title: Synergistic neutralizing antibody response to a dengue virus type 2 DNA vaccine by incorporation of lysosome-associated membrane protein sequences and use of plasmid expressing GM-CSF.
abstract: We have previously shown that a dengue virus type 1 DNA vaccine expressing premembrane (prM) and envelope (E) genes was immunogenic in mice and monkeys and that rhesus monkeys vaccinated with this construct were completely to partially protected from virus challenge. In order to improve the immunogenicity of dengue DNA vaccines, we have evaluated the effect of lysosome targeting of antigens and coimmunization with a plasmid expressing GM-CSF on antibody responses. A dengue virus type 2 candidate vaccine containing prM and E genes was constructed in which the transmembrane and cytoplasmic regions of E were replaced by those of the lysosome-associated membrane protein (LAMP). The modified vaccine construct expressed antigen that was colocalized with endogenous LAMP in lysosomal vesicles of transfected cells, whereas the antigen expressed from the unmodified construct was not. It was hypothesized that targeting of antigen to the lysosomal compartment will increase antigen presentation by MHC class II, leading to stronger CD4-mediated immune responses. Mice immunized with the modified construct responded with significantly higher levels of virus neutralizing antibodies compared to those immunized with the unmodified construct. Coimmunization of mice with a plasmid expressing murine GM-CSF enhanced the antibody response obtained with either the unmodified or the modified construct alone. The highest antibody responses were noted when the modified construct was coinjected with plasmid expressing the GM-CSF gene. These results could form the basis for an effective tetravalent dengue virus DNA vaccine.
authors.affiliation: Virology Program, Infectious Diseases Directorate, Naval Medical Research Center, Silver Spring, Maryland 20910, USA. raviprakashk@nmrc.navy.mil
authors.forename: K
authors.initials: K
authors.lastname: Raviprakash
authors.forename: E
authors.initials: E
authors.lastname: Marques
authors.forename: D
authors.initials: D
authors.lastname: Ewing
authors.forename: Y
authors.initials: Y
authors.lastname: Lu
authors.forename: I
authors.initials: I
authors.lastname: Phillips
authors.forename: K R
authors.initials: KR
authors.lastname: Porter
authors.forename: T J
authors.initials: TJ
authors.lastname: Kochel
authors.forename: T J
authors.initials: TJ
authors.lastname: August
authors.forename: C G
authors.initials: CG
authors.lastname: Hayes
authors.forename: G S
authors.initials: GS
authors.lastname: Murphy
mesh_terms: D016475:3T3 Cells; D000818:Animals; D000914:Antibodies, Viral / Q000276:immunology; D000915:Antibody Affinity; D015703:Antigens, CD / Q000235:genetics / Q000276:immunology*; D002522:Chlorocebus aethiops; D004279:DNA, Viral / Q000276:immunology*; D003716:Dengue Virus / Q000276:immunology*; D004357:Drug Synergism; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000235:genetics / Q000276:immunology*; D006801:Humans; D051907:Lysosomal Membrane Proteins; D008562:Membrane Glycoproteins / Q000235:genetics / Q000276:immunology*; D051379:Mice; D009500:Neutralization Tests; D010957:Plasmids; D019444:Vaccines, DNA / Q000235:genetics / Q000276:immunology*; D014709:Vero Cells; D014759:Viral Envelope Proteins / Q000235:genetics / Q000276:immunology*; D014765:Viral Vaccines / Q000235:genetics / Q000276:immunology*
chemical_list: D000914:Antibodies, Viral; D015703:Antigens, CD; D004279:DNA, Viral; C051821:E-glycoprotein, Dengue virus type 2; D051907:Lysosomal Membrane Proteins; D008562:Membrane Glycoproteins; D019444:Vaccines, DNA; D014759:Viral Envelope Proteins; D014765:Viral Vaccines; C074932:prM protein, Flavivirus; D016178:Granulocyte-Macrophage Colony-Stimulating Factor