The Hunt for HLA-DQ Allogeneic Eplets Is Not Over: Four New Ones, Including a Cross-Chain Eplet

The target of an anti-HLA antibody is an epitope on the surface of the antigen, and in particular the polymorphic eplet of its core, which contains one or a few polymorphic residues accessible to the molecule surface. The HLA Eplet Registry database references more than 550 HLA eplets thus deduced from AA sequence alignments. However, not all eplets have yet been verified and this list is not exhaustive. We performed a systematic analysis of sequence alignment of DQ antigens, to identify polymorphic amino acids so far not proposed as candidate eplets. From this, we describe and validate 3 DQB1 eplets targeted by sera of organ-transplanted patients and explore a new eplet overlapping the DQA1*03 and DQB1*03 chains. Serum antibody profiles using LABScreen, Lifecodes and a complementary DQ Luminex single antigen bead panel all showed a concordant pattern incriminating these residues. Moreover, antibodies were adsorbed and eluted using human splenic mononuclear cells and HLA-DQ transfected murine cell clones, thereby validating these four eplets. Their localisation by 3D modelling revealed uncertain accessibility of some residues implicated. The protrusion and accessibility of an eplet on the HLA surface may ultimately not always be sufficient to predict antibody binding, since the dynamic flexibility of the molecule can modify these parameters. Our strategy, combining Luminex single antigen assays, cell adsorption/elution and in silico prediction of surface exposure, altogether concur to ascertaining the realness of candidate eplets in the highly complex HLA system.