Antibodies-recognising
peptides bound to the major histocompatibility complex (pMHC) represent potentially valuable and promising targets for
chimeric antigen receptor (CAR) T cells to treat patients with
cancer. Here, a human phage-Fab library has been selected using
HLA-A2 complexed with a heteroclitic
peptide variant from an
epitope shared among multiple
melanoma-associated
antigens (MAGEs).
DNA restriction analyses and phage ELISAs confirmed selection of unique antibody clones that specifically bind to
HLA-A2 complexes or HLA-A2-positive target cells loaded with native or heteroclitic
peptide.
Antibodies selected against heteroclitic
peptide, in contrast to native
peptide, demonstrated significantly lower to even negligible binding towards native
peptide or tumour cells that naturally expressed
peptides. The binding to native
peptide was not rescued by phage panning with
antigen-positive tumour cells. Importantly, when
antibodies directed against heteroclitic
peptides were engineered into CARs and expressed by T cells, binding to native
peptides and tumour cells was minimal to absent. In short, TCR-like
antibodies, when isolated from a human Fab phage library using heteroclitic
peptide, fail to recognise its native
peptide. We therefore argue that
peptide modifications to improve antibody selections should be performed with caution as resulting
antibodies, either used directly or as CARs, may lose activity towards endogenously presented tumour
epitopes.