title: Inverted terminal repeat sequences of adeno-associated virus enhance the antibody and CD8(+) responses to a HIV-1 p55Gag/LAMP DNA vaccine chimera.
abstract: The immune responses to an HIV-1 p55Gag vaccine encoded as a DNA chimera with the lysosomal associated membrane protein-1 (LAMP) have been examined for the effect of the addition of the inverted terminal repeat (ITR) sequences of the adeno-associated virus (AAV) to the DNA plasmid construct, and of packaging the LAMP/gag gene as a recombinant AAV vector (rAAV). DNA plasmids encoding Gag and the LAMP/Gag protein chimera were constructed in two vectors, the pcDNA3.1 and a corresponding plasmid containing the ITR sequences (pITR) flanking the expression elements of the plasmid, and the pITR LAMP/gag DNA plasmid was encapsidated in the rAAV vector. Human 293 cells transfected in vitro with LAMP/gag plasmids either in pcDNA3.1 or pITR produced much Gag protein in cell extracts (1.6 and 2.2 ng of Gag/mg of protein, respectively). The immune responses of mice to immunization with these constructs were examined under three protocols: DNA prime/DNA boost, DNA prime/rAAV boost, and a single rAAV immunization. The results demonstrated that under DNA prime/DNA boost protocol, the "naked" DNA vaccines encoding the LAMP/gag chimera, either as pcDNA3.1 or pITR DNA plasmid constructs, elicited strong CD4(+) T cell responses. In contrast, significantly higher levels of CD8(+) and antibody responses were observed with the pITR-DNA constructs. Immunization with the rAAV vector under the DNA prime/rAAV boost protocol resulted in sustained T cell responses and a markedly increased antibody response, predominantly of the IgG(1) isotype resulting from the activation of the Th2 subset of CD4(+) T cells, that was sustained for at least 5 months after immunization.
authors.affiliation: The Johns Hopkins School of Medicine, Baltimore, MD 21205, USA.
authors.forename: Priya
authors.initials: P
authors.lastname: Chikhlikar
authors.forename: Luciana
authors.initials: L
authors.lastname: Barros de Arruda
authors.forename: Shikha
authors.initials: S
authors.lastname: Agrawal
authors.forename: Barry
authors.initials: B
authors.lastname: Byrne
authors.forename: William
authors.initials: W
authors.lastname: Guggino
authors.forename: J Thomas
authors.initials: JT
authors.lastname: August
authors.forename: Ernesto T A
authors.initials: ET
authors.lastname: Marques
mesh_terms: D000818:Animals; D000914:Antibodies, Viral / Q000097:blood*; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology*; D002460:Cell Line; D000229:Dependovirus / Q000235:genetics*; D005260:Female; D015683:Gene Products, gag / Q000235:genetics / Q000276:immunology*; D005822:Genetic Vectors; D006801:Humans; D007115:Immunization Schedule; D051907:Lysosomal Membrane Proteins; D008562:Membrane Glycoproteins / Q000235:genetics / Q000276:immunology*; D051379:Mice; D008807:Mice, Inbred BALB C; D010957:Plasmids; D011498:Protein Precursors / Q000235:genetics / Q000276:immunology*; D011993:Recombinant Fusion Proteins / Q000276:immunology; D020079:Terminal Repeat Sequences / Q000276:immunology*; D019444:Vaccines, DNA / Q000008:administration & dosage / Q000276:immunology*; D052118:Lysosomal-Associated Membrane Protein 1
chemical_list: D000914:Antibodies, Viral; D015683:Gene Products, gag; D051907:Lysosomal Membrane Proteins; D008562:Membrane Glycoproteins; D011498:Protein Precursors; D011993:Recombinant Fusion Proteins; D019444:Vaccines, DNA; C489116:LAMP1 protein, human; D052118:Lysosomal-Associated Membrane Protein 1; C052255:Lamp1 protein, mouse; C059767:p55 gag precursor protein, Human immunodeficiency virus 1