title: Highly efficient production of a dengue pseudoinfectious virus.
abstract: Dengue is a major infectious disease that affects people living in tropical and subtropical regions around the world. The causative agents are dengue virus serotype 1, 2, 3, and 4 (DENV1, 2, 3, and 4). Developing a vaccine for dengue is a high priority for public health, but traditional methods have faced numerous obstacles due to the unique immunopathogenesis of dengue virus infection. Here, we report a novel dengue vaccine candidate based on dengue pseudoinfectious virus (PIV) produced by the incorporation of a dengue subgenomic replicon into viral particles in highly efficient packaging cells. The subgenomic replicon was constructed by deleting the capsid protein (C) gene from the dengue viral genome and optimizing the signal peptide sequence of pre-membrane protein (prM) to facilitate the formation of viral particles. Packaging cells were developed for inducible expression of a bi-protein Cpr, where the protein pr is the "pr" segment of viral protein prM that holds the protein C on the endoplasmic reticulum (ER). When the replicon was introduced into the packaging cells, protein C was released from the bi-protein Cpr by a replicon-encoded viral protease. Coordinate expression of viral structural proteins by the replicon and packaging cells led to the incorporation of the replicon into viral particle to produce PIVs. Animal tests showed that the dengue PIV vaccine was highly immunogenic and the immune response protected mice challenged with a hundred-fold LD50 inoculation of dengue virus. The method described here has the potential to be applied to vaccine development for other flaviviruses.
authors.affiliation: Departments of Oral Pathology, College of Dentistry, Howard University, Washington, DC, United States; Tegen Biomedical Co., Rockville, MD, United States. Electronic address: xpang@howard.edu.
authors.forename: Xiaowu
authors.initials: X
authors.lastname: Pang
authors.affiliation: Oriental Tengen Technology Development Co., Ltd. Biomedicals, Beijing, China.
authors.forename: Yinhan
authors.initials: Y
authors.lastname: Guo
authors.affiliation: Tegen Biomedical Co., Rockville, MD, United States.
authors.forename: Yanfei
authors.initials: Y
authors.lastname: Zhou
authors.affiliation: Oriental Tengen Technology Development Co., Ltd. Biomedicals, Beijing, China.
authors.forename: Wenchuan
authors.initials: W
authors.lastname: Fu
authors.affiliation: Departments of Oral Pathology, College of Dentistry, Howard University, Washington, DC, United States.
authors.forename: Xinbin
authors.initials: X
authors.lastname: Gu
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D036022:Capsid Proteins / Q000235:genetics; D002460:Cell Line; D006224:Cricetinae; D003715:Dengue / Q000517:prevention & control*; D053059:Dengue Vaccines / Q000096:biosynthesis* / Q000276:immunology; D003716:Dengue Virus / Q000235:genetics / Q000276:immunology; D017353:Gene Deletion; D051379:Mice; D008969:Molecular Sequence Data; D012093:Replicon; D014162:Transfection; D015678:Viral Structural Proteins / Q000096:biosynthesis*; D019065:Virus Assembly
chemical_list: D036022:Capsid Proteins; D053059:Dengue Vaccines; D015678:Viral Structural Proteins
keywords: Dengue vaccine; Immunogenicity; Packaging cell; Pseudoinfectious virus; Replicon