Alternatives to the ‘classical’ maturation method (i.e. a cocktail of inflammatory cytokines) for moDC

Our lab generated data on alternatives to the ‘classical’ maturation method (i.e. a cocktail of inflammatory cytokines) for moDC (e.g. ‘clinical grade reagents such as Ampligen TM and Hiltonol TM). Moreover the possibility to co-electroporate moDC with constitutively active TLRs and TAA encoding mRNA was persued. The in vitro charging of monocyte derived DC (moDC) with antigens has been optimised. In contrast to previous studies, the electroporation of TAA encoding mRNA after in vitro maturation of the moDC resulted in a better electroporation efficiency and antigen presentation.

We evaluated the capacity of mRNA-electroporated DC as a vaccine for immunotherapy. First, the early DC maturation kinetics and the effect of different lipopolysaccharide incubation periods on the phenotypic maturation profile of DC were determined. Next, we showed that either immature or mature DC are equally well electroporated and express and present the transgene at a comparable level after electroporation. We point out that the mRNA electroporation results in a negative effect on the interleukin (IL)-12p70, IL-6, and tumor necrosis factor-alpha secretion after maturation. Nevertheless, mRNA-electroporated DC induce an effective cytotoxic T lymphocyte (CTL) response in vivo. Mature electroporated DC are significantly more potent in eliciting an Ag-specific CD8+ CTL response compared to their immature electroporated counterparts. In addition, a significant improvement in CTL response is obtained both in the primary and in the memory effector phases when CD4+CD25+ regulatory T cells (Treg) are depleted in vivo prior to immunization. These findings are further substantiated in tumor protection experiments and hold convincing evidence for the merit of Treg cell depletion prior to immunization with mRNA-electroporated DC.

These results facilitate the preparation of DC-vaccines for clinical use.

A single step procedure for the in vitro maturation and antigen loading of human moDC was optimized. DC were electroporated with mRNA encoding tumor antigens and a putative TLR3 ligand. In vitro stimulation of naïve T cells indicated the superiority of this approach compared to in vitro stimulation with DC matured with inflammatory cytokines followed by electroporation with TAA encoding mRNA.

• molecule type
• lipopolysaccharide,
• cytotoxic T lymphocyte,
• Interleukin-12 p70,
• poly(I:C(12)U),
• tumor associated antigen,
• hiltonol,
• Interleukin-6,
• Tumor necrosis factor alpha,
• Toll like receptor,
• Toll like receptor 3,
• CD25 receptor,
• messenger RNA,
• CD4

• cell type
• regulatory t cell,
• mature dendritic cell,
• immature dendritic cell

• organism type
• Homo sapiens

created over 16 years ago (2 March 2009)    last modified over 12 years ago (6 August 2012)   [ RDF ]   [ RelFinder ]