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Design and Development of Modified mRNA Encoding Core Antigen of Hepatitis C Virus: a Possible Application in Vaccine Production

ABSTRACT: Background:Hepatitis C virus (HCV) is a ?blood-borne pathogen, resulting in liver cirrhosis and liver cancer. Despite of many efforts in development of treatments for HCV, no vaccine has been licensed yet. The purpose of this study was ?to design and prepare a specific mRNA, without 5' cap and poly (A) tail transcribed in vitro capable of coding core protein and also to determine its functionality. Methods:Candidate mRNA was prepared by in vitro transcription of the designed construct consisting of ??5? and 3? untranslated regions of heat shock proteins 70 (hsp70) mRNA, T7 promoter, internal ribosome entry site (IRES) sequences of eIF4G related to human dendritic cells (DCs), and the ?Core gene of HCV. To design the modified mRNA, the ??5' cap and poly (A) tail structures were not considered. DCs were transfected by in vitro-transcribed messenger RNA (IVT-mRNA) and the expressions of green fluorescent protein (GFP), and Core genes were determined by microscopic examination and Western blotting assay. Results:Cell transfection results showed that despite the absence of ??5' cap and poly (A) tail, the structure of the mRNA ?was stable. Moreover, the successful expressions of GFP and Core genes were achieved. Conclusion:Our findings indicated the effectiveness of a designed IVT-mRNA harboring the Core gene of HCV in transfecting and expressing the antigens in DCs. Considering the simple and efficient protocol for the preparation of this IVT-mRNA and its effectiveness in expressing the gene that it carries, this IVT-mRNA could be suitable for development of an RNA vaccine against HCV.

SUBMITTER: Sharifnia Z

PROVIDER: S-EPMC6305823 | biostudies-literature | 2019 Jan

REPOSITORIES: biostudies-literature

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Design and Development of Modified mRNA Encoding Core Antigen of Hepatitis C Virus: a Possible Application in Vaccine Production

Sharifnia Zarin Z Bandehpour Mojgan M Kazemi Bahram B Zarghami Nosratollah N

Iranian biomedical journal 20180730 1

<h4>Background</h4>Hepatitis C virus (HCV) is a ‎blood-borne pathogen, resulting in liver cirrhosis and liver cancer. Despite of many efforts in development of treatments for HCV, no vaccine has been licensed yet. The purpose of this study was ‎to design and prepare a specific mRNA, without 5' cap and poly (A) tail transcribed in vitro capable of coding core protein and also to determine its functionality.<h4>Methods</h4>Candidate mRNA was prepared by in vitro transcription of the designed const ...[more]

PMID: 30056690

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Project description:Background & aimsParticulate hepatitis B core antigen (HBcoreAg) is a potent immunogen used as a vaccine carrier platform. HBcoreAg produced in E. coli encapsidates random bacterial RNA (bRNA). Using the heterologous protein-prime, viral-vector-boost therapeutic hepatitis B vaccine TherVacB, we compared the properties of different HBcoreAg forms. We explored how the content of HBcoreAg modulates antigen stability, immunogenicity, and antiviral efficacy.MethodsbRNA was removed from HBcoreAg by capsid disassembly, followed by reassembly in the absence or presence of specific nucleic acid-based adjuvants poly I:C or CpG. The morphology and structure of empty, bRNA-containing and adjuvant-loaded HBcoreAg were monitored by electron microscopy and nuclear magnetic resonance spectroscopy. Empty, bRNA-containing or adjuvant-loaded HBcoreAg were applied together with HBsAg and with or without nucleic acid-based external adjuvants within the TherVacB regimen in both wild-type and HBV-carrier mice.ResultsWhile HBcoreAg retained its structure upon bRNA removal, its stability and immunogenicity decreased significantly. Loading HBcoreAg with nucleic acid-based adjuvants re-established stability of the capsid-like antigen. Immunization with poly I:C- or CpG-loaded HBcoreAg induced high antibody titers against co-administered HBsAg. When applied within the TherVacB regimen, they activated vigorous HBcoreAg- and HBsAg-specific T-cell responses in wild-type and HBV-carrier mice, requiring a significantly lower dose of adjuvant compared to externally added adjuvant. Finally, immunization with adjuvant-loaded HBcoreAg mixed with HBsAg led to long-term control of persistent HBV replication in the HBV-carrier mice.ConclusionAdjuvant-loaded HBcoreAg retained capsid integrity and stability, was as immunogenic in vivo as externally adjuvanted HBcoreAg, requiring lower adjuvant levels, and supported immunity against co-administered, non-adjuvanted HBsAg. Thus, adjuvant-loaded HBcoreAg represents a promising novel platform for vaccine development.Impact and implicationsHepatitis B core antigen (HBcoreAg) recapitulates the capsid of the HBV that hosts the viral genome. Produced recombinantly, it is not infectious but emerges as a potent immunogen in vaccine development. In this preclinical study, we show that loading HBcoreAg with defined nucleic-acid-based adjuvants on the one hand stabilizes the HBcoreAg with standardized capsid content and, on the other hand, efficiently promotes the immunity of HBcoreAg and a co-administered antigen, allowing for reduced adjuvant doses. Therefore, adjuvant-loaded HBcoreAg not only serves as an encouraging option for therapeutic hepatitis B vaccines, but could also act as an efficient adjuvant delivery system for other types of vaccine.

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Design and Development of Modified mRNA Encoding Core Antigen of Hepatitis C Virus: a Possible Application in Vaccine Production

Publications

Design and Development of Modified mRNA Encoding Core Antigen of Hepatitis C Virus: a Possible Application in Vaccine Production

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