We obtained data on antigen delivery by a vector with ovalbumin and CMVpp65 in mice or human, respectively. We used a MHC II presentation pathway targeting vector originally published by Wang et al. 1999. Proof of principle studies for antigen delivery have been performed with ovalbumin (murine studies) and CMVpp65 (human studies).

Our group obtained data on molecular signaling triggered by LPS in dendritic cells and the role of CD14 in the regulation of the DC life cycle through NFAT activation. Toll-like receptors (TLRs) are the best-characterized pattern recognition receptors (PRRs). Individual TLRs recruit diverse combinations of adapter proteins, and thereby trigger signal transduction pathways leading to the activation of various transcription factors, including nuclear factor (NF)-kB, activation protein (AP)-1, and interferon regulatory factors (IRFs). Interleukin-2 (IL-2) is one of th...

Our lab has data from an interim statistical analysis of the DERMA-ER-DC 04 trial (multipeptide loaded cytokine matured moDC +/- CD40L activation, 70 evaluated patients). This analysis for the first time showed a clinical correlation of induced immune responses as measured in the blood with outcome, as there have been so far statistically significantly less events in stage III patients showing good CTL reponses (IFN-y Elispot) to multiple class I and II peptides compared to low responders of the stage III cohort. Additional in vitro maturation of DC by CD40L (leuci...

We have data from a clinical trial with melanoma patients (stage IV, documented progress) receive DC transfected with RNA encoding MelanA, Mage3 and Survivin +/- E/L-selectin by the i.v. route. This DERMA-ER-DC 05 trial (multipeptide loaded cytokine matured moDC + prior Treg elimination by 3x ONTAK treatment [5µg/kg], 8 evaluated pat.) did not show any enhanced immune reponses compared to the equivalent cohort in the DERMA-ER-DC 04 trial. Patient recruitment into the 2nd phase of the DERMA-ER-DC 06 trial has started (sequential adaptive design). Immunomonitori...

Design of this trial: sequential adaptive design, 2nd cohort, mature DC transfected with RNA encoding MelanA, Mage3 and Survivin +/- E/L-selectin, i.v. route, 13 evaluated patients. Comparison of pre-existing and vaccine-induced immune responses in stage IV melanoma patients. 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. Monocyte-derived and...

BruCells is collaborating with Professor De Witte from ULB for the preclinical study in which circulating tumour-specific CD8+ T-cells will be detected in the blood of newly diagnosed glioblastoma patients and from which data will be obtained. In order to obtain the blood samples and tumour biopsies from the glioblastoma patients, a file has been submitted to the Ethics Committee of the Erasme Hospital and has been recently approved.

We have generated data on the developmental control of translation during DC activation. Our work (Lelouard et al., JCB) demonstrates that DCs down-regulate and change the quality of translation in order to increase their survival, however during this process MHC I antigen processing is affected. This has important consequences for the way antigens should be introduced in DCs and how long after stimulation they should be used during vaccination application. We have also initiated studies on the impact of dsRNA on the regulation of translation and the functional re...

Data were produced in our lab on a broad spectrum of priming events and their impact on the effector and memory phases of the CTL response. Activation of a cytotoxic T lymphocyte (CTL) response in an antigen-exposed lymph node involves a great diversity of encounters between naive CTLs and APCs that differ in both duration and quality. This broad spectrum of priming events instigates a complex blend of CTL developmental pathways. Using an experimental system that allows tight control over CTL priming, we have singled out defined priming events and analyzed the ...

Although many bacteria and protozoa can invade dendritic cells (DC), it is not clear whether infected DC process pathogen-derived antigen and stimulate antigen-specific CD4+ T cells in vivo. To address this issue and obtain data on this question, we have infected susceptible BALB/c (H2-d) mice with a recombinant Leishmania major parasite expressing a fluorescent tracer. We have directly visualized antigen-presenting cells using a mAb reacting to an antigenic peptide derived from the parasite LACK antigen bound to I-Ad MHC class II molecule. I-Ad/LACK complexes wer...

Data was obtained on the infiltration and destruction of solid tumors by cytotoxic T lymphocytes (CTLs). Although the immune system evolved to fight infections, it may also attack and destroy solid tumors. In most cases, tumor rejection is initiated by CD8+ CTLs, which infiltrate solid tumors, recognize tumor antigens and kill tumor cells. We used a combination of two-photon intravital microscopy and immunofluorescence on ordered sequential sections to analyze the infiltration and destruction of solid tumors by CTLs. We show that in the periphery of a thymoma gr...

Our data demonstrate that treatment of primed mice with intact anti-CTLA-4 antibodies induces the development of regulatory T cells expressing high levels of ICOS and producing IL-10. These regulatory T cells inhibit Th1 responses, in vitro and in vivo, and repress experimental intestinal inflammation, by a mechanism involving IL-10 and IDO. These ICOS+ regulatory T cells appear distinct from the naturally occurring regulatory T cells described above, suggesting that two populations can inhibit DC-induced Th1 responses in vivo. Experiments are under way to identi...

We have obtained data addressing the effectiveness of TLR ligand-long peptide conjugates in vivo by either direct vaccination or ex vivo DC loading prior to vaccination. Both approaches show similar effectiveness in CTL priming suggesting that TLRL-peptide conjugates efficiently target DC in vivo.

Knowledge and gene expression data were gained on differentiation patterns of T cells.

Data was obtained on the DC-activating, TLR-4-mediated stimulus, LPS, on rat intestinal and hepatic DC in vivo. DC present peripheral Ags to T cells in lymph nodes, but also influence their differentiation (tolerance/immunity, Th1/Th2). To investigate how peripheral conditions affect DC properties and might subsequently regulate T cell differentiation, we examined the effects of a potent DC-activating, TLR-4-mediated stimulus, LPS, on rat intestinal and hepatic DC in vivo. Our results suggest that any explanation of switching between tolerance and immunity as wel...

We investigated the effects of tumour hypoxia on DC differentiation and maturation and gathered data in mice. This is a an important topic as sites of inflammation are often characterized by both low oxygen tension and an extensive infiltration of inflammatory leukocytes, including DC, which need to move against oxygen gradients. We showed that oxygen availability is necessary to promote full monocyte-derived DC differentiation and maturation. Low oxygen tension (hypoxia) inhibits expression of several differentiation and maturation markers (CD1a, CD40, CD80, CD8...

We obtained data on the T-cell stimulatory capacity of human monocyte-derived DCs co-electroporated with different combinations of CD40L, CD70, and constitutively active toll-like receptor 4 (caTLR4) encoding mRNA. The effectiveness of the dendritic cell (DC) vaccination protocols that are currently in use could be improved by providing the DCs with a more potent maturation signal. We therefore investigated whether the T-cell stimulatory capacity of human monocyte-derived DCs could be increased by co-electroporation with different combinations of CD40L, CD70, and...

In 2007 we established routine monitoring of tumor-specific T-cell subsets by multicolor flow cytometry and obtained immunological data from this. For this purpose we used multimers kindly provided by Pierre Coulie (Brussels) together with surface markers (such as CD8, CD4, CD3, CD45RA, CCR7, CD25, CD27, CD137, CD127) in different 8-color panels on the BD FACSCanto II. Additionally we used functional markers such as CD107a and intracellular cytokine staining for the further analysis of individual T-cell populations and T-cell clones. Monitoring of the ongoing vacc...

Our lab has gathered data in establishing a per-clinical model of breast cancer.

We obtained data on the Pharmacological compound 1alpha,25-Dihydroxyvitamin D3 [1,25(OH)2D3], the biologically active metabolite of Vitamin D3, and its analogues, and its effect on the DC differentiation/activation pathway induced by type I IFN. 1,25(OH)2D3 prevented the generation of IFN-DCs when added to freshly isolated monocytes, and is capable to redirect already differentiated IFN-DCs toward a more immature stage. Interestingly the suppressive effect of 1,25(OH)2D3 was associated with a potent impairment of DC migration in response to inflammatory and lymph ...

Data is available from an evaluation of different stimulation conditions for the cytometric assessment of antigen-specific Th-cells based on CD154 expression induced during short-term in vitro activation, a method established by us during previous DC-THERA activities. This is important and essential for implementation of the technology into clinical immune monitoring in e.g. vaccination trials. Moreover, the affect of stimulation length was determined. Differences with respect to the induced frequencies of antigen-specific, activated CD154+ Th-cells were minor. I...