The Endogenous Thrombin Potential as a Monitoring Parameter in a Patient with an Acquired Hemophilia A

The Endogenous Thrombin Potential as a Monitoring Parameter in a Patient with an Acquired Hemophilia A

U. Scholz, A. Siegemund, T. Siegemund, S. Petros and L. Engelmann

Introduction

Acquired hemophilia is a rare bleeding disorder of unknown etiology that mainly affects elderly patients. The bleeding diathesis mostly involves soft tissues and the severity is variable, from mild subcutaneous to life threatening internal bleeding episodes. The management of this bleeding disorder is not yet standardized.

Laboratory diagnosis and monitoring is based on the finding of prolonged activated partial thromboplastin time (APTT), marked reduction of the coagulation factor involved (mostly factor VIII) and presence of an inhibitor on Bethesda assay [1–3].

However, these parameters do not correlate with the severely of bleeding [4].

Based on our current understanding of the coagulation system, measurement of thrombin generation (TG) in platelet-rich plasma may present a possibility to bet- ter define the hemostatic status of a given individual. A recent study by our group [5] has shown that TG is decreased in patients with hereditary hemophilia A and B, and this is the result of a reduced factor Xa-generation on the surface of activated platelets due to the deficiency of the factor VIIIa-IXa (tenase) complex.

Based on these findings, the TG assay was used in a patient with an acquired hemophilia A in order to define the interaction of coagulation factors and platelets and improve the coagulation monitoring.

Patient

A 62-year-old woman was referred to our center due to a massive uncontrolled macrohematuria of a sudden onset. Other than a type 2 diabetes mellitus, no serious disease state was identified. She never had a bleeding diathesis prior to the present admission and family history was not remarkable.

On admission, factor VIII activity was 5.2% and factor VIII inhibitor was 7.6 Bethesda units. Bleeding was managed with activated prothrombin complex (FEIBA, Baxter) or recombinant activated factor VII (NovoSeven, NovoNordisk).

Initial treatment for inhibitor eradication included immunoadsorption, followed by administration of the anti-CD20 monoclonal antibody MabThera (Rituximab) and finally immunosuppression with cyclophosphamide and prednisolone. After 6 months of immunosuppressive treatment, a complete elimination of the inhibitor could be achieved.

I. Scharrer/W. Schramm (Ed.)

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Hemophilia Symposium Hamburg 2003

” Springer Medizin Verlag Heidelberg 2005

Method

Coagulation factor VIII activity was measured with the Behring Coagulation System (Dade Behring, Marburg) using a one-stage clotting assay and factor-defi- cient plasma. Factor VIII inhibitor was measured with the Bethesda assay.

258 U. Scholz et al.

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F-V III:C (% )

0,0 0,5 1,0 1,5 100,0 150,0 200,0

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FVIII-Inhibitor (Bethesda Units)

0 20 40 60 80 100 120 140

I R C+P

Fig. 1. Factor VIII activity and inhibitor concentration during the course of inhibitor treat- ment. (I = immunoadsorption; R = administration of Rituximab; C = cyclophosphamide;

P = prednisolone)

TG was measured in a platelet-rich plasma (PRP) as previously described [5]. In short, freshly drawn citrated blood was centrifuged for 10 minutes at 170 g. 25 µl of the test reagent (fluorogenic substrate, dimethyl sulfoxide, isotonic saline, CaCl 2 , tris puffer and exogenous and endogenous activator) was added to 100 µl of PRP.

Fluorescence intensity was measured with the Fluorescan Ascent 2.2 (LabSystems, Helsinki, Finland) at wavelengths of 340 nm (excitation) and 440 nm (emission). The measurement lasted 120 minutes and data are given in arbitrary fluorescence units The Endogenous Thrombin Potential as a Monitoring Parameter 259

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ETP (FU)

0 50 100 150 200 250 300

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Tim e to peak (s)

0 500 1000 1500 2000 2500 3000

Fig. 2. ETP and time to peak measurements during the course of inhibitor treatment

(FU). Not only the area under the TG curve (i.e., the endogenous thrombin potenti- al, ETP), but the time to peak is also presented to define the coagulation capacity [5].

Results

Figures 1 shows the plasma activity of factor VIII and the inhibitor during the course of treatment. Simultaneous TG measurements are shown to be useful for treatment monitoring (Fig. 2). TG was increased immediately after immunoadsorption and administration of rituximab. This corresponded with the clinical observation of ces- sation of bleeding, although factor VIII activity was still low and inhibitor titer high.

Further observation under immunosuppressive treatment demonstrated inhibitor eradication and normalization of factor VIII activity and TG.

Discussion

The thrombin generation assay in PRP allows monitoring the contribution of the plasmatic and the cellular (platelets) system in the coagulation process. In contrast to the measurement of individual coagulation factors or PTT or prothrombin time (PT), TG represents the total coagulation capacity.

Acquired hemophilia A is a rare disorder that may result in serious bleeding epi- sodes. The measurement of factor VIII activity or inhibitor level does not correlate well with the extent of bleeding, thus not sufficient enough for monitoring efficacy of acute care. In the present case, improvement in thrombin generation was shown after immunoadsorption and administration of rituximab, although no improvement in factor VIII activity or inhibitor level was observed. Particularly, inhibitor level mark- edly increased after application of rituximab (the mechanism is not known) without any deterioration in the hemostatic status of the patient. Therefore, the TG assay may be a useful tool to define the coagulation state of such patients and monitor treat- ment. Further investigation is necessary to verify these findings and establish the method in the clinical routine. The fact that PRP should be processed immediately and the long period of measurement required are problems yet to be solved.

References

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3. Grünewald M et al. 2001: Acquired haemophilia: experiences with a standardized approach.

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4. Scharrer I., Großmann R. 2000: Acquired inhibitory body hemophilia. Anaesthesist 49: 34–42 5. Siegemund T et al. 2003: Thrombin generation in severe haemophilia A and B: the endoge-

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