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1063 Structural Basis for Quinine-Dependent Antibody Binding to Platelet Integrin alphaIIbbeta3

Disorders of Platelet Number or Function
Program: Oral and Poster Abstracts
Session: 311. Disorders of Platelet Number or Function: Poster I
Saturday, December 5, 2015, 5:30 PM-7:30 PM
Hall A, Level 2 (Orange County Convention Center)

Jianghai Zhu, PhD1,2*, Jieqing Zhu, PhD3, Daniel W. Bougie, PhD3, Richard H. Aster, MD3 and Timothy A. Springer, PhD1,2*

1Department of Biological Chemistry and Molecular Pharmacology, and Department of Medicine, Harvard Medical School, Boston, MA
2Program in Cellular and Molecular Medicine, Children’s Hospital Boston, Boston, MA
3Blood Research Institute, BloodCenter of Wisconsin, Milwaukee, WI

Drug-induced immune thrombocytopenia (DITP) is caused by antibodies that react with specific platelet membrane glycoproteins when the provoking drug is present. More than 100 drugs including quinine have been implicated as causes of immune thrombocytopenia, a relatively common, sometimes life-threatening, disorder. The cause of DITP in most cases appears to be a drug-induced antibody that binds to a platelet membrane glycoprotein only when the drug is present. Quinine, originally used as a prophylactic against malaria, is used at lower concentrations to impart the bitter flavor to tonic water, and is still used occasionally for the prevention of nocturnal leg cramps. For unknown reasons, quinine is much more likely to cause DITP than other drugs, with the exception of heparin, which acts by a distinctly different mechanism. The hallmark of DITP caused by drugs other than heparin is an antibody that is non-reactive in the absence of the sensitizing drug but binds tightly to a platelet glycoprotein, usually integrin αIIbβ3 (GPIIb/IIIa) when the drug is present. In contrast to drugs that act as a hapten to induce hypersensitivity, drugs that cause DITP appear not to bind covalently to the target antigen and do not inhibit antibody binding at high concentration. Nor has it been possible to show that drug binds noncovalently to an autologous target and somehow primes it for antibody binding. A mechanism recently proposed to explain drug-dependent antibody binding in DITP proposes that DDAbs are derived from a pool of naturally-occurring immunoglobulins that react weakly with autologous targets and that drug reacts at the antibody-antigen interface to increase the Ka for binding (“sandwich model”). Studies to define the mechanism(s) responsible for drug-dependent antibody (DDAb) binding to platelets have been handicapped by the requirement to use human antibodies, which are polyclonal and often in short supply. Recently developed murine monoclonal antibodies 314.1 and 314.3, which are specific for the β-propeller domain of αIIb integrin and closely mimic the behavior of antibodies found in patients with quinine-induced thrombocytopenia in vitro and in vivo, have provided new tools for characterizing this interaction at a molecular level. In this report, we demonstrate specific, high-affinity binding of quinine to the complementarity-determining regions (CDR) of these antibodies and define in crystal structures the changes induced in the CDR by this interaction. Since in previous studies, no detectable binding of quinine to the target integrin could be demonstrated, the findings indicate that a hybrid paratope consisting of quinine and reconfigured antibody CDR plays a critical role in recognition of its target epitope by antibody and suggest that, in this type of drug-induced immunologic injury, the primary reaction involves binding of drug to antibody CDR, causing it to acquire specificity for a site on a platelet integrin. This previously undescribed mechanism for drug-dependent antibody-target interaction could have implications for other types of drug sensitivity.

Disclosures: Aster: BLOODCENTER OF WISCONSIN: Patents & Royalties: A patent application has been filed based partly on these findings (Method of detecting platelet activating antibodies that cause heparin-induced thrombocytopenia/thrombosis; PCT/US14/62591) .

*signifies non-member of ASH