An MHC restricted antibody based chimeric antigen receptor requires TCR like affinity to maintain antigen specificity
Chimeric antigen receptors (CARs) are synthetic receptors that usually redirect T cells to surface antigens independent of human leukocyte antigen (HLA) Here, we investigated a T cell receptor like CAR based on an antibody that recognizes HLA A*0201 presenting a peptide epitope derived from the cancer testis antigen NY ESO 1. We hypothesized that this CAR would efficiently redirect transduced T cells in an HLA restricted, antigen specific manner. However, we found that despite the specificity of the soluble Fab, the same antibody in the form of a CAR caused moderate lysis of HLA A2 expressing targets independent of antigen owing to T cell avidity. We hypothesized that lowering the affinity of the CAR for HLA A2 would improve its specificity. We undertook a rational approach of mutating residues that, in the crystal structure, were predicted to stabilize binding to HLA A2. We found that one mutation (DN) lowered the affinity of the Fab to T cell receptor range and restored the epitope specificity of the CAR. DN CAR T cells lysed native tumor targets in vitro, and, in a xenogeneic mouse model implanted with two human melanoma lines (A2+/NYESO+ and A2+/NYESO DN CAR T cells specifically migrated to, and delayed progression of, only the HLA A2+/NY ESO 1+ melanoma. Thus, although maintaining MHC restricted antigen specificity required T cell receptor like affinity that decreased potency, there is exciting potential for CARs to expand their repertoire to include a broad range of intracellular antigens.
Molecular therapy oncolytics