Perumal D, Papagatsias T, Roberts M, Athanasopoulos T, Gray C, Dickson G, Gotch F, Patterson S; Conference on Retroviruses and Opportunistic Infections.

9th Conf Retrovir Oppor Infect Feb 24 28 2002 Wash State Conv Trade Cent Seattle Wash Conf Retrovir Oppor Infect 9th 2002 Seattle Wash. 2002 Feb 24-28; 9: abstract no. 281-T.

Imperial Coll. Sch. of Sci., Technology and Med., London, UK

BACKGROUND: The unique ability of dendritic cells (DC) to induce primary cellular immune responses makes them optimal candidates for vaccination protocols. In this study we are using in vitro derived dermal dendritic cells (DDC) and Langerhans-type cells (LC) as a tool to analyse T-cell response to new adenoviral vaccine constructs based on HIV-1 clade C env, pol, or gag genes obtained from isolates in South Africa. Here we report on antigen expression studies in DDC and LC infected with an adenoviral vector carrying the HIV-1 gag gene.METHODS: DDC and LC were derived from monocytes of healthy donors. The gag gene was cloned into an adenovirus vector through homologous recombination processes in bacteria. HIV-1 gag gene expression was initially examined by plasmid transfection studies using 911 cells. In vitro generated DDC and LC were transduced with the adenovirus vectors and assessed for gag and GFP expression by flow cytometry. The ability of the antigen-pulsed DC types to stimulate allogeneic T cells was determined by MLR.RESULTS: 911 cells transfected with AdTrack-CMV-gag construct exhibited a low HIV-1 gag level (22%) when compared to GFP expression (74%) despite both genes being present on the same plasmid. Co-transfection with pCMV-rev did not show any improvement in HIV-1 gag expression while GFP fluorescence remained high. In vitro generated immature DC infected with pAdEasy-DELTATP/DELTAPol-gag at an MOI of 100 exhibited GFP fluorescence 4 hours post-infection. After 48 hours, DDC exhibited 31% GFP and 15% gag while LC showed higher levels at 47% and 42%, respectively. These differences may be due to the phenotypic and functional properties of the 2 antigen-pulsed DC types. HIV-1 gag-pulsed DDC stimulated stronger T-cell proliferation than HIV-1 gag-pulsed LC or uninfected DDC.CONCLUSIONS: Here we show that in vitro derived DCs can be transduced by replication defective recombinant adenovirus encoding for HIV-1 clade C gag. These results are encouraging for the development of modified DC expressing other HIV-1 clade C genes for use in mapping CTL epitopes in uninfected individuals.

Publication Types:

• Meeting Abstracts

Keywords:

• Adenoviridae
• Dendritic Cells
• Genes, gag
• HIV-1
• In Vitro
• Langerhans Cells
• South Africa
• T-Lymphocytes
• T-Lymphocytes, Cytotoxic
• genetics

Other ID:

• GWAIDS0024385

UI: 102264009

From Meeting Abstracts