Identification of antigens recognised by T cells from melanoma patients responding to immunotherapy
We are collecting immunological data from a melanoma patient, aiming to identify the antigen(s) recognized by T cells from this patient that experienced an outstanding response to immunotherapy. This patient received several adoptive T cell transfers and regressed from stage IV melanoma to a disease free state and has remained tumor free for several years. In a collaboration with Dr. O. Winqvist, Karolinska Institute, we attempted to improve the adoptive transfer protocol by stimulating T cells with monocyte derived DC pulsed with tumor lysate, instead of simply adding tumor lysate into PBMC cultures. T cells raised exhibited some tumor reactivity and outgrowth of a distinct T cell population could be observed. We intented to identify the target antigen recognized by the tumor reactive T cells in order to further improve T cell stimulations and to possibly complement treatment with DC vaccination. We then evaluated the capacity of autologous DC and HEK293 cells transfected with relevant HLA alleles to function as T cell targets in Elispot assays upon transfection with tumor antigen encoding plasmids. Due to strong background from the autologous DC we decided that HEK293 transfected with one HLA-allele at a time plus simultaneously transfected with up to 5 tumor antiges would be optimal to screen for antigen specificity in the patients T cells. However, during a second T cell culture to expand large numbers of T cells for this screening procedure a drastic expansion of regulatory T cells was observed. Since no sufficient number of T cells could be expanded the screening was not performed. The reason for the altered phenotype/function of the expanded T cells from the second bleeding of this patient is currently not known.
“Functional gap junctions facilitate melanoma antigen transfer and cross-presentation between human dendritic cells”
Elispot assays using autologous DC/HEK293 cells as T cell targets
Expanding patient derived T cells into tumor reactive CTLs
HEK293 cells transfected with HLA alleles
Monocyte derived DC pulsed with tumor lysate
T cells stimulated with monocyte derived DC pulsed with tumor lysate