Surface display of an anti-DEC-205 single chain Fv fragment in XXXX increases internalization and plasmid transfer to dendritic cells in vitro and in vivo.

@entity1336 @entity23838 @entity19
BACKGROUND: @entity23838 ( @entity23838 ) are promising vehicles for delivery of a variety of medicinal compounds, including antigens and cytokines. It has also been established that @entity23838 are able to deliver cDNA to host cells. To increase the efficiency of @entity23838 -driven DNA delivery we have constructed @entity1336 strains targeting DEC-205, which is a receptor located at the surface of dendritic cells (DCs). The purpose was to increase uptake of bacterial cells, which could lead to improved cDNA delivery to immune cells. RESULTS: Anti-DEC-205 antibody (aDec) was displayed at the surface of @entity1336 using three different anchoring strategies: (1) covalent anchoring of aDec to the cell membrane (Lipobox domain, Lip); (2) covalent anchoring to the cell wall (LPXTG domain, CWA); (3) non-covalent anchoring to the cell wall (LysM domain, LysM). aDec was successfully expressed in all three strains, but surface location of the antibody could only be demonstrated for the two strains with cell wall anchors (CWA and LysM). Co-incubation of the engineered strains and DCs showed increased uptake when anchoring aDec using the CWA or LysM anchors. In a competition assay, free anti-DEC abolished the increased uptake, showing that the internalization is due to specific interactions between the DEC-205 receptor and aDec. To test plasmid transfer, a plasmid for expression of GFP under control of an eukaryotic promoter was transformed into the aDec expressing strains and GFP expression in DCs was indeed increased when using the strains producing cell-wall anchored aDec. Plasmid transfer to DCs in the gastro intestinal tract was also detected using a @entity19 model. Surprisingly, in @entity19 the highest expression of GFP was observed for the strain in which aDec was coupled to the cell membrane. CONCLUSION: The results show that surface expression of aDec leads to increased internalization of @entity1336 and plasmid transfer in DCs and that efficiency depends on the type of anchor used. Interestingly, in vitro data indicates that cell wall anchoring is more effective, whereas in vivo data seem to indicate that anchoring to the cell membrane is preferable. It is likely that the more embedded localization of aDec in the latter case is favorable when cells are exposed to the harsh conditions of the gastro-intestinal tract.
