Note: Descriptions are shown in the official language in which they were submitted.
21S69~1
EE~ W~K~ AKTIE~Gl~SEI,I,SC~laFT 1994/~013-Ma lC37
Dr. Bo/hg
Compo3itions which are 3uitable for employment as
antidotes to blood anticoagulants, and their use
The invention relate3 to antidote3 to blood
anticoagulant3 .
Blood coagulation is a complex process in which pla3ma
proteins and cells are involved. Formation of a wound
10 closure is initiated by rapid adhesion of blood platelets
to the damaged 6urfaces and by proteases and contact
factors which are activated successively in a cascade-
like reaction sequence. This course of events leads to
the production of thrombin which, inter alia, catalyzes
15 activation of fibrinogen to form fibrin.
Proteins such as factors VIII and V contribute, as
accelerators, to the ~ff;r~ ney of this system. In
addition to the protein C system, inhibitors of the
proteases involved essentially regulate the coagulation
20 proce3ses in such a way that, while wound clo3ure takes
place, excessive reaction3 are avoided. Di3tur~ance of
thi3 finely tuned equilibrium can lead to fibrin
depo3ition or the formation of thrombose3. A3 i3 known,
the vessel occlusions which result from thi3 are the
25 cause of many pathophy3iological proce33es 3uch as
cardiac infarction, in particular. This problem assumes
particular importance Ln tho3e patient3 who are 3ubject
to increased risk of thrombosis, which often represents
a po3t-operative complication.
30 Variou3 approache3 are taken for preventing the3e
pathophysiological proce3se3. Thu3, anticoagulant3 which
intervene in dif f erent way3 in the event of coagulation
are often used in clinical practice to prevent the
formation of a thrombu3 or el3e to 3upport its di3so-
35 lution (thrombolysi3). In addition to reducing or pre-
venting thromoocyte functions (aggregation, adhe3ion and
activation and the 3ynthesi3 and 3ecretion of activation
21~991
-- 2
products ), inhibitors o participating coagulation
protease3, in particular of the thromboplastin/factor
VI~a complex, of factor Xa and, in particular, of
thrombin (factor IIa) are being investigated and applied.
5 E~irudin, which was originally isolated from the salivary
gland3 of the leech E~irudo m~r~ n~ , is a particularly
specific and potent inhibitor of thrombin. In the
interim, a variety of dif f erent variant3, analogs,
fragments and mutants of hirudin have become known and
10 are being investigated for their suitability. While
hirudin is sulfated in its naturally occurring form, it
has been observed that ~he advantageous biological
activity is also retained in the non-sulfated form. As a
conse~uence, hirudin or its derivatives can conveniently
15 be prepared by means of genetic manipulation.
The blood anticoagulants also include heparin, and its
fragments and variants, as well a~ synthetic inhibitors
which inhibit the catalytic activity of thrombin and
other coagulation proteases, for example factor Xa. These
20 components can also be employed in the prophylaxis and/or
therapy of thrombotic or pre-thrombotic complications.
The search for ever more specific and potent
anticoagulant~ and antithrombotic agents ref lects the
unsatisfactory situation that the ~1;nil-~lly established
25 substallces such as heparins or vitamin K antagonist ii are
either insufficiently effective or else exhibit side
e~ects which are undesirable and in some cases serious.
Eirudill is currently the most powerful known thrombin
inhibitor which does not al~o interfere with the other
3 0 enzymes of the blood coagulation cascade . To date,
hirudin has not been observed to have any appreciable
side effects on pul3e rate, respiration, blood pres~ure,
thrombocyte count or the content of ~ibrinogen or
hemoglobin. For this reason, hirudin i9 a preferred blood
35 anticoagulant.
The u~e of anticoagulants or antithrombotic agents in a
215~991 '
.
-- 3 --
patient naturally entails the risk of inGreasing the
tendency to bleed which, in extreme situation~, for
example in a3sociation with improper use or dosing, can
lead to uncontrolled hemorrhages. In order to be able to
counteract the undesirable tendency to bleed in such a
situation, it is pos3ible rapidly to remove ~30me
substances, whose constitution (molecular weight and
structure) permits it, from the blood plasma of the
patient by dialysis. However when this is not possible
lp due to the lack of appropriate apparatus or Eor other
reasons, an antidote must be made available.
Some substances or preparations have already bee~ pro-
posed as antidotes. Thus, the thrombins and their blocked
variants exhibit a certain hirudin-neutralizing e~fect;
however, on the one hand, the use of active thrombin in
patients who are being treated with blood anticoagulants
is contraindicated as a rule and, on the other, it is
problematic to prepare inactive thrombin using low
molecular weight inhibitors which enter into covalent
2 0 bond3, owing to the toxicity o~ these substances . In
addition, there is a lack of experience with regard to
whether these complexes possibly have an antigenic
effect. Owing to their nature, the3e complexes are not
suitable for neutralizing low molecular weight
(synthetic1 antithrombins or antithrombotic agents. While
meizothrombin, an int~r~ 3te which arises briefly when
prothrombin i3 activated to form thrombin, po33es3e3 a
certain antagoni3tic potency, it can tran3form rapidly
into thrombin in vivo and, as 3uch, i3 contraindicated as
3 0 a rule .
Investigation o~ factor VII as a hirudin antidote in
appropriate models failed to provide satisfactory
results. European Patent Application EPA 89.810733.9
describes the u3e o~ Eactor VIII, or fragment3 of factor
VIII, a3 antidote3 to blood anticoagulant3. It i3 al30
known that 3ubstances, 3uch as, for example, desmopre33in
(Mannucci, P.M., ~rogress in E~aemo3tasi3 and Thrombosi3,
~, 21~6ggl
8 (lg86), pp. 19-45), can be u~ed which increase the
concentration of factor VIII in the blood.
~owever, 3ince even these antidote3 are not adequate for
all area3 of application, there i3 3till the need to make
5 available antidote3 to blood anticoagulant3, anti-
thrombotic agent3 and platelet antagoni3t3 which are more
effective and have fewer side effect3.
Within the 3cope of the pre3ent application, anti-
coagulatory or antithrombotic sub3tance3 are under3tood
to mean glyco3aminoglycan3 and 3ulfated poly3accharide3,
including heparin3, in particular unractionated and low
molecular weight (L~W) pento3an poly3ulfate, heparan
3ulfate, dermatan 3ulfate or chondroitin 3ulfate, and
al30 hirudin3 and their ragment3, for example hirugen
(J. Biol. Chem. 265 (1990), 13484-13487; EP-A-0 333 356),
analog3 or mutant3, for example l~lirulog~ (W0 92/13952),
coupled hirudin3, f or example PEG-hirudin or hirutonin3,
natural or 3ynthetic inhibitor3 of the thrombo-
pla3tin/factor VIIa complex, of factor Xa or o thrombin,
2 ~ components of the protein C sy3tem, namely activated
protein C, protein S and thlu..~, l;n, inhibitor3 of
fibrin aggregation or cro331inking, 3ub3tance3 from the
vitamin K antagoni3t group, 3ub3tance3 which have an
effect on platelet activation and/or platelet aggregation
25 or el3e platelet adhesion, 3uch a3, in particular,
f ibrinogen receptor antagoni3t3, or which are able to
increa3e the endogenou3 capacity for fibrinoly3i3, 3uch
a3, in particular, PAI-l inhibitor3, or atimulator3 of
the 3ynthe3i3 and 3ecretion Of pl ~pm; nogen activator
30 inhibitors, and thus have an indirect antithrom.botic
ef ect .
It i~ particularly important to provide a 3uitable
antidote to the 3aid coupled hirudin3, for example PEG-
hirudin, 3ince the3e co,upled hirudin3 can only be removed
35 rom the organi3m with difficulty.
-
_ 5 _ 21~;6991
The present invention therefore relate3 to compositions
which are used as antidotes to blood anticoagulants, it
being possible for these blood antieoagulants to be
selected from the group consisting of glycosaminoglycans
5 and sulfated polysaccharides, such as hirudins, and their
fragments, analogs or mutants, natural or synthetie
inhibitors of the thromboplastin/factor VIIa complex, of
faetor Xa or of thrombin, eomponents of the protein C
systeIn, namely aetivated protein C, protein S and
10 thL~ ~vl,Ludulin, inhibitors of fibrin aggregation or
erosslinking, substances from the vitamin 1~ antagonist
group, substances which have an effeet on platelet
aetivation and/or platelet aggregation or else platelet
adhesion, sueh as, in particular, fibrinogen reeeptor
15 antagonists, or which are able to inerease the endogenous
eapaeity for fibrinolysis, such as, in particular, PAI-l
inhibitors, or s~; m~ lrs of the synthesis and seeretion
of plasminogen activator inhibitors, whieh eompositions
exhibit a prothrQmbin complex concentrate and at least
20 one additional eomponent which promote~ blood
coagulation .
The prothrombin eomplex concentrates which are used in
aeeordanee with the invention are isolated from the
eorresponding blood prepzLrations by means of enriehment.
25 The prothrombin eomplex eoneentrates eontain, in par-
tieular, faetors II, VII, IX and X. The prothrombin
eomplex eoneentrates (ppss) ean be present in non--
activated f orm or in aetivated f orm with activation
eustomarily being effeeted by reaeting the prothrombin
3 0 eomplex eoncentrate with thrombin or eontact activators,
f or example . The activated prothrombin eomplex eoneen-
trates therefore reaet eonsiderably faster and are
eorre~pondingly more diffieult to eontrol. ~owever, the
non-aetivated prothrombin complex concentrates are
3~ preferred within the scope of the present invention.
In a preferred embodiment, the additional component which
promotes blood coagulation is factor VIII, with the
21~6991
additional component which promotes blood coagulation
particularly preferably being a combination of factor
VIII and the von Willebrand factor (vWF~.
In other, likewise preferred, embodiments, the additional
5 component which promotes blood coagulation is factor V,
factor Va, von W~ hr~n~ factor, factor XI, factor XII
or factor XIII, in particular factor3 V and VIII. Where
appropriate, mixtures of these components can al30 be
used .
10 These components can also be present in the form of
active fragments or variants which can, in particular, be
prepared by genetic manipulation . In another pref erred
embodiment of the present invention, the additional
component which promotes blood coagulation i8 at least
15 one lipid or a mixture of lipids, with negatively charged
lipids or lipid mixtures being preferred.
In another preferred embodiment of the present invention,
the additional component which promotes blood coagulation
is at least one diuretic agent.
20 Compoæitions are particularly preferred which comprise a
pro~hrt ; n complex concentrate and at least two
additional components - as described above - which
promote blood coagulation.
In the compositions according to the invention, the
25 prothrombin complex concentrates are employed in a
concentration of ~rom 1 to 2 0 0 0 international units ( IU )
per kilogram of the bodyweight of the patient to be
treated, with concentrations of the prothrombin complex
concentrate ln a quantity of irom 10 to 1000 IU/kg being
3 C particularly pref erred .
The additional component which promote~ blood coagulation
is customarily present in a concentration of in each case
from 1 to 2000 IU/kg, with concentrations of from 10 to
2156~91
-- 7
1000 IU/kg being particularly preferred in this case as
well .
The diuretic agent and/or the lipid or lipid mixture is
customarily present in a concentration of from 0.01 to
5 100 mg/kg of patient bodyweight, with concentrations of
the diuretic agent and/or the lipid or the lipid mixture
ln a quantity of from 0.1 to 10 mg/kg being particularly
pref erred .
In a further embodiment, the present invention ralates to
10 compositions which are used as antidotes to blood anti-
coagulants, it being possible for the blood anticoagu-
lants to be 3elected from the group consisting of glycos-
aminoglycans and sulfated polys~ h~ri ~e3, 3uch as
hirudins, and their fragments, analogs or mutants,
15 natural and synthetic inhibitors of the thrombo-
plastin/factor VIIa complex, of factor Xa or of thrombin,
components of the protein C 3ystem, namely activated
protein C, protein S and th~ lulin~ inhibitors of
fibriLI aggregation or crosslinking, substances from the
20 vitamin ~ antagonist group, substances which have an
effect on platelet activation and/or platelet aggregation
or else platelet adhesion, such as, in particular,
fibrinogen receptor antagonists, or which are able to
increase the endogenous capacity for fibrinolysis, such
25 as, in particular, PAI-1 inhibitors or stimulators of the
3ynthesis and secretion of pl~3Tn;no~en activator
inhihitors, which compositions comprise, on the one hand,
at least one ~- _ nn~nt selected from the group lipid,
lipid mixture and diuretic agent, and, on the other hand,
30 at least one component selected from the group pro-
thrombin complex concentrate, one of factors II, V, Va,
VIII, IX, X, XI, XII or XIII, and von Willebrand factor,
as a single factor, or as mixtures of these factors.
In a further ~mho~ -nt, the present invention relates to
35 a composition which is used as an antidote to blood
anticoagulants, it being possible f or the anticoagulants
~ 21~6991
-- 8 --
to be selected from the group consisting of glycosamino-
glycans and sulfated polysaccharides, such as hirudins,
and their fragment3, analogs or mutants, natural and
synthetic inhibitors of the thromboplastin/factor VIIa
5 complex, of factor Xa or of thrombin, components of the
protein C system, namely activated protein C, protein S
and th~ 1; n, inhibitors of f ibrin aggregation or
crosslinking, substances from the vitamin K antagoni3t
group, substances which have an ef f ect on platelet
10 activation and~or platelet aggregation or else platelet
adhesion, such a3, in particular, fibrinogen receptor
antagonists, or which are able to increase the endogenous
capacity for fibrinolysis, such as, in particular, PAI-1
inhibitors or stimulators of the synthesis and secretion
15 of plasminogen activator inhibitors, which compositions
each comprise at least one c~ n~T3t selected from one of
the t~o groups A and B, where group A comprises f actors
II, V, Va, VIII and IX, and group B comprises factors II,
V, Va, VII, VIIa and VIII, von Willebrand factor, and
20 factors IX, X, XI, XII and XIII, with the proviso that
the component or the components from group A is/are not
identical to the component or components f rom group B .
The invention also relates to the use of one of the
abovedescribed compositions as an antldote to blood
25 anticoagulants.
rhe present invention is further illustrated by the
following examples.
~xample 1
In this in vitro experiment, the potential of F VIII
30 (Beriate'!g), a non-activated prothrombin complex concen-
trate ~PeSB/heparin-free Beriplex'!g), and a combination of
the two, to antagonize the coagulation-inhibiting effect
of hirudin was tested in standard human plasma lS~P).
For this purpose, 100 ~Ll volumes of citrate-SllP were
35 brought, after the addition of in each case 2 ~1 of an
approp~iately concentrated stock solution, to f Lnal
21S6991
- - 9
concentration~ of 3 and 20 ,ug/ml hirudin, respectively.
After mixing together with 50 1~l of a concentrated
solution containing F VIII and/or PPSB, and adding 100 1ll
of Pathromtin<9/kaolin suspension, the whole was incubated
5 at 37C for 2 min. The reaction was started by i~1m;~in~
50 ~1 of a 50 mM solution of CaCl2. The (mo~;f;ed) acti-
vated partial thromboplastin time (aPTT) was det~-rm;n-
~in accordance with Schnitger and Gross.
The coagulation times listed in Table 1 make clear that
10 the extension in the aPTT (as compared with the control)
which was brought about by 3 ~g/ml hirudin was shortened
by 2 IU of F VIII. The extension produced by PPSB on its
own may possibly be attributed to an in vitrQ effect
which has not previously been completely elucidated. The
15 combination of the 2 compounds was f ound to be particu-
larly effective. This effect became all the more apparent
when very high hirudin concentrations (20 ~g/ml) were
employed which prevented non-antagonized plasma f rom
clotting even after 2000 seconds. l~ere! 1 IU of PPSB on
20 its own exhibited a certain shortening effect while, on
the other hand, 2 IU of F VIII on its own was not able to
elicit any clotting. By contrast, the combination
F VIII/PPSB was once again found to have a markedly
~etter ef f ect than the individual components .
2 5 Table
aPTT ( sec . )
~iru~lin Control 2 IU F VIII 1 IU PPss 2 IU F VIII
conc. /ml /ml + 1 IU
~g/ml ~ PPSB/ml
036.4 31.5 31.4 24.7
30 3 9~.1 76.0 121.1 57.7
20>2000 >2000 313.6 92.7
Table 2 show~3 different combinations of F VIII/PPSB
concentrations which supplement the above experimental
2ls699l
-- 1 0
series and confirm the results (using 10 ~g/ml hirudin
~nd comp~rlng w~tt th~ control~.
2ls6
/-
-- 1 1 --
Table 2
PPSB
(IU ml) 0 0.25 0.5 1.0
F VIII
5( IU~ml )
037.734.1 31.9 31.4
170 . 1 181 . 5 196 . 6 225 . 5
1.033.8 28.0 27.1 26.7
140.0101.1 83.9 86.2
2.031.7 26.1 24.2 24.2
123.289.6 67.8 69.7
4.028.g 22.9 21.3 22.1
107.286.1 54.6 56.6
10 normal: (control~, without hirudin
bold: 10 ~g/ml hirudin
Fxa~le 2
a) In a corresponding manner to that employed in
Example 1, rat citrate plasma to which 10 l~g/ml
hirudin were added was used in the aPTT test to
. investigate the antagonizing effect of F VIII, PPSB
and combinations of the two. The coagulation times
shown in Table 3 demonstrate once again that the
combination produces a more effective n~ l; 7~tion
2 0 than do the individual components .
21~6991
-- 12 --
Table 3 Rat citrate plasma/aPTT
Pl?SB
(IU/ml) 0 0.25 0.5 l.0
F VIII
5,( I~J/ml )
025.9 31.0 33.8 35.1
111.0 133.6 138.6 141.5
1.0 23.0 23.2 25.4 27.2
76.1 49.8 52.6 54.1
2.0 21.3 21.6 23.8 25.1
62 . 1 42 . 6 45 . 8 47 . l
4.0 19.9 20.0 20.8 23.0
56.0 38.7 37.6 23.1
normal: without hirudin (control~
bold: 10 ~g/ml hirudin
b) Experiments with rat whole blood, which was investi-
gated in a thromboelastograph (TEG) in order to
approach more closely to the in vivo situation,
confirm the results obtained in the aPTT test
(Table 4 ) . The r value (reaction time) es#entially
denote3 the time until clotting begins i . e . the time
from initiation of the reaction (blood removal or
r~lr;f;~ation) to the widening of the TEG curve by
l mm. The k value (clot forming time) corresponds to
the time which elapses from the endpoint of the r
value until the curve has widened to 20 mm.
For this purpose, 200 ,ul of rat citrate whole blood
were mixed with 5 0 ~Ll of an appropriately concen-
trated stock solution of hirudin (in diethyl bar-
~iturate buf f er, pE~ 7 . 6; 1 96 human serum albumin )
and with 50 ~l of antidote (F VIII, PPSB or F VIII
+ PPSB) and subjected to preincubation at 37C for
3 min. The reaction was started by adding 50 ~l of
a 100 mM solution of CaCl2.
21 56991
-- 13 --
Table 4: Rat whole blood/TEG
Elirudin Control 2 IU 1 IU 1 I~ PPSB
(llg/ml) F VIII/ml PPSB/ml 2 IIJ
F VIII/ml
10 r>40.0 min. >40.0 min. 3.7 min. 2.0 min.
k>4 0 . 0 min >4 0 . 0 min . 2 . 4 min . 1. 5 min .
5 Example 3
The antagoni~ing effect of the combination of factor V
and PPS33 was tested in a manner corresponding to that
employed in Example 1. Once again, it was found that use
of the combination was superior to use of the individual
10 components. The results are presented in Table 5.
Tahle 5
aPTT ( sec. )
~irudin Control 2 IU F V 1 I~ PPSB Z IU F V +
conc. /ml /ml 1 IU
(~g/ml) PPSB/ml
15 0 36.4 3~.3 31.4 28.9
20 >2000 >2000 313.6 156.3
Example 4
In principle, an undesirable, excessive anticoagulatory
reaction which is o~tained using a low molecular weight
20 thromoin inhibitor, for example argatroban (MD 805,
(2R,4R) -4-methyl-1-{N-{3-methyl-1,2,3,4-tetrahydro-8-
quinolinyl) sulfonyl}-L-argininyl}-2-pip~r; ~; nerA~hoxylic
acid), can also be countered using the described com-
ponents; however the combination o~ the two is most
25 effective. Typical eifects on the aPTT of S~P
( implementation as described in Example 1 ) are presented
in Table 6.
21S6991
-- 1 4 --
Table 6
PPSB aPTT tsec. )
IU/ml )
F VIII 0 0.25 0.5
5( IU/ml )
036.5 32.6 31.2
217.7 216.5 228.3
l.Q 33.4 26.9 26.1
18g . 8 164 . 4 161 . 7
2.0 31.1 24.2 23.8
177 . 2 137 . 4 135 . 2
4.0 28.1 21.6 20.0
164 . 1 122 . 4 114 . 8
10 normal: without 2aD 805, control
bold: lO ~g/ml M~ 805
A similar ef f ect can also be achieved when bivalent
thrombin inhibitors of the ~Iiruloge~!3 type, such as D-Phe-
Pro-Arg-Pro-Gly-Gly-Gly-Gly-Asn-Gly-Asp-Phe-Glu-Glu-Ile-
15 Pro-Glu-Glu-Tyr-Leu (analog), are uged, as tl~.ri.~ y
way of Example in Table 7.
Table 7
aPT~ ( 3ec. )
Analog Control 2 IU 1 IU PPSB 2 IU F VIII
( ~g/ml ) F VIII/ml /ml 1 IU
PPSB/ml
203.0 g7.0 77.0 107.1 66.6
Example 5
In a corresponding manner to that employed in Example 1,
Sh~ to which 10 ~g/ml hirudin were added was investigated
in the aPTT test for the antagonizing effect of F VIII,
25 F IX and the combination of the two. The coagulation
2~56991
-- 15 --
times shown in rra~le 8 demonstrate once again that, while
F IX also has a certain antagonizing effect, the
combination of the individual romronl~ntS is superior.
Table 8
F IX aPTT ( sec . )
- ( IU~
F VIII 0 1 2 4
( IU/IIL1 ~
0137.5 121.7 113.0 107.3
10 2 103.3 90.6 86.2 78.3
~irudin concentration: 10 ~g/ml
Example 6
In a corresponding manner to that employed in Example 1,
S~IP to which 20 ~g/ml hirudin were added was investigated
15 in the aPTT test for the antagonizing effect of F II,
F V, F VII, F VIII, F IX and F X and combi3rations of
these compounds. The coagulation times shown in Table 9
demonstrate oncc again that the combinations of the
compounds are superior to the individual components. The
20 combination of all 6 components was found to be the most
ef f ecti~e .
Table 9
aPTT ( sec )
2 IU factor V i 2000.0
25 2 IU factor V ~ -
2 IU factor II 182 . 0
2 IU f actor V +
2 IU f actor II +
2 IU factor VII 152 . 0
-
2~6991
-- 16 --
Continuation of Table 9
aPTT ( ~ee )
2 IU f aetor V +
2 IU f aetor II +
52 IU faetor VII +
2 IU faetor VIII 53 . 0
2 IU faetor V +
2 IU faetor II +
2 IU faetor VIr +
2 IU faetor VIII +
2 IU f actor X 3 9 .1
2 IU f aetor V f
2 IU f aetor II +
2 IU faetor VII +
2 IU faetor VIII +
2 IU f aetor IX +
2 IU factor X 29 . 6
2 IU f aetor V +
2 0 2 IU f aetor II +
2 IU faetor VIII 105 . 0
2 IU f aetor V +
2 IU faetor II +
2 IU faetor VIII +
2 IU faetor IX 51. 0
2 IU f actor V +
2 IU faetor II +
2 IU faetor VIII +
2 IU factor IX +
2 IU faetor X 32 . 0
2 IU f aetor V +
2 IU f aetor II +
2 IU faetor IX +
3!~ 2 IU faetor X 91.3
21~6991
-- 17 --
Continuation of Table 9
aPTT ( 3ec )
2 IU factor V +
2 IU factor VII > 2000 . 0
5 2 IU f actor V +
2 IU factor VIII i 2000 . O
2 IU factor V +
2 IU factor VIII +
2 IU factor IX > 2000.0
2 IU factor V +
2 IU f actor VIII +
2 IU factor IX +
2 IU factor X > 2000.0
2 IU factor VII ~ > 2000 . 0
2 IU f actor VII +
2 IU factor II 202.8
2 IU factor VII +
2 IU factor II +
2 IU factor VIII 141.0
2 IU factor VII +
2 IU f actor II +
2 IU factor IX 173 . 5
2 IU f actor VII +
2 IU factor II +
2 IU factor X 173 . 0
2 I~ factor VII +
2 IU factor VIII > 2000 . 0
2 IU factor VII +
2 IU factor IX > 2000.0
21~69gl
-- 18 --
C--nt i n~ tion of Table 9
aPTT ( see )
2 IU factor VII +
2 IU faetor X > 200~ . 0
5 2 IU faetor VIII +
2 IU faetor II +
2 IU faetor IX +
2 IU faetor X 84 . 0
10 2 IU fae~or IX +
2 IU faetor II 210.3
2 IU faetor IX +
2 IU f actor II +
2 IU faetor X 148.0
2 IU f actor X > 2 0 0 0 . 0
2 IU factor X +
~ lU ~actor II 196,0
