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.

Commercially available siRNA Smart Pool from Dharmacon targeted against the STAT3 transcription factor have been successfully transfected in human MDDCs.

In order to identify the APCs that were responsible for the development of this state of immune tolerance, we aimed to generate a transgenic mouse that express a recombinant fluorescent protein in the mammary glands, an approach that should result in high levels of expression of the fluorescent protein in the milk. We thus should be able to detect the cells that capture, degrade and present this antigen to T cells. Since we had previously generated a monoclonal antibody, 2C44, that reacts to a peptide derived from the Leishmania LACK protein bound to I-Ad MHC ...

We have shown recently that the T cell stimulatory capacity of DCs pulsed with tumorantigen-derived peptides can be considerably increased by activating the DCs through electroporation with CD40L, CD70 and constitutively active TLR4 encoding mRNA (TriMix DCs). This vaccine will also be tested in the multicentre trial.

The peptides are used in the generation of monocyte-derived DC (pulsing of the cells at a concentration of 5 µg/mL in approximately 5 mL and at a cell density of 20 x 10e6/mL for 1 h.).

We established tumor cell lines from melanoma in our laboratory.

We established tumor cell lines from pancreatic cancer in our laboratory.

We established tumor cell lines from renal cell cancer in our laboratory.

This tumor lysate is from patients with advanced (stage III or IV) melanoma. It is used for the generation of a vaccine and loading of DC.

These monocyte-derived dendritic cells are loaded with tumor lysate generated from the excised metastatic lesion of stage III/IV melanoma patients. They will be used for vaccine generation.

This tumor RNA was used to transfect matured DC with the final aim to produce a DC vaccine.

The immunotherapy potential of DC, pulsed with tumor antigens, was evaluated in EG7-OVA and P815 tumor models. Depletion of natural regulatory T cells in these tumor bearing mice resulted in rejection of P815 cells, but not EG7-OVA, suggesting that regulatory T cells have a stronger impact on immune responses against weakly immunogenic or auto-antigen (P1A).

Melanoma patients were vaccinated with 4 peptides (MAGE-A3, gp100, NA17 and tyrosinase) presented by HLA-A2 and monitored. We measured responses to these peptides injected with Montanide, but did not know whether the Montanide adjuvant was important for these T cell responses. We therefore injected the same peptides without adjuvant, and no response was observed. We conclude that Montanide had a clear adjuvant effect for the CD8 T cell responses measured against these four peptides.

These T cells are to be purified using MACS cell separation to contain only CD8+ T cells. The latter are then co-cultured with electroporated DC, stimulated and become evaluated on their cytokine secretion, cytotoxicity and % of antigen specific T cells.

Untouched naive CD4+ T cells are co-coltured with fungi -pulsed DCs to prime naive T cells.

Vectors were engineered to express distinct chemokines at the local skin site of mice. In particular, attention has been focussed on the recruitment of distinct subsets of DC to skin.

These constructs were prepared for a per-clinical model of breast cancer.

Yeast cells in different conditions of culture (along with R848, Curdland, yeast RNA and LPS) were used to induce DC activation.

In a process of total RNA extraction from S. cerevisia, the cells were collected and the pellet were resuspend in 3.6 ml AE Buffer.

Yeast RNA (along with R848, Curdland, LPS and yeast cells in different conditions of culture) was used to induce DC activation.