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2002;
DOIuse
10.1182/blood-2002-04-1055
Anti-hirudin antibodies following low - dose subcutaneous
treatment with desirudin for thrombosis prophylaxis after hipreplacement surgery: incidence and clinical relevance.
A.Greinacher1, P.Eichler1, D.Albrecht1, U.Strobel1, B.Pötzsch2, B.I.Eriksson3
1. Institute for Immunology and Transfusion Medicine, Ernst-Moritz-ArndtUniversity Greifswald
2. Institute for Experimental Haematology and Transfusion Medicine, University
Bonn
3. Department of Orthopedic Surgery, Sahlgrenska University Hospital/ÖSTRA,
Göteborg University, Sweden
Word counts: abstract: 146
manuscript: 1193
Correspondence address:
Prof.Dr.med.Andreas Greinacher
Institut für Immunologie und Transfusionsmedizin
Ernst-Moritz-Arndt-Universität
Sauerbruchstr./Diagnostikzentrum
17487 Greifswald
Germany
Tel.: +49-(0)-3834-865482
Fax: +49-(0)-3834-865489
Email: [email protected]
running head: desirudin-antibodies incidence/relevance
scientific heading: thrombosis
Copyright (c) 2002 American Society of Hematology
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Abstract
Recombinant hirudin has been found to be immunogenic in patients treated with
lepirudin following heparin-induced thrombocytopenia (HIT). We assessed the
incidence of IgG-anti-hirudin antibodies by ELISA in 112 patients enrolled in a dose
finding study with desirudin. Patients received desirudin subcutaneously following
orthopedic hip surgery 10 mg b.i.d. (n=17), 15 mg b.i.d. (n=75), 20 mg b.i.d. (n=20).
11/112 patients (9.8%) developed anti-hirudin antibodies independently of the dose
(17.6%, 8.0%, 10.0%, n.s.). The rate of immunization did not differ from that
observed in HIT patients treated with lepirudin (p=0.113). Plasma concentrations of
desirudin did not differ between anti-hirudin antibody positive and negative patients.
Anti-hirudin antibodies had no impact on incidences of deep vein thrombosis and/or
pulmonary embolism, allergic reactions, hemorrhage. However, the total number of
immunized patients observed was low, and so infrequent, but severe, effects of antihirudin antibodies cannot be excluded.
Email: [email protected]
Introduction
Two recombinant hirudins are approved in the European Union, desirudin (Revasc,
Aventis) and lepirudin (Refludan, Schering). Desirudin is approved for thrombosis
prophylaxis following orthopedic hip replacement surgery and lepirudin for
anticoagulation in patients with heparin-induced thrombocytopenia (HIT) and
thromboembolic complications. Both compounds differ from each other in their Ntermini: desirudin possesses a valine – valine and lepirudin a leucine – threonine
structure. Otherwise the molecules are very similar with molecular weights of 6.96
kDa and 6.98 kDa, respectively.
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Rhesus monkeys did not show a specific immune response following intravenous
boluses of desirudin1. In contrast, balb/c mice showed a 100% anti-hirudin antibody
response after immunization by albumin- conjugated hirudin or hirudin fragments2,3.
Also, dogs generated anti-hirudin antibodies following immunization with desirudin4.
In healthy volunteers, a few allergic reactions were observed following intravenous as
well as subcutaneous applications of desirudin5. Allergic reactions occurred in three
volunteers, in another four the skin test became positive. In one volunteer anti-hirudin
antibodies were detected.
In none of the major clinical studies of patients with acute coronary syndromes was
an immune reaction towards one of the two hirudins reported6,7,8,9,10 . However, these
patients were usually treated for 2-3 days only, a too short period for a B-cell
response.
In 1996, we reported that a high proportion of patients receiving lepirudin for HIT
developed anti-hirudin antibodies11,12, a finding confirmed by others13,14. During a
mean duration of treatment of 14.4 days, 44% of 198 patients treated with lepirudin
developed anti-hirudin antibodies of the IgG class, which occurred as early as day 4
(peak at day 8-9). Of 87 patients with detectable anti-hirudin antibodies, 61 (70.1%)
developed during treatment, 23 (26.4%) occurred after cessation of lepirudin, three
became positive during a second treatment course with lepirudin only12.
In another study with lepirudin given to patients with HIT(up to 26 days), up to 74% of
patients developed anti-hirudin antibodies of the IgG-, IgA-, and/or IgM-class13,14.
None of these patients devoloped allergic reactions.
Desirudin is approved for thrombosis prophylaxis following orthopedic hip
replacement surgery based on three prospective trials15,16,17. These patients usually
require treatment for 7-10 days, which is within the time frame for induction of antihirudin antibodies. The purpose of the present study was to assess whether desirudin
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given subcutaneously in prophylactic dose in patients following orthopedic hip
replacement surgery is immunogenic, and to compare the outcomes of patients
testing positive and negative for anti-hirudin antibodies.
Materials and Methods
Patients:
Patients were enrolled in a prospective dose-finding study, as described elsewhere15.
Serum was obtained between days 6 and 11 after start of desirudin (mean day: 8 ±1).
Samples were frozen at -30°C or below.
Hirudins:
Desirudin was provided by Novartis Pharma, Nürnberg, Germany, lepirudin by
Aventis, Frankfurt, Germany.
Laboratory assays:
Anti-hirudin antibody ELISA was performed as described12. Microtiter wells (Covalink,
Nunc, Germany) coated with desirudin or lepirudin were incubated with patient
plasma (1:50, 60 min, 37°C), and IgG binding assessed by anti-human IgGperoxidase labelled antibodies. Sera of HIT patients with known anti-hirudin
antibodies served as positive controls, plasma of normal blood donors served as
negative controls. The assay was performed by personnel blinded for the dose of
desirudin given to the patients.
Concentrations of free desirudin in patient plasma samples were assessed by a
chromogenic assay using the thrombin substrate S2238, as described before18.
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Incidences of deep vein thrombosis and pulmonary embolism, allergic reactions, and
hemorrhages were compared by Fisher’s exact test (2-tailed). Occurrence of antihirudin antibodies over time was calculated by Kaplan-Meier statistics. Plasma
concentrations of anti-hirudin antibody positive patients and of anti-hirudin antibody
negative patients were compared by Wilcoxon rank-sum test.
Results
Samples from 112 patients were studied: 20 patients received a dose of 20 mg
desirudin s.c. b.i.d., 75 patients received a dose of 15 mg b.i.d. and 17 patients
received a dose of 10 mg b.i.d..
In 11/112 patients (9.8%), anti-hirudin antibodies were detectable. Results were
identical whether desirudin or lepirudin was used in the ELISA, indicating a 100%
cross-reactivity. Seroconversion rates were similar among the three dosing groups:
2/20 patients (10%) who received 20 mg desirudin b.i.d., 6/75 patients (8.0%) who
received 15 mg b.i.d., and 3/17 patients (17.6%) who receivd 10 mg b.i.d.. Compared
to the occurrence of anti-hirudin antibodies in HIT patients treated with lepirudin, no
differences were observed (p=0.113, log-rank test, figure 1).
Desirudin-plasma concentrations of anti-hirudin antibody positive patients did not
differ from those of anti-hirudin antibody negative patients within the three dosing
groups (10 mg, 0.1 µg/ml vs. 0.094 µg/ml [p=1.0]; 15 mg, 0.17 µg/ml vs. 0.28 µg/ml
[p=0.33] and 20 mg, 0.1 µg/ml vs. 0.17 µg/ml [p=0.63]).
Clinical endpoints could be evaluated in 109 patients. Outcomes of anti-hirudin
antibody positive patients did not differ compared to anti-hirudin antibody negative
patients: deep vein thrombosis and/or pulmonary embolism, 2/11 (18.2%) vs. 11/101
(10.9%), p=0.61; allergic reactions, 0/11 vs 1/101, p=1.0; hemorrhage, 1/11 (9.1%)
vs. 6/101 (5.9%), p=0.53.
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Discussion
The present study shows that subcutaneous desirudin is immunogenic in patients
after major orthopedic surgery. At day 8 of treatment, approximately 10% of patients
are immunized. Numerically this rate is much lower than the rate observed in patient
with heparin-induced thrombocytopenia receiving lepirudin12,13,14. However, using
Kaplan-Meier calculation, the risk of immunization per time exposed seems to be
identical in both patient populations (figure 1). Thus the total rate of immunization
depends on treatment duration and time point of blood sampling.
The two hirudins differ in the structure of the N-terminal part. However, antibodies of
patients treated with desirudin showed a 100% cross-reactivity to lepirudin and vice
versa, and therefore must recognize structures common to both hirudins with the
same immunogenicity.
Also different routes of application, intravenous vs. subcutaneous are of no relevance
for immunization, in the present study patients received hirudin only s.c., whereas
lepirudin was only given i.v..
As the dose was higher in lepirudin treated patients ( 250 mg per day in a 70 kg
patient) as compared to the present study, immunization is not dose related at least if
10 mg b.i.d. are given.
The result is surprising as the two patient groups are very different. We suspected
HIT patients to be more likely to form anti-hirudin antibodies, since they have already
formed antibodies towards platelet factor 4/heparin complexes. This indicates that
HIT patients do not have an increased tendency to form antibodies to other drugs.
Anti-hirudin antibody-desirudin complexes cannot be filtered by the glomeruli.
However, neither a difference in desirudin plasma levels, nor in bleeding
complications, could be observed between anti-hirudin antibody-positive and –
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negative patients. Also we found no evidence for inhibitory antibodies which have
been described in HIT patients, receiving lepirudin
12,19
. Due to the small numbers of
immunized patients, this observation must be considered preliminary.
In conclusion, desirudin given in prophylactic dose in patients after orthopedic hip
replacement surgery is immunogenic. The overall risk of immunization seems to be
the same as in HIT patients receiving lepirudin in therapeutic dose. As the treatmentand observation period was shorter in the present study, the percentage of
immunized patients was lower. The overall number of immunized patients, however,
is still small, and thus less frequent severe adverse effects might have been
overlooked up to now. Patients under treatment should be carefully observed for
adverse events.
Acknowledgements:
The present study has been supported by Deutsche Forschungsgemeinschaft DFG
Gr1096/2-3a and by a grant of Novartis Pharma.
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Legend
Figure 1
Occurrence of anti-hirudin-antibodies in relation to treatment duration
desirudin -treated patients
(n=112)
lepirudin -treated patients
(n=198)
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From www.bloodjournal.org by guest on June 18, 2017. For personal use only.
Prepublished online October 17, 2002;
doi:10.1182/blood-2002-04-1055
Anti-hirudin antibodies following low-dose subcutaneous treatment with
desirudin for thrombosis prophylaxis after hip-replacement surgery:
incidence and clinical relevance
Andreas Greinacher, Petra Eichler, Dorothea Albrecht, Ulrike Strobel, Bernd Poetzsch and Bengt I
Eriksson
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