Antigen-loading strategies for DC: comparison of lipofection of tumor-antigen-encoding RNA and RNA electroporation into DC; RNA electroporation with loading of DC with anti-DEC205-antigen constructssignalling dataset
We developed additional antigen-loading strategies for DC and obtained data from first experiments on which constructs lead to antigen presentation and which do not.
On the one hand we compared lipofection of tumor-antigen-encoding RNA to RNA electroporation into DC, and found that electroporation results in antigen expression in all cells, while lipofection results in higher antigen expression only in a portion of the cells (other cells are negative). Interestingly, the priming capacity of MelanA/HLA-A2-specific autologous CD8+ T cells by lipofected DC was higher compared to electroprated DC. On the other hand, we compared RNA electroporation to loading of DC 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). To quantify presentation of the epitopes by DC, we used bulk T cells electroporated with TCR-encoding RNA in stimulation assays. Cytokine production by these T cells was used as a read-out.
First experiments showed that loading of DC with the anti-DEC-205scFv-MAGE-A3-KKL construct led to antigen presentation, whereas heat-inactivated constructs or control constructs did not.
- molecule type
- ribonucleic acids,
- melanoma antigen MAGE-3,
- anti-DEC205 antigen,
- Major histocompatibility complex class II,
- Melan A
- experimental design type
- cellular modification design: RNA interfering experiment