Circulation. 1998;98:813-820
(Circulation. 1998;98:813-820.)
© 1998 American Heart Association, Inc.
Protective Effect of Oral Xemilofiban in Arterial Thrombosis in Dogs
Increased Activity in Combination With Aspirin
Leo G. Frederick, MS;
Osman D. Suleymanov, MS;
James A. Szalony, MS;
Beatrice B. Taite, AA;
Anita K. Salyers, BS;
Lucy W. King, BS;
Larry P. Feigen, PhD;
; Nancy S. Nicholson, MBA, MS
From the Thrombosis Research Department, Searle, Skokie, Ill.
Correspondence to Leo G. Frederick, Senior Research Investigator, Searle, 4901 Searle Pkwy, Skokie, IL 60077.
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Abstract
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Background—Inhibition of
platelet aggregation by preventing the binding of fibrinogen to
glycoprotein (GP) IIb/IIIa on activated
platelets results in antithrombotic activity. We report on the
antithrombotic effect of xemilofiban (SC-54684A), an oral GP IIb/IIIa
antagonist, administered alone or with aspirin (ASA) in an
acute thrombosis model.
Methods and Results—Conscious dogs were treated with xemilofiban
(1.25, 2.5, 5.0, or 6 mg/kg, n=6); low-dose (LD, 81 mg) ASA, n=7;
high-dose (HD, 162 mg) ASA, n=6; xemilofibran 1.25 mg/kg plus LD ASA,
n=6; xemilofibran 1.25 mg/kg plus HD ASA, n=6; or placebo, n=7. Dogs
were anesthetized 60 minutes later, and the effects
of the treatments were evaluated after electrolytic injury (250 µA
for 180 minutes) in the left circumflex coronary artery.
Bleeding time (BT) was assessed in a separate study. Incidence of
thrombosis was reduced (P<0.05) by xemilofiban 
2.5
mg/kg, HD ASA, or xemilofiban 1.25 mg/kg plus HD ASA compared with
placebo. Xemilofiban 2.5 mg/kg or xemilofiban 1.25 mg/kg plus HD ASA
significantly increased time to occlusion, inhibited ex vivo
platelet aggregation to collagen >90%, and prevented or decreased
(P<0.05) cyclic flow variations (CFVs) compared with
placebo. BT was increased (P<0.05) with xemilofiban
2.5 mg/kg but not with xemilofiban 1.25 mg/kg plus HD ASA.
Conclusions—Xemilofiban 2.5 mg/kg, HD ASA, or xemilofiban 1.25
mg/kg plus HD ASA significantly reduced the incidence of thrombosis.
These doses of xemilofiban or xemilofiban 1.25 mg/kg plus HD ASA
increased time to occlusion, inhibited ex vivo platelet aggregation
by >90%, and prevented or reduced CFVs. Xemilofiban 2.5 mg/kg but
not xemilofiban 1.25 mg/kg plus HD ASA significantly increased BT.
Key Words: platelet aggregation inhibitors • glycoproteins • thrombosis
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Introduction
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The binding of
adhesion protein ligands, such as fibrinogen and von Willebrand
factor, to activated platelets has been identified as the
final step in platelet aggregation, and this binding is completely
mediated by GP IIb/IIIa.1 There is abundant
evidence that the formation of platelet aggregates plays an
essential role in the pathogenesis of acute arterial
thrombosis and is associated with such vascular diseases as unstable
angina, transient ischemic attack, and myocardial
infarction.2 Thus, effective inhibition of
platelet activity and platelet aggregation is a primary target
for the development of drugs designed to treat various
cardiovascular diseases. Consequently, the continual
search for and development of antithrombotic agents that block
fibrinogen–GP IIb/IIIa binding on activated platelets is a
major goal of cardiovascular research.
Xemilofiban (SC-54684A; Figure 1
) is an
orally active antithrombotic agent that is currently in clinical
trials. After administration, xemilofiban is rapidly metabolized to
SC-54701. The hydrochloride salt (SC-54701A) of the active moiety of
xemilofiban is a potent inhibitor of fibrinogen binding to
platelets.3 This compound is highly selective
for the GP IIb/IIIa receptor compared with other integrins sharing the
same ß3-subunit.3 ASA is
the most convenient and widely tested antiplatelet agent, but this
drug possesses limited antithrombotic efficacy. Clinical studies show
that the incidence of acute myocardial infarction is reduced 40% by
ASA,4 yet the event may occur during ASA therapy
in 1% of individuals free of prior myocardial
infarction,5 in 4% of patients with unstable
angina,6 and in 24% of patients with prior
myocardial infarction.7 After successful
thrombolytic therapy, rethrombosis may occur in
patients even though the treatment regimen includes oral ASA and
anticoagulation with heparin.8 Thus, there is a
need for newer orally active agents with antiplatelet and
antithrombotic properties that would add to the therapeutic
armamentarium for thrombosis-related disorders. Previously, we have
shown that intravenous ASA and concomitant administration
of heparin reduce the dose of intravenous SC-54701A that
provides antithrombotic efficacy in a canine model of occlusive
coronary artery thrombosis.9
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Figure 1. Chemical structure of xemilofiban: ethyl
3S-[[4[[4(aminoiminomethyl)phenyl]amino]-1,4-dioxobutyl]amino]-4-pentynoate
monohydrochloride. Xemilofiban is also known as SC-54684A.
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Initial animal studies with xemilofiban demonstrated its
effectiveness as an antiplatelet agent.10
Accordingly, the present investigation determines the
antithrombotic activity of orally administered xemilofiban in a canine
model of occlusive coronary artery
thrombosis9 and tests the hypothesis that
targeted levels of inhibition of platelet aggregation by
xemilofiban prevent thrombotic occlusion. Moreover, the present
experiments determine the antithrombotic effect of a minimally
effective dose of xemilofiban combined with oral ASA.
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Methods
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Reagents
Xemilofiban was synthesized at Searle. Xemilofiban was
administered as the hydrochloride salt (SC-54684A), and the doses were
calculated on the basis of the free base (SC-54684, MW=358.4). A
1-mg/kg dose of the ester is 2.79 µmol/kg, which is equivalent
to 1.10 mg/kg of hydrochloride salt. SC-54701B, the trifluoroacetate
salt of the active moiety, was used in the in vitro experiments.
Concentrations of SC-54701B were determined on the basis of the free
base (SC-54701, MW=330.3). Generic oral ASA was purchased locally, and
soluble lysine ASA was obtained from Synthélabo. Collagen from
equine tendon was purchased from Chrono-log Corp. ADP, U46619 (a
thromboxane mimic), and sodium citrate were purchased from
Sigma Chemical Co. Arachidonic acid and
epinephrine were obtained from Bio/Data Corp.
TXB2, used as an indirect measurement of
TXA2 production, was measured by the
Bioanalytical Laboratory at Searle by use of a
TXB2 Enzyme Immunoassay Kit obtained from Cayman
Chemical Co.
Experimental Procedures
A schematic of the protocol for the experiments performed in
this study is presented in Figure 2.
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Figure 2. Schematic of experimental protocol used in study.
Xemilofiban (Xe), ASA, Xe plus ASA, or placebo (n=6 or 7 per group) was
administered before electrolytic induction of thrombosis in LCx.
Incidence of occlusive thrombus formation, time to arterial
occlusion, and inhibition of ex vivo platelet aggregation were
monitored.
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Oral Drug Administration
Male or female mongrel dogs (14 to 27 kg) were preselected on
the basis of general health and a platelet count of
200x103 to
250x103/µL. The dogs were fasted for 12 to 18
hours before the start of testing and were divided into the following
treatment groups: xemilofiban (single dose of 1.25, 2.5, 5.0, or 6
mg/kg, n=6/group); LD (81 mg) ASA, n=7; HD (162 mg) ASA, n=6;
xemilofiban 1.25 mg/kg plus LD ASA, n=6; and xemilofiban 1.25 mg/kg
plus HD ASA, n=6. Xemilofiban or placebo (sucrose, n=7) was
administered to conscious dogs by soft gelatin capsules. Throughout
this article, the dose of xemilofiban is 1.25 mg/kg when used in
combination with LD ASA or HD ASA.
Surgical Procedures
One hour after treatment, the experimental model was used
essentially as previously described.9 Briefly,
the dogs were anesthetized with sodium pentobarbital (30 mg/kg
IV, supplemented as needed) and placed on a respiratory pump
(Biological Research Apparatus). The right femoral artery
and vein were cannulated for monitoring arterial blood
pressure (Micron Instruments) and for withdrawing blood samples,
respectively. The heart was exposed, and the LCx was isolated.
The artery was instrumented with a Doppler flow probe for
monitoring blood flow velocity (Crystal Biotech), a stimulation
electrode, and a Goldblatt clamp. The clamp was adjusted to reduce the
reactive hyperemic response without affecting resting LCx blood
flow. The stimulation electrode was connected in series with a 12- to
112-k
variable resistor to the positive terminal of a 9-V
battery. The negative terminal of the battery was connected to an
electrode placed in a subcutaneous site. Thrombotic occlusion of the
LCx was initiated 15 minutes after surgical preparation by
application and maintenance of 250 µA of continuous current
to the stimulation electrode. The current lasted for 180 minutes unless
the dog died after LCx occlusion. Thrombotic occlusion occurred when
the LCx blood flow decreased to and remained at zero for a minimum of
30 minutes. During electrical stimulation, before an occlusive thrombus
was formed, the number and frequency (No./min) of CFVs were
recorded. The frequency of CFVs in coronary blood flow
preceding the formation of an occlusive thrombus was expressed as x100
because of the small values obtained. At the end of the experiment, the
dogs were euthanized with an overdose of sodium pentobarbital. Light
microscopy was used to assess mural thrombosis at the site of injury in
the LCx of selected dogs treated with xemilofiban. The section of the
LCx chosen for this procedure had no observable thrombus in the lumen
of the vessel.
Ex Vivo Platelet Aggregation and Platelet Counts
Before treatment, blood was drawn from the cephalic vein of the
dogs to determine platelet counts (S-Plus IV cell counter, Coulter)
and ex vivo collagen-induced platelet aggregation. Blood used for
platelet counts was collected into evacuated tubes containing 7.5%
EDTA, and blood for platelet aggregation was collected into
evacuated tubes containing 3.8% sodium citrate.9
Assays for the determination of inhibition of platelet aggregation
were repeated 1 hour after treatment while the dogs were conscious and
at 1-hour intervals while the dogs were anesthetized. The blood
was centrifuged for 6 minutes at 266g (Sorvall
Technospin R centrifuge; H-5094 rotor) at room temperature
(24°C), and PRP was removed. The remaining blood was
centrifuged for 10 minutes at 2000g at room
temperature, and PPP was removed. Samples were assayed on a PAP-4
platelet aggregometer (Bio/Data Corp) with PPP as the blank.
Platelet aggregation was performed by adding 50 µL of collagen
(33 or 40 µg/mL final concentration) to 450 µL of PRP and measuring
aggregation for 3 minutes. (The 2 concentrations of collagen were used
to produce maximum aggregation with different lots of collagen.)
Collagen was used because it provided a better relationship between
protection and inhibition than ADP in this model.
Bioassay for Plasma Levels of the Active Moiety of
Xemilofiban
The procedures of the bioassay were described
previously.9 Briefly, the bioassay used plasma
from treated dogs as the source of inhibitor to be tested
in vitro against normal (naive) platelets from donor dogs. PRP from
nontreated dogs was added to wells containing plasma samples from
treated dogs in a 96-well microtiter plate. ADP (20 µmol/L) was
added to the platelet suspension in each well to induce
aggregation. Optical density at 405 nm was measured on all wells
simultaneously in a plate reader (Molecular Devices). The
results were quantified by comparison to a standard inhibition curve
prepared in plasma with the use of known amounts of SC-54701A.
Template BT
To assess BT with the treatments that prevented occlusive
thrombus formation, a separate study was conducted in a separate group
of beagle dogs. Xemilofiban (2.5 or 6 mg/kg, n=4 per treatment) or
xemilofiban plus HD ASA (n=4) was administered to fasted, conscious
dogs as described above. Sixty minutes after treatment, the dogs were
anesthetized with sodium pentobarbital (30 mg/kg IV,
supplemented as needed). The effect of treatment on upper-lip BT was
observed after an additional 120 minutes (comparable to the time
allowed for treatment and surgery with the other dogs in the main
study). BT was measured with a Simplate IIR device (Organon Teknika
Corp). Duplicate tests were performed on each dog.
In Vitro Effect of Xemilofiban+ASA
In a separate study, PRP and PPP were prepared as previously
described. PRP (400 µL) was added to cuvettes containing 50 ng/mL
SC-54701B with or without ASA (100 µg/mL) or ASA alone. The choice of
the concentration of SC-54701B was based on plasma concentrations
measured at the time of arterial occlusion or at the end of
the experiment (see Table 2, xemilofiban 1.25 mg/kg). The concentration
of ASA used was based on calculations using HD ASA, average dog body
weight, and estimated plasma volume of the dogs. Controls without
SC-54701B or ASA were also carried out. After the addition of the
agonist, platelet aggregation was monitored for 2 minutes with ADP
(20 µmol/L) or arachidonic acid (0.2 mg/mL) and
for 3 minutes with collagen (40 µmol/L) or U46619 (100
µmol/L). [Maximal aggregation was achieved with 100 µmol/L
U46619 after priming with epinephrine (60 µmol/L).] The
contents of the cuvette were transferred to Eppendorf
centrifuge tubes containing 10 µL of ASA to prevent further
TXA2 production, and the tubes were
centrifuged at 12 000g for 3 minutes. Supernatants
were aspirated and frozen for determination of
TXB2 levels.
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Table 2. Plasma Levels of the Active Moiety of Xemilofiban
With the Related Ex Vivo Inhibition of Collagen-Induced Platelet
Aggregation
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Data Analysis
Unless otherwise stated, values are expressed as mean±SEM. The
1-tailed ¯
2 trend test was applied
to the rank-transformed data. Tukey's pairwise test (2-tailed) was
used for additional comparisons. The Mantel-Haenszel test was applied
only for the response "incidence of occlusive thrombus," which is
by its nature a discrete valued variable. An independent
t test was used to compare BT. A value of P<0.05
was considered to be statistically significant.
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Results
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Group Characteristics
Platelet counts and platelet aggregation were not
statistically different among the 9 groups of dogs before treatment.
Ventricular fibrillation occurred after complete thrombotic
occlusion in a dog treated with xemilofiban plus LD ASA. This dog was
not excluded from data analysis.
Prevention of Occlusive Thrombus
The effects of xemilofiban, ASA, or both on incidence of LCx
occlusion are shown in Table 1. Incidence of occlusion
was significantly reduced by 2.5, 5, or 6 mg/kg of xemilofiban. The LCx
remained patent in all dogs receiving 2.5 or 6.0 mg/kg of xemilofiban
and in 5 of 6 dogs treated with 5 mg/kg of the compound. The LCx
remained patent in only 1 dog treated with 1.25 mg/kg of xemilofiban
and 3 of 6 dogs treated with HD ASA. When the low dose of xemilofiban
was combined with HD ASA, no occlusive thrombus was formed during the
entire 180-minute study period. Typically, patent vessels, as expected,
showed disruption of the endothelial surface and
discontinuities in the internal elastic lamina (Figure 3). However, other than occasional
marginated neutrophils, platelets, and red blood cells, the lumens
were clear. All dogs treated with placebo developed complete thrombotic
occlusion. Figure 4 shows a schematic of
the patency status of the LCx for selected treatment groups.
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Figure 3. Patent section of coronary vessel after
electrical endothelial damage.
Endothelium and portions of internal elastic lamina
have been lost. On this surface are marginated neutrophils and red
blood cells but very few platelets or fibrin strands.
Notwithstanding these occasional accumulations of cells along damaged
vessel wall, this artery was patent at time of fixation. Toluidine
blue–stained 1-µm section, x480. Lumen is at top of figure.
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Figure 4. Schematic of LCx patency status in dogs treated
orally with selected doses of xemilofiban (Xe), Xe plus ASA, or placebo
control. Each horizontal bar represents response of 1 dog to
180 minutes of continuous anodal current. The current is started
approximately 3 hours after treatment. The open portions
represent periods of measurable blood flow continuously
monitored by a pulsed Doppler system, and the shaded portions
represent periods of LCx occlusion.
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Figure 5A shows the effect of the
treatments on inhibition of platelet aggregation at the time of
complete occlusion or at the end of the experiment (if LCx occlusion
did not occur). Inhibition of platelet aggregation was
significantly increased by xemilofiban (1.25 mg/kg, 59±15%; 2.5
mg/kg, 94±4%; 5 mg/kg, 98±2%; 6 mg/kg, 98±2%), ASA (LD, 30±12%;
HD, 78±9%), and the combined treatment (xemilofiban+LD ASA, 95±4%;
xemilofiban+HD ASA, 99±0%) compared with placebo (2±1%). Notably,
platelet aggregation was inhibited to a greater extent with
xemilofiban plus HD ASA than when either treatment was used alone.
Inhibition of aggregation by the 2 highest doses of xemilofiban or the
combined treatment was significantly greater than that of the 1.25
mg/kg dose of xemilofiban. All treatments, except 1.25 mg/kg of
xemilofiban or HD ASA, significantly increased inhibition of
platelet aggregation compared with LD ASA. Only 5 mg/kg of
xemilofiban produced a statistically significant increase in the
inhibition level relative to HD ASA.
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Figure 5. Effects of oral administration of xemilofiban
(Xe), ASA, Xe plus ASA, or placebo on inhibition of ex vivo
collagen-induced platelet aggregation (A) and on corresponding time
to occlusion of LCx (B). Time to zero flow of 180 minutes
represents maximum time of current application with no
thrombotic occlusion. n=6 or 7 per treatment group. Values are
mean±SEM. *P<0.05 vs placebo; +P<0.05
vs LD ASA; #P<0.05 vs HD ASA;  P<0.05
vs Xe 1.25 mg/kg.
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The effect of the treatments on time to zero flow in the LCx is shown
in Figure 5B
. Dogs with a patent LCx at the end of the experiment were
assigned a value of >180 minutes (censored) for time to zero flow, and
180 minutes was used for comparative purposes. The LCx of placebo dogs
occluded in 68±10 minutes. Compared with placebo, significant
prolongation of time to zero flow was noted in the dogs treated with
xemilofiban (2.5 mg/kg, >180 minutes; 5 mg/kg, 160±20 minutes; 6
mg/kg, >180 minutes) and in the group of dogs receiving the combined
treatment (xemilofiban+LD ASA, 139±25 minutes; xemilofiban+HD ASA,
>180 minutes).
CFVs During Anodal Stimulation of the LCx
A summary of the mean CFVs observed during stimulation of the LCx
is also presented in Table 1. Five of 6 dogs treated with
placebo had CFVs; the data on CFVs were not obtained from the remaining
dog because of technical problems. CFVs were not observed with dogs
treated with either 2.5 or 6 mg/kg xemilofiban. The CFV observed with 5
mg/kg xemilofiban was attributed to the dog that had a thrombotic
occlusion (see Table 1 and "Plasma Levels of the Active Moiety"
below). Neither ASA alone nor xemilofiban plus LD ASA significantly
reduced CFVs compared with placebo. No CFVs were observed with the
treatment of xemilofiban plus HD ASA.
Plasma Levels of the Active Moiety
Table 2 shows
that the active moiety was increased in a dose-dependent manner at the
time of thrombotic artery occlusion or at the end of the experiment. In
general, protection against occlusive thrombus formation was achieved
in all dogs when levels of the active moiety were >100 ng/mL (Figure 6). The dog with the occlusive thrombus
and the CFVs after treatment with 5 mg/kg xemilofiban had a maximum
xemilofiban plasma level of 31 ng/mL.
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Figure 6. Time to zero flow (ie, time to vessel occlusion)
and plasma levels of active moiety of xemilofiban for dogs treated with
xemilofiban. Each symbol represents individual dog. Time to
zero flow of 180 minutes represents maximum time of current
application with no thrombotic occlusion.
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Template BT
BT was assessed in duplicate in dogs treated with 2.5 or 6
mg/kg of xemilofiban and dogs treated with xemilofiban plus HD ASA
(Table 3). Both doses of
xemilofiban significantly prolonged BT relative to baseline. However,
the increase in BT by the lower dose of xemilofiban (the lowest dose
that provided antithrombotic protection in all dogs in the main study)
was modest. Three of the 4 dogs treated with 6 mg/kg of xemilofiban
exhibited a BT of >30 minutes. By contrast, BT was not significantly
lengthened by the combination treatment compared with baseline.
Mechanism of Increased Activity of Xemilofiban With ASA
Table 4 summarizes the
effects of SC-54701B alone, SC-54701B in combination with ASA, or ASA
alone on TXA2 production and platelet
aggregation with several agonists. SC-54701B inhibited
TXA2 production. SC-54701B was not as
effective as ASA alone in decreasing the level of
TXA2, but addition of ASA to SC-54701B increased
the inhibitory effect of the latter on
TXA2 production with all agonists tested.
ASA partially inhibited platelet aggregation induced by ADP,
collagen, and arachidonic acid, whereas SC-54701B alone
or in combination with ASA blocked aggregation.
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Table 4. TXA2 Production and Platelet Aggregation
with SC-54701B, ASA, SC-54701B in Combination With ASA or
Control
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Discussion
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The formation of thrombi, which usually constitutes the primary
event of occlusive cardiovascular disease, is mediated
by the binding of adhesion protein ligands, such as fibrinogen and von
Willebrand factor, to activated
platelets.11 Effective inhibition of
platelet aggregation and subsequent thrombus formation therefore
represents a target for pharmacological
intervention.12 The binding of fibrinogen to GP
IIb/IIIa receptors on activated platelets
represents the final common pathway for platelet
aggregation.13 Therefore, blockade of the
platelet fibrinogen receptor GP IIb/IIIa is an effective
antithrombotic strategy. Monoclonal antibodies,14
peptide mimetics,15 and nonpeptide
mimetics16 have been developed to antagonize this
complex. These compounds have to be administered
intravenously, which limits their therapeutic use to acute
thrombotic situations. An orally active compound would be desirable
because of ease of administration and greater patient compliance, and
treatment could be administered outside of the setting of a medical
facility. Recently, several orally active GP IIb/IIIa receptor
antagonists have been reported that inhibit platelet
aggregation and inhibit arterial thrombus
formation.17 18 19
Xemilofiban, currently undergoing clinical trials, is a prodrug of a
nonpeptide mimetic of the amino acid sequence
arginine-glycine–aspartic acid that is recognized by the GP IIb/IIIa
receptor. The active moiety of xemilofiban is a potent, specific
inhibitor of fibrinogen binding that blocks platelet
aggregation to all known stimuli.3 A previous
study demonstrated that administration of xemilofiban to conscious dogs
resulted in dose-dependent antiplatelet activity and a high level
of bioavailability.10 In this report, we assessed
for the first time the antithrombotic effectiveness of orally
administered xemilofiban or xemilofiban with ASA in a canine model of
coronary artery thrombosis. This experimental model allowed us
to determine the efficacy of the test agents for prevention of
occlusive thrombus formation in response to electrolytic damage of
the intimal surface of the LCx.
Administration of xemilofiban resulted in a dose-related inhibition of
ex vivo platelet aggregation in response to the agonist collagen.
This ability of xemilofiban resulted in the dose-dependent sustained
antithrombotic effect achieved in this study. The lowest dose of
xemilofiban that provided protection from occlusive thrombus
formation was 2.5 mg/kg. This dose of the compound was associated with
a 94% inhibition of platelet aggregation. The thrombotic occlusion
observed in 1 of the dogs treated with 5 mg/kg of xemilofiban was
attributed to poor absorption of the compound, because plasma levels of
the active moiety of xemilofiban measured 23 to 31 ng/mL throughout the
experiment. (These levels were substantially less than the 194 to 339
ng/mL measured in the remaining dogs in the group. No known factors
would increase the clearance of the active moiety and thereby result in
low plasma levels, and there are no reasons to suspect that there would
be altered metabolism or increased clearance in this
setting.) Selected arteries that remained patent after treatment with
xemilofiban alone were examined by light microscopy for the presence or
absence of mural thrombus. In general, the arteries had minimal mural
thrombosis consisting primarily of red blood cells, activated
platelets, and/or marginated monocytes.
The efficacy of ASA as an antiplatelet agent has been extensively
investigated, and it remains the most widely used and cost-effective
drug in the prevention of platelet
aggregation.20 ASA, however, is not universally
effective at inhibiting platelet-dependent coronary artery
thrombosis, because non–cyclooxygenase-dependent
platelet aggregation commonly occurs in vivo, and ASA activity is
limited to inhibition of the cyclooxygenase
pathway.21 The finding that ASA (the terms
"low" and "high" doses of ASA are used here to compare the 2
doses with each other and are not meant to refer to treatment levels of
ASA in either dogs or humans) was essentially ineffective in preventing
thrombosis in the LCx was expected, because we9
and others22 using this arterial
thrombosis model have previously shown that the effects of
intravenous ASA were limited in the prevention of
arterial thrombosis. This is probably because ASA works by
inhibiting cyclooxygenase and therefore blocks the
synthesis of the endoperoxides,
prostaglandins, and thromboxanes. ASA will not
eliminate all possible proaggregatory mechanisms, whereas inhibition of
fibrinogen binding to platelets represents the final common
pathway and blocks all platelet aggregation and thrombus formation.
The ineffectiveness of ASA as an antithrombotic agent has also been
reported in other canine models.23 24
Because patients requiring antithrombotic therapy often take ASA as
adjunctive treatment, we tested the effect of low-dose xemilofiban in
combination with oral ASA. Treatment with 1.25 mg/kg of xemilofiban or
HD ASA prevented LCx occlusion in 
50% of the dogs. When both
treatments were combined, the observed antithrombotic effect was
increased. Because the mechanisms of the
platelet-inhibitory effect of xemilofiban and ASA
differ, and each agent inhibited platelet aggregation to some
extent, one might expect that the combination of a suboptimal dose of
xemilofiban and ASA would result in an increased antiaggregatory
response. Increased antiaggregatory activity from the administration of
a GP IIb/IIIa receptor antagonist combined with ASA has
been reported by others.25 In the in vitro
mechanism study, SC-54701B inhibited TXA2 (a
potent agonist of platelet aggregation) production in a
dose-related manner. When a suboptimal dose of SC-54701B was combined
with ASA, increased inhibition of TXA2 was
observed. The ability of xemilofiban to prevent platelet
aggregation, resulting in an indirect decrease of
TXA2, coupled with the additional inhibition of
aggregation and TXA2 release by ASA, probably
contributed substantially to the mechanism by which high-dose ASA
potentiated the effect of 1.25 mg/kg of xemilofiban in dogs.
Although BT was significantly increased by xemilofiban in the
present study, BT does not predict surgical
bleeding.26 (BT was modestly lengthened by 2.5
mg/kg of xemilofiban, the minimum antithrombotic dose. A further
prolongation of BT was observed with 6 mg/kg of xemilofiban.) Results
from clinical trials have demonstrated that GP IIb/IIIa blockade
reduced ischemic/thrombotic events after coronary
intervention, but at the cost of an increased risk of bleeding
complications.27 This increased risk was partly
due to the concomitant administration of heparin. Concerns about
excessive bleeding in the setting of treatment with a GP IIb/IIIa
receptor antagonist (with ASA and heparin) have been
allayed by careful titration of heparin along with prudent vascular
access site management.28 Ticlopidine is an
example of an antiplatelet drug that prolongs BT without increasing
operative blood loss.29 Results from our main
study showed that a suboptimal dose of xemilofiban plus ASA did not
significantly alter BT, suggesting that administration of xemilofiban
with ASA may be used in the clinic for prevention of acute thrombotic
events without impairment of hemostasis.
Unlike the 67% (4 of 6) of the dogs with CFVs after treatment with the
1.25 mg/kg dose of xemilofiban or HD ASA, the combination of the two
treatments provided increased protection against CFVs. This effect was
similar to that produced by the 2.5- or 6-mg/kg dose of xemilofiban.
The mechanism of CFVs has been associated with the release of such
vasoactive substances as serotonin and
TXA230 and with periodic acute
occlusive platelet thrombus formation followed by
embolization.31
Orally active compounds are expected to be more readily employed for
preventive antithrombotic therapy. Xemilofiban has the potential to
meet this goal. The degree of platelet inhibition necessary for
this agent to be efficacious in the patient population is currently
being assessed in a variety of clinical trials. Sustained inhibition of
platelet aggregation with xemilofiban has been demonstrated in
patients with unstable angina32 and in patients
after coronary stent deployment.33
In summary, xemilofiban, a novel GP IIb/IIIa receptor
antagonist, inhibits ex vivo platelet aggregation,
prevents occlusive thrombus formation, and prevents CFVs in response to
continuous electrical injury in a canine model of arterial
thrombosis. In addition, coadministration of HD ASA provides an
observed increase in the antithrombotic response and in the level of
protection against CFVs by a dose of xemilofiban that when administered
alone has limited efficacy in this model.
|
Selected Abbreviations and Acronyms
|
|---|
| ASA |
= |
aspirin |
| BT |
= |
bleeding time |
| CFV |
= |
cyclic flow variation |
| GP |
= |
glycoprotein |
| HD |
= |
high-dose |
| LCx |
= |
left circumflex coronary artery |
| LD |
= |
low-dose |
| PPP |
= |
platelet-poor plasma |
| PRP |
= |
platelet-rich plasma |
| TX |
= |
thromboxane |
|
|
Acknowledgments
|
|---|
The authors would like to acknowledge the assistance of Dr David
Baron of the Department of Product Safety, Searle, Skokie, Ill, for
the histological assessment of mural thrombosis in the
patent arteries.
Received January 6, 1998;
revision received February 19, 1998;
accepted March 17, 1998.
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Abstract
Introduction