PB0142 - FVIII-containing immune complexes interrupt platelet aggregation in an ex-vivo model of primary hemostasis

Sunday, June 25, 2023

18:30 – 19:30 ET

Room: Exhibition

Presenting Author(s)

Olga Oleshko, MD (she/her/hers)

PhD student
Department of Hematology, Hemostasis, Oncology and Stem Cell Transplantation, Hannover Medical School, Hannover, Germany
Hannover, Niedersachsen, Germany

Co-Author(s)

AK

Annika Klingberg

Technical assistants
Department of Hematology, Hemostasis, Oncology and Stem Cell Transplantation, Hannover Medical School, Hannover, Germany
Hannover, Niedersachsen, Germany
UC

Ute Curth

Professor
Institute for Biophysical Chemistry, Hannover Medical School, Hannover, Germany
Hannover, Niedersachsen, Germany
SW

Sonja Werwitzke

Specialist in laboratory medicine
Department of Hematology, Hemostasis, Oncology and Stem Cell Transplantation, Hannover Medical School, Hannover, Germany
Hannover, Niedersachsen, Germany
AS

Amrita Sarkar

Research Associate
Division of Hematology, Children's Hospital of Philadelphia, Philadelphia, PA
Philadelphia, Pennsylvania, United States
LR

Lubica Rauova, MD, PhD

Associate Professor
Division of Hematology, Children's Hospital of Philadelphia, Philadelphia, PA
Philadelphia, Pennsylvania, United States
MP

Mortimer Poncz

Professor
Division of Hematology, Children's Hospital of Philadelphia, Philadelphia, PA
Philadelphia, Pennsylvania, United States
Andreas Tiede, MD PhD (he/him/his)

Professor of Hemostasis and Thrombosis
Department of Hematology, Hemostasis, Oncology and Stem Cell Transplantation, Hannover Medical School, Hannover, Germany
Hannover, Niedersachsen, Germany

Background: Acquired hemophilia A (AHA) caused by neutralizing autoantibodies against coagulation factor VIII (FVIII) can lead to life-threatening bleeds. With extensive subcutaneous hematomas, bleeding after venipuncture, and frequent muscle bleeds, the clinical picture of AHA differs from congenital hemophilia A. The underlying reason is unknown. Recently, we showed that FVIII and anti-FVIII antibodies form immune complexes (IC) incorporating von Willebrand factor (VWF). FVIII-IC were also demonstrated to disturb platelet function in a collagen-coated flow chamber model. Here, we describe the impact of FVIII-IC on primary hemostasis under more physiological conditions.

Aims: To examine the impact of VWF-containing FVIII-IC on primary hemostasis in whole blood from healthy volunteers using an endothelial cell (EC)-coated flow chamber model.

Methods: FVIII-IC, formed with monoclonal anti-FVIII IgG antibodies directed against the A2 or C2 domain of FVIII were characterized by analytical ultracentrifugation (AUC). FVIII-IC were then generated with these antibodies in citrated whole blood. Human umbilical vein EC were seeded into Bioflux 1000 flow channels and treated with localized photochemical injury. Subsequent to flow experiments, channels were stained for VWF and anti-FVIII antibodies and imaged under the confocal microscope.

Results: The formation of VWF-containing FVIII-IC by anti-A2 or anti-C2 FVIII antibodies was shown by AUC (Figure 1). In the presence of antibodies, platelet adhesion under shear was reduced by 28 to 71% as compared with controls. VWF and anti-FVIII antibodies were found to be co-localized (Figure 2) demonstrating these were deposited at the injury site.

Conclusion(s): VWF-containing FVIII-IC disturb primary hemostasis in an EC-based flow chamber model. This suggests that primary hemostasis impairment contributes to the unique bleeding pattern in AHA and may be an important therapeutic target.

The study was supported by the “Early Career Research Grant 2021” from the Society of Thrombosis and Haemostasis Research (GTH e.V.).