Inverted terminal repeat sequences of adeno-associated virus enhance the antibody and CD8+ responses to a HIV-1 p55Gag/LAMP DNA vaccine chimera

Priya Chikhlikar, Luciana Barros De Arruda, Shikha Agrawal, Barry Byrne, William Guggino, J. Thomas August, Ernesto T.A. Marques

Research output: Contribution to journalArticlepeer-review


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 IgG1 isotype resulting from the activation of the Th2 subset of CD4+ T cells, that was sustained for at least 5 months after immunization.

Original languageEnglish (US)
Pages (from-to)220-232
Number of pages13
Issue number2
StatePublished - Jun 1 2004


  • AAV
  • APC
  • CD4
  • CD8
  • DNA vaccine
  • HIV-1 Gag
  • Inverted terminal repeat
  • LAMP
  • MHC
  • MHC II
  • MHC class II-enriched compartment
  • MIIC
  • RRE
  • Rev-responsive elements
  • antigen presenting cell
  • lysosome-associated membrane protein
  • major histocompatibility complex
  • rAAV

ASJC Scopus subject areas

  • Virology

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