The effect of recombinant IgG antibodies against the leucine-33 form of the platelet β3 integrin (HPA-1a) on platelet function

Lotta Joutsi-Korhonen; Sandy Preston; Peter A. Smethurst; Martin IJsseldijk; Elisabeth Schaffner-Reckinger; Kathryn L. Armour; Nicholas A. Watkins; Michael R. Clark; Philip G. de Groot; Richard W. Farndale; Willem H. Ouwehand; Lorna M. Williamson

doi:10.1160/TH03-07-0484

Thrombosis and Haemostasis, Table of Contents

Thromb Haemost 2004; 91(04): 743-754
DOI: 10.1160/TH03-07-0484

Platelets and Blood Cells

Schattauer GmbH

The effect of recombinant IgG antibodies against the leucine-33 form of the platelet β3 integrin (HPA-1a) on platelet function

Authors

Lotta Joutsi-Korhonen

¹Departments of Haematology, Biochemistry and Pathology, University of Cambridge, UK

²National Blood Service, Cambridge, UK
Sandy Preston

¹Departments of Haematology, Biochemistry and Pathology, University of Cambridge, UK

²National Blood Service, Cambridge, UK
Peter A. Smethurst

¹Departments of Haematology, Biochemistry and Pathology, University of Cambridge, UK
Martin IJsseldijk

³Utrecht University Hospital, Utrecht, The Netherlands
Elisabeth Schaffner-Reckinger

⁴Laboratoire de Biologie et Physiologie Integree, University Center, Luxembourg
Kathryn L. Armour

¹Departments of Haematology, Biochemistry and Pathology, University of Cambridge, UK
Nicholas A. Watkins

¹Departments of Haematology, Biochemistry and Pathology, University of Cambridge, UK
Michael R. Clark

¹Departments of Haematology, Biochemistry and Pathology, University of Cambridge, UK
Philip G. de Groot

³Utrecht University Hospital, Utrecht, The Netherlands
Richard W. Farndale

¹Departments of Haematology, Biochemistry and Pathology, University of Cambridge, UK
Willem H. Ouwehand

¹Departments of Haematology, Biochemistry and Pathology, University of Cambridge, UK

²National Blood Service, Cambridge, UK
Lorna M. Williamson

¹Departments of Haematology, Biochemistry and Pathology, University of Cambridge, UK

²National Blood Service, Cambridge, UK

Abstract

Summary

Recombinant HPA-1a antibodies with Fc, mutated to remove destructive effector functions, have been developed as a potential therapy for fetomaternal alloimmune thrombocytopenia (FMAIT), via blockade of binding of human HPA-1a polyclonal antibodies to fetal HPA-1a1b platelets. We have assessed the effect of the IgG1 HPA-1a antibody B2G1 and two mutated derivatives in various functional assays in resting and agoniststimulated platelets of the three HPA-1 genotypes. With HPA-1a1b platelets (fetal genotype), the antibodies did not activate signalling or CD62P expression in resting platelets, did not change in vitro bleeding time (IVBT), and did not inhibit platelet adhesion to collagen in flowing blood. Adhesion of HPA-1a1b platelets to fibrinogen was reduced by 20%, and aggregation induced by ADP by 50%, but collagen-related peptide (CRP-XL)-induced aggregation was normal. With HPA-1a1a platelets, aggregation to both ADP and CRP-XL was inhibited, with total blockade of adhesion to fibrinogen and of IVBT responses. Interestingly, a monovalent antibody fragment with identical specificity had no inhibitory effect on aggregation. In static adhesion assays using human αIIbβ3 or αVβ3 transfectants of HPA-1a (Leu³³) phenotype, attachment to fibrinogen of the latter but not of the former was completely blocked by the HPA-1a antibodies. These observations are best explained by antibody-mediated blockade of the RGD binding site on β3 by a mechanism of steric hindrance. As the effect on platelet function is modest with HPA-1a1b (fetal type) platelets, the mutated HPA-1a antibodies described here could be developed further for FMAIT therapy.

Keywords

HPA-1a - platelet function - β3 integrin

Full Text

References

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