Abstract
BackgroundAntigen presentation by non professional antigen presenting cells (APC) can lead to anergy. In genetic vaccines, targeting the macrophages and APC for efficient antigen presentation might lead to balanced immune response. One such approach is to incorporate APC specific promoter in the vector to be used.MethodsThree promoters known to be active in macrophage were selected and cloned in mammalian expressing vector (pAcGFP1-N1) to reconstruct (pAcGFP-MS), (pAcGFP-EMR) and (pAcGFP-B5I) with macrosialin, EmrI and Beta-5 Integrin promoters respectively. As a positive control (pAcGFP-CMV) was used with CMV promoter and promoterless vector (pAcGFP-NIX) which served as a negative control. GFP gene was used as readout under the control of each of the promoter. The expression of GFP was analyzed on macrophage and non-macrophage cell lines using Flow cytometry and qRT-PCR with TaqMan probe chemistries.ResultsAll the promoters in question were dominant to macrophage lineage cell lines as observed by fluorescence, Western blot and quantitative RT-PCR. The activity of macrosialin was significantly higher than other macrophage promoters. CMV promoter showed 1.83 times higher activity in macrophage cell lines. The expression of GFP driven by macrosialin promoter after 24 hours was 4.40 times higher in macrophage derived cell lines in comparison with non macrophage cell lines.ConclusionsBased on this study, macrosialin promoter can be utilized for targeting macrophage dominant expression. In vivo study needs to be carried out for its utility as a vaccine candidate.
Highlights
Antigen presentation by non professional antigen presenting cells (APC) can lead to anergy
Regardless of the observed difference between mRNA or protein level, our finding clearly shows that macrosialin dominantly govern the expression in macrophage derived cells
To determine whether APC expressing promoters could be useful in terms of its specificity and activity, we compared with the CMV immediate early promoter in macrophage and non-macrophage derived cells
Summary
Antigen presentation by non professional antigen presenting cells (APC) can lead to anergy. In genetic vaccines, targeting the macrophages and APC for efficient antigen presentation might lead to balanced immune response. One such approach is to incorporate APC specific promoter in the vector to be used. The DNA elicits both the arms of immune response following in vivo expression of the antigen [1]. In DNA vaccines, expression of antigen in non APC cells might lead to such an outcome. For the treatment of HIV-1, APC have been targeted through ex vivo priming by expressed antigen and reinoculation [16] Another approach is to target the expression to APC without expression in non APC cells, which could be achieved by using promoters active only in APC [17]. Lentiviral vectors were studied to deliver the gene into
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