccine typically used for tuberculosis prevention, has been increasingly utilized as a vector for the expression of recombinant proteins that will induce specific humoral and cellular immune responses. In this study, recombinant BCG (rBCG) was engineered to express a HCV multi-epitope antigen CtEm, and HLA-A2.1 transgenic mice were immunized with rBCG-CtEm. High levels of specific anti-HCV antibodies targeted to mimotopes of HVR1 were detected in the serum. HCV-specific lymphocyte proliferation assay, cytokine determination and cytotoxicity assay indicated that HCV epitope-specific cellular immune responses were elicited in vitro. The rBCG-CtEm immunization conferred protection against infection with the recombinant vaccinia virus (rVV-HCV-CNS) in vivo. These results suggest that rBCG expressing multi-epitope antigen may serve as an effective vaccine against HCV infection.
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Affiliation(s)
- S-H Wei
- The State Key Discipline and Department of Microbiology, Fourth Military Medical University of PLA, 17 Changlexi Road, 710032 Xi'an, China
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Deng Q, Zhai JW, Michel ML, Zhang J, Qin J, Kong YY, Zhang XX, Budkowska A, Tiollais P, Wang Y, Xie YH. Identification and characterization of peptides that interact with hepatitis B virus via the putative receptor binding site. J Virol 2006; 81:4244-54. [PMID: 17192308 PMCID: PMC1866126 DOI: 10.1128/jvi.01270-06] [Citation(s) in RCA: 36] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/23/2022] Open
Abstract
A direct involvement of the PreS domain of the hepatitis B virus (HBV) large envelope protein, and in particular amino acid residues 21 to 47, in virus attachment to hepatocytes has been suggested by many previous studies. Several PreS-interacting proteins have been identified. However, they share few common sequence motifs, and a bona fide cellular receptor for HBV remains elusive. In this study, we aimed to identify PreS-interacting motifs and to search for novel HBV-interacting proteins and the long-sought receptor. PreS fusion proteins were used as baits to screen a phage display library of random peptides. A group of PreS-binding peptides were obtained. These peptides could bind to amino acids 21 to 47 of PreS1 and shared a linear motif (W1T2X3W4W5) sufficient for binding specifically to PreS and viral particles. Several human proteins with such a motif were identified through BLAST search. Analysis of their biochemical and structural properties suggested that lipoprotein lipase (LPL), a key enzyme in lipoprotein metabolism, might interact with PreS and HBV particles. The interaction of HBV with LPL was demonstrated by in vitro binding, virus capture, and cell attachment assays. These findings suggest that LPL may play a role in the initiation of HBV infection. Identification of peptides and protein ligands corresponding to LPL that bind to the HBV envelope will offer new therapeutic strategies against HBV infection.
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Affiliation(s)
- Qiang Deng
- State Key Laboratory of Molecular Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes of Biological Sciences, Chinese Academy of Sciences, and Ruijin Hospital, Department of Infectious Diseases, Shanghai, China
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Abstract
Systematic approaches to directed evolution of proteins have been documented since the 1970s. The ability to recruit new protein functions arises from the considerable substrate ambiguity of many proteins. The substrate ambiguity of a protein can be interpreted as the evolutionary potential that allows a protein to acquire new specificities through mutation or to regain function via mutations that differ from the original protein sequence. All organisms have evolutionarily exploited this substrate ambiguity. When exploited in a laboratory under controlled mutagenesis and selection, it enables a protein to "evolve" in desired directions. One of the most effective strategies in directed protein evolution is to gradually accumulate mutations, either sequentially or by recombination, while applying selective pressure. This is typically achieved by the generation of libraries of mutants followed by efficient screening of these libraries for targeted functions and subsequent repetition of the process using improved mutants from the previous screening. Here we review some of the successful strategies in creating protein diversity and the more recent progress in directed protein evolution in a wide range of scientific disciplines and its impacts in chemical, pharmaceutical, and agricultural sciences.
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Affiliation(s)
- Ling Yuan
- Department of Plant and Soil Sciences, and Kentucky Tobacco Research and Development Center, University of Kentucky, Lexington, KY 40546, USA.
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Deng Q, Zhuang M, Kong YY, Xie YH, Wang Y. Screening for PreS specific binding ligands with a phage displayed peptides library. World J Gastroenterol 2005; 11:4018-23. [PMID: 15996026 PMCID: PMC4502097 DOI: 10.3748/wjg.v11.i26.4018] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM: To construct a random peptide phage display library and search for peptides that specifically bind to the PreS region of hepatitis B virus (HBV).
METHODS: A phage display vector, pFuse8, based on the gene 8 product (pVIII) of M13 phage was made and used to construct a random peptide library. E.coli derived thioredoxin-PreS was purified with Thio-bond beads, and exploited as the bait protein for library screening. Five rounds of bio-panning were performed. The PreS-binding specificities of enriched phages were characterized with phage ELISA assay.
RESULTS: A phage display vector was successfully constructed as demonstrated to present a pVIII fused HBV PreS1 epitope on the phage surface with a high efficiency. A cysteine confined random peptide library was constructed containing independent clones exceeding 5±108 clone forming unit (CFU). A pool of phages showing a PreS-binding specificity was obtained after the screening against thio-PreS with an enrichment of approximately 400 times. Five phages with high PreS-binding specificities were selected and characterized. Sequences of the peptides displayed on these phages were determined.
CONCLUSION: A phage library has been constructed, with random peptides displaying as pVIII-fusion proteins. Specific PreS-binding peptides have been obtained, which may be useful for developing antivirals against HBV infection.
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Affiliation(s)
- Qiang Deng
- Institute of Biochemistry and Cell Biology, 320 Yue-Yang Road, Shanghai 200031, China
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