Andrea Cortese Hassett, Ph.D.
Scientific Director of the Coagulation Laboratory, Reference Laboratory Alliance,
Heparin is widely used for the prevention and treatment of
thromboembolic diseases. The anticoagulant response to heparin varies widely among
patients, possibly because of variations in the plasma concentrations of heparin-binding
proteins. There is evidence that the clinical efficacy of heparin is optimized if the
anticoagulant effect is maintained above a defined minimal level and that the risk of
bleeding is increased as the dose of heparin increases. For these reasons, the therapeutic
use of unfractionated (UF) heparin requires laboratory control, and activated partial
thromboplastin times (APTT) and thrombin clotting times (TCT) are most commonly employed.
There are clinical situations that make it difficult to monitor heparin by conventional
methods and in recent years, low molecular weight (LMW) heparin preparations have
increased in clinical use. LMW heparin preparations have lost the ability to prolong the
clotting time tests therefore, therapeutic monitoring by the APTT and TCT can no longer be
used. Monitoring of the new heparins and difficult clinical situations can be achieved in
an anti-Xa system.
Heparin, a naturally occurring polymeric mucopolysaccharide with
anticoagulant and antithrombotic properties, is effective in the prevention and treatment
of a variety of venous and arterial thromboembolic disorders. It is used as prophylaxis
for postoperative thrombosis, in conjunction with the initiation of oral anticoagulant
therapy, in the treatment of unstable angina and acute myocardial infarction and to
prevent clotting during cardiac bypass.
Commercial preparations of heparin are heterogeneous, their components
having molecular weights (mw) ranging from 3,000 to 30,000 (mean, 15,000). Only about one
third of the heparin binds to antithrombin III and this fraction is responsible for most
of its anticoagulant effect. The remaining two thirds has minimal anticoagulant activity
at therapeutic concentrations.
The heparin-antithrombin III complex inactivates a number of
coagulation enzymes, including thrombin and activated factors X, XII, XI and IX. Thrombin
and activated factor X (factor Xa) are the most sensitive to inactivation. The rate of
inactivation , under normal conditions, is slow but can be increased several thousand-fold
by heparin. This mechanism accounts for the anticoagulant effect of unfractionated
heparin. Low molecular weight heparin preparations (<5400 mw) are unable to bind
thrombin and antithrombin III (ATIII) simultaneously and therefore are unable to
accelerate the inactivation of thrombin by ATIII, but retain the ability to catalyze the
inhibition of factor Xa by ATIII.
Monitoring By APTT
Laboratory monitoring of heparin therapy is desirable to ensure that an
appropriate antithrombotic effect is obtained, while guarding against bleeding
complications of an overdosage. Currently, the APTT is the most common test used to
monitor heparin therapy. Monitoring by APTT evaluates heparin's overall activity
throughout the entire coagulation system i.e., inactivation of thrombin, Xa, XIIa, XIa and
IXa. Heparin treatment is usually monitored to maintain the ratio of the patient's APTT to
the mean control APTT within a defined range of approximately 1.5 to 2.5, referred to as
the therapeutic range. Laboratory and clinical studies have established a therapeutic
range that is equivalent to a heparin level of 0.2 to 0.4 U per milliliter (mL) by
protamine titration, or 0.35 to 0.7 U per mL according to the level of anti-Xa activity.
It should be noted that the responsiveness of the reagents used in APTT tests can vary
widely. The therapeutic range for any given APTT reagent should therefore be established
in the clinical laboratory to correspond to a heparin level of 0.2 to 0.4 U/mL by
One of the most common problems encountered is lack of an adequate
response to an "adequate" dose. There are two major causes for this; an
inadequate dose of heparin or an inadequate response of the APTT to adequate level of
Monitoring By Anti-Xa Assay
An alternative approach is to assay for heparin exploiting its
catalysis by antithrombin III inhibition of coagulation enzymes, particularly factor Xa.
The factor Xa inhibition test (anti-Xa assay) is the most useful test for assaying the
widest variety of therapeutic heparin preparations. In this method, both factor Xa and
antithrombin III are present in excess and the residual factor Xa activity is inversely
proportional to the heparin concentration. The assumption is made that the patient has a
normal concentration of antithrombin III. For a patient with ATIII deficiency a heparin
concentration is measured, but this does not necessarily correspond to the anticoagulant
capacity in vivo. It is recommended to also measure the antithrombin III level for all
patients under heparin therapy when using this type of assay to ensure normal ATIII
activity. The therapeutic range of the anti-Xa assay in the treatment of thromboembolic
disease established by laboratory and clinical studies for unfractionated heparin is 0.35
to 0.7 anti-Xa Units/mL. The therapeutic range for LMW heparins has not been well
established at this time.
The aim of therapy is to produce truly stabilized levels of circulating
heparin capable of maintaining an effective hypocoagulability. There are several clinical
situations in addition to the use of LMW heparins, where the specific measurement of
heparin levels using the anti-factor Xa method may be necessary. Patients receiving
heparin but demonstrating an inadequate APTT response can be evaluated for heparin by the
anti-Xa assay. Monitoring of heparin is difficult by conventional methods when the
baseline APTT is prolonged as seen in patients with lupus anticoagulants and deficiencies
of factor XII (Hagemen factor), prekallikrein (Fletcher factor) and high molecular weight
kininogen (Fitzgerald factor). A quantitative anti-Xa assay makes heparin monitoring
possible in these clinical situations.
In pregnant women requiring anticoagulation, heparin is the drug of
choice because it does not cross the placenta or produce untoward effects in the fetus or
newborn when administered to the mother. Studies have also shown that LMW heparin does not
cross the placenta. The anti-Xa heparin assay can provide accurate monitoring to ensure
therapeutic levels are maintained since the long term use of high doses of heparin is
associated with osteoporosis.
It is necessary to re-evaluate the clinical monitoring of the heparin
effect, since it has been shown that global clotting assays such as APTT and specific
assays such as anti-Xa measure different entities. For standard unfractionated heparins
and uncomplicated clinical situations, monitoring by APTT is sufficient. Clinical
situations such as a lupus anticoagulant, specific factor deficiencies and pregnancies
requiring long term anticoagulation must be monitored by the anti-Xa assay. LMW heparins
have high specific anti-Xa activity and low APTT activity. It will not be analytically
feasible to monitor heparin therapy with APTT methods. Currently, these drugs can only be
monitored by anti-Xa assays.
1. Hirsh, J. Drug Therapy, N. Engl. J. Med. 1991, 234: 1565-1574.
2. Hirsh, J., Levine, M.N. Blood. 1992, 79: 1-17.
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