Suppression of FVIII Antibody Production By FVIII BAR T-Cell in Vitro in the Presence of Inhibitors

To modulate B-cell responsiveness to FVIII, we previously generated cytotoxic cells expressing FVIII C2 or A2 immunodominant domains as chimeric receptors. We termed these antigen-expressing engineered T cells, "BARs", for B-cell Antibody Receptor. These CD8 T cells directly interact and kill FVIII-specific B cells and anti-FVIII hybridomas in prophylactic experiments in vitro and in vivo. It was not known whether these BAR CD8s could function or would be blocked in the presence of circulating antibodies to the expressed BAR domains. To test this, we cultured FVIII C2 or A2 BAR CD8 T cell with a mixture of monoclonal antibodies specific for these domains (up to 10 BU), and then added them to spleen cells from FVIII-immunized mice. These spleen cells were then re-stimulated with FVIII and the antibody response was determined after 5 days. Our results showed that these BAR CD8 T cells were not blocked in their ability to suppress the antibody response to FVIII under these conditions. Coupled with the observation that BAR-T cells can be stimulated to proliferate by anti-FVIII monoclonals, these results suggest that BAR cytotoxic activity may still be effective in the presences of inhibitors. (Supported by NIH grant R01 HL126727)