STRUCTURAL BASIS OF THE REACTIVITY OF HUMAN MONOCLONAL ALLOANTIBODIES WITH HLA CLASS I ANTIGENS

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TITLE: STRUCTURAL BASIS OF THE REACTIVITY OF HUMAN MONOCLONAL ALLOANTIBODIES WITH HLA CLASS I ANTIGENS

Rene J. Duquesnoy,¹ Arend Mulder,² Geziena M.Th Schreuder,² Ilias I. N. Doxiadis,² Frans J. H. Claas.²

¹Division of Transplant Pathology, University of Pittsburgh Medical Center, Pittsburgh, PA; ²Department of Immunohematology, Leiden University Medical Center, Leiden, ZH, Netherlands

This report describes a new approach to analyze the reactivity and specificity of HLA alloantibodies. We have studied 64 human monoclonal antibodies (mAbs) tested by direct lymphocytotoxicity against a large panel (N > 700) of cells typed at the DNA allelic level during the 13th International Histocompatibility Workshop. These antibodies originated from pregnant women and for each one, we determined from the HLA types of the antibody producer and immunizer the array of non–self amino acid triplets (i.e. the so–called triplotype) of the immunizing antigen. This analysis was based on the premise that the complementarity–determining regions (CDRs) of an antibody make contact with multiple sites of an antigen and that antigen–antibody binding must release sufficient energy for a conformational change in the antibody molecule necessary to initiate the Complement cascade leading to cytotoxicity. Most mAbs reacted with HLA antigens expressing one triplet found on the immunizing triplotype, i.e. these antibodies had apparently one CDR that conveyed the antibody specificity for this triplet. The lymphocytotoxic activity of some mAbs seemed to require the interaction between a second CDR with another triplet on the antigen and sometimes, this involved a self–triplet. Other mAbs showed specificity for several triplets on the immunizing triplotype. As an example, antibody producer OK had yielded two similarly highly reactive mAbs to the immunizing HLA–A3 triplotype: 62Qe,144tKr,151aHe,163dT. Analysis of reactivity patterns with informative panel cells revealed specific recognition of three triplets: 62Qe, 144tKr and 151aHe, apparently by three different CDRs of these mAbs. Each triplet could be recognized independently by these mAbs but lymphocytotoxicity required always the interaction between the self–triplets 80gTl and 82lRg and apparent fourth CDR. These findings provide a new understanding of the structural basis of the interactions between HLA antigens and alloantibodies.