We used dendritic cells loaded with anti-DEC205-antigen constructs, consisting of a single-chain variable fragment (scFv) directed against DEC-205, genetically linked to different parts of the MAGE-A3 model antigen (i.e. the KKl and EVDPIGHLY peptides), in a study to compare antigen-loading strategies.

We have finalized analysis of the receptors on DCs responsible for capture of exosomes secreted by mature DCs. We have shown that expression of LFA-1, but not of Mac-1, integrin, on DCs is required for capture of ICAM-1-bearing exosomes.

Lentiviral transduction of shRNAs: to this aim, shRNA specific for IRF4 and STAT3 are in the process to be cloned into the pLVTHM lentiviral vector (obtained from the repository Addgene). This vector has been already successfully used to generate self-inactivating lentiviral vectors previously tested in DCs for transfection efficiency and absence of effects on their phenotype.

We are currently performing gene silencing experiments in human monocyte-derived DCs (MDDCs) with the dual aim of generating relevant knowledge on (i) the transcriptional regulation of DC differentiation, and (ii) the safe and rationale in vitro manipulation of gene expression in DCs.

We observed that ICAM-1 expression by mature DCs is critical for long-lasting contacts with CD8+ T cells, but dispensable for short-lived antigen-specific interactions. Serial brief T cell-dendritic cell contacts induced early CD8+ T cell activation, proliferation and effector CTL differentiation in the first few days after immunization.

We decided to analyze the protein changes that occur at the cell surface of a maturing DC, induced by a prototypical maturation stimulus such as LPS. Results obtained from the LC-MS/MS analysis of the integral plasma membrane proteome of D1 cells stimulated for 24 hours with LPS have been compared to the results obtained from immature D1 cells.

For a transcriptional profiling of dendritic cells, DC were stimulated with PolyI:C, pretreated or not with PP2, and microarray analysis was performed.

The exosomes were used to immunize mice, hybridomas were generated that produce antibodies recognizing at least the human exosomes.

It has been shown that DCs express specific receptors for exosomes. Since ICAM-1 on exosomes is required for their ability to activate T cells via DCs in vitro, the role of Mac-1 and LFA-1 (the major ligands for ICAM-1) as potential receptors for exosomes has been investigated. A new role for LFA-1 on DCs, as a receptor for exosomes has been proposed.

These multipeptide loaded cytokine matured moDC +/- CD40L activation were used for the DERMA-ER-DC 04 trial.

The DC were matured with TLR ligand R848 and polyI:C. Using these DC, we obtained best results in our evaluation whether antigens could be rerouted from the endosomal into the cytosolic compartment by combining antigen with cell penetrating peptides to further enhance cross-presentation.

We have efficiently transfected DC with RNA encoding a functional protein (E/L-selectin), which allows entry of DC into LN from HEV. These DC rolled in vitro on sialyl-LewisX-coated slides, and in vivo, mouse E/L-selectin-transfected DC homed to LN after i.v. application.

We have investigated the in vivo capacity of resting (and of DC-primed) NK cells to reach the draining lymph nodes.

Since LPS-activated DC are potent inducers of NK cell priming and lymph nodes appear to be a key place in which DC and NK cell interactions occur, we have investigated the in vivo capacity of resting and DC-primed NK cells to reach the draining lymph nodes.

To quantify presentation of the epitopes by DC, we used bulk T cells electroporated with TCR-encoding RNA in stimulation assays.

These patients receive DC transfected with RNA encoding MelanA, Mage3 and Survivin +/- E/L-selectin by the i.v. route. 2nd phase of the DERMA-ER-DC 06 trial.

We have determined mechanisms for DC-induced Th1 development and the molecular pathways for inducing Th1 cells producing IFN-gamma solely, or Th1 cells producing IFN-gamma and IL-10, which have important implications for regulation of the immune response to eradicate pathogens with minimum pathology.

Using pharmacological inhibitors, or macrophages and DC from mice carrying a null mutation in MAP kinase signalling molecule(s), we have evidence for a role of certain MAP kinases in the regulation of IL-10 and IL-12, IFN-gamma production.

Using pharmacological inhibitors, or macrophages and DC from mice carrying a null mutation in MAP kinase signalling molecule(s), we have evidence for a role of certain MAP kinases in the regulation of IL-10 and IL-12, IFN-gamma production.

Using pharmacological inhibitors, or macrophages and DC from mice carrying a null mutation in MAP kinase signalling molecule(s), we have evidence for a role of certain MAP kinases in the regulation of IL-10 and IL-12, IFN-gamma production.