Substitution of Glutamic Acid at Position 71 of DRβ1*04:01 and Collagen-Specific Tolerance Without Alloreactivity

The DRB1 locus is strongly associated with both susceptibility and resistance to rheumatoid arthritis (RA). DRB1 alleles encoding the VKA or VRA epitope in positions 11, 71, and 74 confer the highest risk of developing RA, whereas the allele encoding VEA is protective. We therefore investigated the feasibility of creating antigen-specific tolerance without inducing alloreactivity by replacing lysine with glutamic acid at position 71 in DRβ1*04:01.

Individual DRB1 alleles and the DRB1*04:01^K71E allele were cloned into T2 cell lines to measure binding of biotinylated peptides. Transgenic animals expressing DRB1*04:01, DRB1*01:01, or DRB1*04:01^K71E were injected with collagen to measure T cell proliferation. Skin and bone marrow transplants between DRB1*04:01^K71E and DRB1*04:01 mice were performed to determine if the single amino acid change at position 71 would be recognized as foreign. DRB1*04:01 mice transplanted with DRB1*04:01^K71E bone marrow were injected with collagen to test if resistance to collagen sensitization could be transferred.

Replacing lysine (K) at position 71 in DRβ1*04:01 with glutamic acid (E) blocked collagen peptide binding and rendered the DRB1*04:01^K71E mice resistant to collagen sensitization. Skin and bone marrow transplants from DRB1*04:01^K71E mice were not rejected by DRB1*04:01 mice, suggesting the single E⁷¹ difference was not recognized as allogeneic. Bone marrow from DRB1*04:01^K71E mice adoptively transferred antigen-specific tolerance to collagen to DRB1*04:01 mice.

These studies demonstrate that editing a single amino acid in DRβ1*04:01 blocks collagen peptide binding without inducing alloreactivity and could therefore represent a gene therapy approach to induce antigen-specific passive tolerance.