The majority of HLA-DR3 alloreactive T cells is peptide specific, but does not recognize known DR3-bound sequences

Rejection of transplants is frequently caused by activation of alloreactive T cells that recognize HLA/peptide differences between patient and graft. This T-cell response can be directed towards the HLA molecule, the HLA-bound peptide or towards a combination. More insight in the involvement of peptides in this process may help to find ways to avoid rejection using for example antagonist peptides. In recent years many naturally processed HLA-bound peptides have been identified. This raises the question of whether these, presumably abundant, peptides are involved in class II-specific allorecognition. To investigate this, we first determined the proportion of peptide-specific alloreactive T cells in the alloresponse against HLA-DR3. For this purpose we have tested a panel of DR3-specific alloreactive T-cell clones against a DM-mutant (i.e. peptide loading deficient) cell line. We found that 59 out of 64 alloreactive T-cell clones were dependent upon the presence of DM for an optimal response. However, only 2 DM-dependent T-cell clones recognize known peptide sequences. Thus we conclude that most DR3-specific alloreactive T-cell clones are peptide specific and that the currently known DR3-bound peptides are not the main target for allorecognition. Finally, we identified 4 T-cell clones that recognized the DM-mutant better than the wild-type cell line. The response against the wild-type cell line could not be restored with invariant chain derived peptides (CLIP). This provides additional evidence that DM can negatively select self-peptides other than CLIP, which can result in selection against peptides involved in allorecognition.