Background
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The Lipo5 lipopeptides included in the ANRS vaccine trials have been initially designed on the basis of the immunodominant CD8+ T cell epitopes but have been shown to induce both CD4+ and CD8+ T cell response in healthy vaccinees.  In order to characterize the CD4+ T cell response specific for the Lipo5 sequences, we derived Lipo5 specific T cell lines using in vitro stimulation and characterized their specificity. 

Methods
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Specific CD4+ T cell responses were induced in vitro against Lipo5 peptides on 8 HLA unrelated naive donors.  Purified CD4+ T cells were stimulated once a week during 4 weeks, by mature DC loaded with Lipo5 peptides.  The CD4+ T cell response was revealed by IFN-γ ELISpots. 

Results
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All 8 donors developed an important specific CD4+ T cell response.  Globally, we found a mean of 32 specific CD4+ precursors per twenty million of peripheral CD4+ T cells.  Fifty seven per cent of the Lipo5-specific response was against Gag 253--284, 150 T cell lines being specific for this peptide.  T cell response specific for Pol 325--355, Gag 17--35, Nef 116--154 and Nef 66--97 represented 21%, 11%, 6% and 5% of the Lipo5 specific response, respectively.  In agreement with its binding capacity, we demonstrated that the strong immunogenicity of Gag 253--284 was due to its presentation by multiple HLA DR molecules.  All the 51 specific CD4+ T cell lines, with the exception of two, were stimulated by the clade B and C consensus sequences. 

Conclusion
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In agreement with the observations made in vaccinees, Lipo5 sequences introduced in the ANRS vaccine elicit a strong CD4+ T cell response in vitro and contain multiple CD4 epitopes.  We therefore confirmed the good immunogenicity of the Lipo5 peptides and demonstrated their capacity to elicit a multiepitopic CD4+ response cross reacting with clade B and C sequences.