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Serial clopidogrel dose adjustment after platelet function testingimproves outcome of acute coronary syndrome patientsundergoing percutaneous coronary intervention with highon-treatment platelet reactivity
Jure Samardzic • Miroslav Krpan • Bosko Skoric •
Marijan Pasalic • Mate Petricevic • Davor Milicic
� Springer Science+Business Media New York 2014
Abstract High on-treatment platelet reactivity (HTPR)
on clopidogrel correlates with adverse outcomes in patients
treated with percutaneous coronary intervention (PCI).
Whether HTPR is a modifiable risk factor for future events
is not clear. We evaluated the effect of serial clopidogrel
dose adjustment based on platelet function testing (PFT)
during 12 months of dual antiplatelet therapy (DAPT)
using Multiplate� analyzer in patients with HTPR after PCI
in acute coronary syndrome on clinical outcome. Eighty-
seven patients were randomized to interventional (n = 43)
and control group (n = 44). Blood samples for PFT were
drawn at day 1, 2, 3, 7, 30 and at month 2, 3, 6, 9 and 12.
Clopidogrel dose was modified at each point of PFT in the
interventional group with patients taking up to two addi-
tional 600 mg loading doses and a range of 75–300 mg
maintenance dose to achieve and maintain optimal platelet
reactivity (19–46 U). The incidence of the primary end-
point (composite of cardiovascular death, non-fatal myo-
cardial infarction, target vessel revascularization and
ischemic stroke) was significantly higher in the control
group (36.3 vs 16.2 %; p = 0.034). There were no differ-
ences in total bleeding events (6.8 vs 4.6 %, p = ns).
Patients in the interventional group maintained better
P2Y12 inhibition during follow-up. We hypothesize that
targeting the therapeutic window of platelet reactivity
continuously throughout DAPT by dose adjustment of
P2Y12 inhibitor may lead to better platelet reactivity con-
trol, and thus reduce the rate of ischemic complications in
this high risk group of patients.
Keywords Clopidogrel � Platelet reactivity � Acute
coronary syndrome � Tailoring therapy � Outcome
Introduction
Dual antiplatelet therapy (DAPT) with aspirin and P2Y12
receptor antagonist during 12 months presents cornerstone
treatment in acute coronary syndrome (ACS) patients
undergoing percutaneous coronary intervention (PCI) [1–
4]. Despite the use of DAPT 9–12 % of patients still
develop new ischemic event after myocardial infarction
(MI) [5–7]. Clopidogrel is the most widely used P2Y12
inhibitor [8] despite it’s limitations that include highly
variable P2Y12-receptor inhibition which causes wide
interindividual platelet reactivity variations [9]. Since high
on-treatment platelet reactivity (HTPR) on clopidogrel is
strongly associated with adverse events [10], antiplatelet
therapy tailoring has been vastly investigated to determine
whether individualized approach could improve outcomes.
New P2Y12 inhibitors like prasugrel and ticagrelor produce
more potent and more uniform antiplatelet effect and thus
reduce the risk of adverse ischemic events in ACS patients
undergoing stent implantation better than clopidogrel but at
the cost of increased risk of major, non CABG related
bleeding [6, 7]. In the time of progressive personalized
approach to therapy, effective strategies are needed to
minimize the risk of ischemic adverse events without
increasing the risk for bleeding. We conducted a
J. Samardzic (&) � M. Krpan � B. Skoric � M. Pasalic �D. Milicic
Department of Cardiovascular Diseases, University Hospital
Center Zagreb, University of Zagreb School of Medicine,
Kispaticeva 12, 10000 Zagreb, Croatia
e-mail: [email protected]
M. Petricevic
Department of Cardiac Surgery, University Hospital Center
Zagreb, University of Zagreb School of Medicine,
Kispaticeva 12, 10000 Zagreb, Croatia
123
J Thromb Thrombolysis
DOI 10.1007/s11239-014-1087-0
Page 2
prospective, randomized study with the aim to investigate
whether serial clopidogrel dose adjustment according to
platelet reactivity measurements using Multiplate� ana-
lyzer (Roche Diagnostics, Mannheim, Germany) could
decrease the rate of adverse events in ACS patients treated
with PCI and with determined HTPR.
Methods
This study was approved by Ethics Committees of Uni-
versity Hospital Centre Zagreb and University of Zagreb
School of Medicine. All patients gave informed consent
before enrollment. Inclusion criteria were predefined as
follows: ACS patients aged 18–80, treated with aspirin,
clopidogrel and successful coronary stenting and with
HTPR phenotype determined by multiple electrode aggre-
gometry (MEA) 12–24 h after PCI. ACS was diagnosed
based on clinical presentation, ECG changes and troponin
T values. Exclusion criteria were continuous postinter-
ventional glycoprotein (GP) IIbIIIa receptor inhibitor per-
fusion, thrombocytopenia (\150 9 109/L), significant
renal insufficiency (creatinine [ 200 lmol/L), anemia
(Htc \ 30 %), hemorrhagic diathesis, history of intracra-
nial bleeding or ischemic cerebrovacular insult 6 months
before, major operation 6 weeks before, concomitant
chronic anticoagulation therapy and advanced age
([80 years). All patients were treated with aspirin and
clopidogrel as newer P2Y12 inhibitors were unavailable.
Patients presenting with STEMI (60.9 %) were subjected
to primary PCI and received 600 mg loading dose (LD) of
clopidogrel prior to the procedure. Patients presenting with
NSTEMI (22.9 %) were predominantly treated with
300 mg LD of clopidogrel and early invasive strategy
(\24 h). Patients presenting wih unstable angina—UA
(16.1 %) were predominantly treated with DAPT for a
couple of days before PCI. Patients with STEMI and
NSTEMI received loading dose of aspirin—300 mg prior
to PCI. Patients with UA received aspirin 100 mg/day
several days prior to PCI. All patients received 100 mg/day
maintenance dose of aspirin.
Platelet activity was measured 12–24 h following suc-
cessful PCI using MEA—one of several recommended
platelet function tests (PFT) [11]. Multiplate� analyzer is a
point of care device that uses whole blood sample for PFT.
Depending on the test that is used it can measure platelet
reactivity through different signaling pathways and thus,
measure the effect of different types of antiplatelet drugs.
Measuring platelet response to antiplatelet therapy using
MEA is based on the impedance, where a change of
resistance between two electrodes immersed into the test-
ing blood is determined after the addition of saline and a
specific aggregation agonist—in this case adenosine
diphosphate (ADP) with a final concentration of 6.4 lM.
The blood samples were taken for analysis in most cases
from the antecubital vein in the official MEA tubes which
are coated with hirudin, a direct thrombin inhibitor, which
does not interfere with platelet activation. After blood
collection, the tubes were immediately gently inverted
several times and tested after at least 30 min. According to
the manufacturer’s recommendations, all samples were
tested between 30 min and three hours after blood collec-
tion. The volume of blood required for performing one test
is 300 lL and the result is available in approximately
10 min (3 min for incubation and 6 min for the post-
stimulation measurement).
We screened 461 consecutive ACS patients. All patients
without exclusion criteria (341) underwent PFT. We
identified and enrolled 87 patients with HTPR in the study
(Fig. 1). Assignment to study groups was performed by
random allocation using randomization software (Research
Randomizer). All patients enrolled in the study were
hemodynamically stable at randomization. None of the
patients enrolled in the study had cardiogenic shock nor
IABP implanted at randomization. This is because patients
who presented on arrival with cardiogenic shock either
died before enrollment, had unsuccessful PCI or had
developed exclusion criteria on screening (i.e. renal
insufficiency). Furthermore, some patients with cardio-
genic shock who were eligible for PFT 12–24 h after
successful PCI were either in no condition to give inform
ACS patients treated with PCIn = 461
Patients with exclusion criteria n = 120
MEAtestingn = 341
MEA platelet reactivity≤ 46 U
n = 254 (74.5%)
HTPRn = 87 (25.5%)
Interventional groupn = 43
Control groupn = 44
Fig. 1 Flowchart of study design and patient selection (ACS acute
coronary syndrome, HTPR high on-treatment platelet reactivity, MEA
multiple electrode aggregometry, PCI percutaneous coronary
intervention)
S. Jure et al.
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consent for PFT (i.e. patients with out of hospital cardiac
arrest undergoing therapeutic hypothermia) or decided not
to participate in the study.
The enrolled patients were randomily assigned either to
clopidogrel tailoring group (interventinal group) or stan-
dard clopidogrel dose group (control group). Clopidogrel
dose was adjusted just as PFT results were obtained.
Control PFT and tailoring regimen in the first seven days
were performed in the morning before taking the therapy
according to the protocol described in Fig. 2. Cut off
values for HTPR and enchanced platelet response
were set according to the consensus statement at [ 46 U
and \ 19 U, respectively [12]. The patients in the inter-
ventional group recieved up to two additional loading
doses of 600 mg clopidogrel and up to 300 mg mainte-
nance dose of clopidogrel (150 mg maximum for patients
older than 70). Clopidogrel dose adjustment at 30 days and
on following PFT controls were performed in the same
manner as on day 7. The control group underwent PFT in
the same intervals but without clopidogrel dose adjustment.
Compliance to treatment and major adverse cardiac and
cerebrovascular events (MACCE) occurence were assesed
on control visits by interview and by counting the returned
clopidogrel tablets given to the patient at the last visit.
Primary endpoint was the composite of death, non-fatal
MI, target vessel revascularization and ischemic stroke.
Secondary endpoints were bleeding events according to
Bleeding Academic Research Consortium (BARC) defini-
tion [13].
Statistical methodology
All collected data were analyzed by descriptive statistics.
Categorical variables are shown as frequencies and prev-
alence. Metric variables are shown as arithmetic means
with standard deviations. Data are supplemented by tables
and figures. Kolmogorov–Smirnov test was used to test for
normal distribution of continuous data. Appropriate para-
metric and non-parametric tests were used according to the
results. Differences in individual characteristics between
the study and control groups were tested by Student’s t test
for independent samples metric variables and v2 test for
categorical variables. The difference in clinical outcome
MD decreaseif 300 mg decrease to 150 mgif 150 mg decrease to 75 mg
if 75 mg continue 75 mg
No changes in the MDMD increaseif 75 mg raise to 150 mgif 150 mg raise to 300 mgif 300 mg continue 300mg
(for patients > 70 years MD 150 mg max)
Day 7 - MEA testing
clopidogrel75 mg MD
clopidogrel150 mg MD
clopidogrel300 mg MD
clopidogrel 75 mg MDclopidogrel 150 mg MD< 19 U
19-46 U
19-46 U> 46 U
< 19 U
< 19 U≥ 19 U
≤ 46 U
> 46 U
> 46 U
Interventional group
Day 1 - clopidogrel 600 mg LD
clopidogrel 150 mg MDDay 2 - MEA testingclopidogrel 600 mg LD
Day 3 - MEA testing
Fig. 2 Flow chart of the personalized treatment with clopidogrel in the interventional group during the first week after coronary
revascularization (LD loading dose, MEA multiple electrode aggregometry, MD maintenance dose)
Serial clopidogrel dose adjustment
123
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between the two groups was tested by v2 test. The differ-
ences in the data between study groups were tested by
Student’s t test of independent samples or ANOVA for
multiple independent samples. Survival to MACCE and the
rate of ischemic events are shown by Kaplan–Meier curve.
P values \ 0.05 were regarded statistically significant.
Statistical analysis was performed using SPSS software
version 21 (IBM Corporation, USA).
Results
Eighty-seven patients were enrolled in the study: 43 in the
interventional group and 44 in the control group. Most
patients presented with STEMI (60.9 %), while NSTEMI
and UA were present in 22.9 % and 16.1 % of patients,
respectively. Most of the patients were clopidogrel naive at
admission (95.4 %). Aside from the extension of coronary
artery disease the groups did not differ significantly in
concomitant medical therapy, CYP2C19 genotype, demo-
graphic and procedural characteristics (Tables 1, 2). Pro-
portion of patients treated with lipophilic statin who
presented with STEMI, NSTEMI and UA and were
receiving initially intensified statin dose was 76.3, 66.6 and
14.3 %, respectively. During the follow-up 7 and 16
patients reached the primary outcome in the interventional
and control group, respectively (16.2 vs 36.3 %;
p = 0.034). Two patients were lost to follow-up in both
groups. Three and four patients died in the control and
interventional group, respectively. Three stent thromboses
(ST) occurred in the control group, while there was only
one ST in the interventional group (p = ns). Two patients
in the interventional and one in the control group experi-
enced ischemic stroke. There were no differences in total
bleeding outcomes; 6.8 and 4.6 % in the control and
interventional group, respectively (p = ns). One BARC
type 2 and type 5 bleeding event occurred in the inter-
ventional group, while there were two type 2 and one type
3 event in the control group. Patients in the interventional
group had a significantly better outcome and survival to an
adverse event (ischemic or bleeding) as shown in Fig. 3.
The proportion of patients with HTPR decreased in both
groups after randomization, but significantly more in the
interventional group (Fig. 4). Mean platelet reactivity of
patients in both groups is shown in Fig. 5. Intraindividual
platelet reactivity was noted in both study groups. Mean
difference between minimal and maximal platelet reactiv-
ity in each patient was 50 U (SD ± 17.47) and 53.46 U
(SD ± 16.71) in the control and interventional group,
respectively (Fig. 6). Platelet inhibition was mostly not
consistent in relation to the cut off point for HTPR as
52.3 % of patients crossed from a responder to a non-
responder at least three times (Tables 3, 4).
Discussion
To the best of our knowledge this is the first randomized
study that performed serial clopidogrel dose adjustment
during 12 months of DAPT to maintain the proposed range
of ideal platelet inhibition recommended for MEA [12]
based on receiver-operator characteristic (ROC) curve
analysis performed by Sibbing et al. that determined
Table 1 Baseline demographic characteristics of study patients
Patients’ characteristics Interventional
group
(n = 43)
Control
group
(n = 44)
p
Age, mean (SD) 63.37 (12.56) 63.48 (12.15) 0.968
Men, n (%) 22 (51.1) 27 (61.4) 0.338
Unstable angina, n (%) 8 (18.6) 6 (13.7)
0.801NSTEMI, n (%) 10 (23.3) 10 (22.7)
STEMI, n (%) 25 (58.1) 28 (63.6)
Arterial hypertension, n (%) 28 (65.1) 27 (61.4) 0.717
Hyperlipidemia, n (%) 25 (58.1) 20 (45.5) 0.236
Diabetes mellitus, n (%) 14 (32.6) 12 (27.3) 0.590
Smokers, n (%) 9 (20.9) 12 (27.3) 0.489
Family history for
CAD, n (%)
7 (16.3) 6 (13.6) 0.730
Previous MI, n (%) 6 (13.9) 3 (6.8) 0.314
Previous PCI, n (%) 6 (13.9) 2 (4.5) 0.157
Previous CABG, n (%) 0 (0) 0 (0) 1.000
Previous stroke, n (%) 1 (2.3) 4 (9.1) 0.360
PAD, n (%) 0 (0) 1 (2.3) 1.000
BMI, kg/m2, mean (SD) 28.93 (4.45) 27.90 (4.98) 0.313
Medical therapy
Clopidogrel naıve patients
prior to randomization,
n (%)
40 (93.1) 43 (97.7) 0.360
Statins, n (%) 40 (93.0) 31 (93.2) 0.689
Lipophilic statin, n (%) 21 (48.8) 30 (68.2) 0.150
PPIs, n (%) 14 (32.5) 22 (50.0) 0.151
Calcium channel
blocker, n (%)
6 (13.9) 7 (15.9) 1.000
CYP2C19*2, n (%)
Carrier homozygote 0 (0) 2 (4.8)
0.317Carrier heterozygote 16 (37.2) 13 (30.9)
Others 27 (62.8) 27 (64.3)
CYP2C19*17, n (%)
Carrier homozygote 4 (9.3) 4 (9.5)
0.996Carrier heterozygote 14 (32.5) 14 (33.3)
Others 25 (58.1) 24 (57.1)
BMI body mass index, CABG coronary artery bypass graft, CAD
coronary artery disease, MI myocardial infarction, NSTEMI non ST-
elevation myocardial infarction, PAD peripheral artery disease, PCI
percutaneous coronary intervention, PPI proton pump inhibitor, SD
standard deviation, STEMI ST-elevation myocardial infarction
S. Jure et al.
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optimal range of platelet reactivity on clopidogrel for MEA
[14, 15].
As the benefit of DAPT is consistent during 12 months
post PCI [16], we assumed that the serial PFT and dose
adjustment to prevail HTPR during 12 months would lead
to a better outcome. Our results suggest that serial clopi-
dogrel dose adjustment according to PFT throughout
12 months of DAPT leads to better platelet inhibition in
patients with initial HTPR after coronary stenting in ACS
as we observed that the control group had a significantly
larger rate of HTPR phenotype during 12 months of fol-
low-up (p \ 0.01). As the patients in the interventional
group had a better outcome, this might imply that PFT
might be useful in guiding antiplatelet therapy in patients
Table 2 Procedural characteristics of study patients
Angiographic and
PCI characteristics
Interventional
group
Control
group
p Angiographic and
PCI characteristics
Interventional
group
Control
group
p
Mean procedure
duration, min (SD)
36.05 (17.98) 38.23 (18.16) 0.575 Lesion type
De novo, n (%) 37 (86.0) 42(95.4)
0.226Bifurcation, n (%) 4 (9.3) 2 (4.5)
ISR/ST, n (%) 2 (4.6) 0 (0)
Treated arteries Stent type
One, n (%) 41 (95.3) 42(95.5)
0.981
BMS, n (%) 37 (86.0) 42 (95.4)0.157
Two, n (%) 2 (4.7) 2 (4.5) DES, n (%) 6 (13.9) 2 (4.5)
Treated lesions Thrombectomy, n (%) 7 (16.3) 8 (18.2) 1.000
One, n (%) 34 (79.1) 36 (81.8)0.748
Two, n (%) 9 (20.9) 8 (18.2)
CAD Mean referent artery
diameter, cm (SD)
3,05 (0.05) 3.18 (0.07) 0.153
Single vessel, n (%) 24 (55.8) 11 (25.0)
0.001Two vessel, n (%) 13 (30.2) 14 (31.8)
Three vessel, n (%) 6 (14.0) 19 (43.2)
Culprit lesion Mean lesion length,
mm (SD)
19,51 (9.86) 19.89 (7.84) 0.845
LAD, n (%) 16 (37.2) 16 (36.5) 0.935
LCx, n (%) 10 (23.2) 8 (18.2) 0.559
RCA, n (%) 19 (44.2) 22 (50.0) 0.587
Tortuosities, n (%) 2 (4.6) 5 (11.4) 0.434 Stenosis, % (SD) 95,26 (8.82) 94.80 (10.04) 0.821
Dominance TIMI flow before wire
Right, n (%) 34 (79.1) 30 (68.2)
0.488
0 17 (39.5) 21 (47.7)
0.791Left, n (%) 2 (4.7) 4 (9.1) 1 0 (0) 1 (2.3)
Equilibrated, n (%) 7 (16.2) 10 (22.7) 2 8 (18.6) 2 (4.5)
3 18 (41.8) 20 (45.4)
Stents implanted TIMI flow after wire
One, n 31 28
0.470
0 5 (11.6) 10 (22.7)
0.699Two, n 8 12 1 4 (9.3) 3 (6.8)
Three, n 2 3 2 10 (23.2) 6 (13.6)
Four, n 2 1 3 24 (55.8) 25 (56.8)
Mean width of the
narrowest
stent, mm (SD)
2.99 (0.064) 3.11 (0.082) 0.232 TIMI flow, final
0 0 (0) 0 (0)
0.6281 0 (0) 0 (0)
2 3 (6.9) 2 (4.5)
3 40 (93.0) 42 (95.4)
Mean total stent length,
mm (SD)
26.30 (14.39) 28.73 (15.86) 0.457
BMS bare metal stent, CAD coronary artery disease, DES drug eluting stent, ISR in-stent restenosis, LAD left anterior descending, LCx left
circumflex, PCI percutaneous coronary intervention, RCA right coronary artery, SD standard deviation, ST stent thrombosis, TIMI thrombolysis
in myocardial infarction
Serial clopidogrel dose adjustment
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presenting with ACS. The routine use of PFT is currenlty
not recommended for any PCI patients. It might be con-
sidered only in guiding antiplatelet therapy for patients
undergoing high-risk PCI or those with a known history of
stent thrombosis (ST) [11, 12].
Hazarbasanov et al. showed using MEA that additional
600 mg LD of clopidogrel and 150 mg clopidogrel main-
tenance dose (MD) during the first month after PCI
improved platelet inhibition in HTPR patients and their
outcome after six months. Their research group, however,
enrolled both ACS (57 %) and stable angina patients [17].
ACS patients’ platelet response change differs from stable
coronary artery disease (CAD) patients as ACS is a highly
hypercoagulabile state [18–20]. The degree of platelet
response to clopidogrel is influenced by more factors
beside ACS such as underdosing, compliance, comorbidi-
ties, concomitant therapy or CYP2C19 genotype [21–28].
Large, randomized studies addressing the intriguing issue
0102030405060708090
100
1 21 41 61 81 101
121
141
161
181
201
221
241
261
281
301
321
341
361
Control group
Interventional group
p=0.044
0
2
4
6
8
10
12
14
16
181 20 39 58 77 96 115
134
153
172
191
210
229
248
267
286
305
324
343
362
p=0.034
ba
Time (days)Time (days)
Pat
ient
s (n
o.)
Pat
ient
s (%
)
Control group
Interventional group
Fig. 3 a Kaplan–Meier analysis of cumulative ischemic events in both groups. b Kaplan–Meier survival analysis to an ischemic or a bleeding
event in both groupsP
atie
nts
with
HT
PR
(%
)
Interventional vs control group p<0.01
Interventional group
Control group
Day 1 Day 2 Day 3 Day 7 Day 30 Mo. 2 Mo. 3 Mo. 6 Mo. 9 Mo. 12
90
70
50
30
10
0
20
40
60
80
100Fig. 4 Rates of high on-
treatment platelet reactivity in
both groups of patients during
follow-up. HTPR high on-
treatment platelet reactivity
The
rape
utic
win
dow
AD
P -
U (
Uni
ts)
Day 1 Day 2 Day 3 Day 7 Day 30 Mo. 2 Mo. 3 Mo. 6 Mo. 9 Mo. 12
Interventional group
Control group
0
10
20
30
40
50
60
70
80Fig. 5 Mean platelet reactivity
in both groups of patients during
follow-up
S. Jure et al.
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of PFT in giuding antiplatelet therapy using VerifyNow�
(Accumetrics Inc., San Diego, California, USA) brought
negative results and setbacks in PFT [29, 30]. As those
trials recruited mostly stable coronary artery disease
patients, testing usefulness of PFT in tailoring therapy is
being switched to ACS patients—a population that really
might benefit with this strategy as they are at a higher risk
for adverse events. Currently, there are no completed large
randomized studies that have addressed this issue in ACS
patients. Previous studies have shown that platelet reac-
tivity decreases with time and that certain proportion of
patients still exhibit HTPR phenotype [29, 31]. In
GRAVITAS study, 62 % of patients with initial HTPR in
the standard-dose clopidogrel group remained low
responders to clopidogrel after 30 days [29]. Around 50 %
of patients in our control group kept HTPR phenotype
during the late post PCI period, as well. We also observed
significant intraindividual temporal variations in platelet
reactivity in both study groups. This implies that HTPR and
a personalized approach in antiplatelet treatment should
continue to be further extensively investigated. Even though
HTPR was shown to be an independent predictor of ische-
mic adverse events [32, 33] there is still no answer to the
question whether it is a modifiable factor. Our results imply
that maintaining proposed optimal platelet inhibition
throughout early and late period after coronary stenting in
ACS leads to a better clinical outcome. In the RECLOSE
2-ACS study, Parodi et al. showed using light transmission
aggregometry (LTA) that HTPR phenotype in ACS patients
after PCI was associated with increased risk of adverse
ischemic events at both short- and long-term follow-up
despite increased clopidogrel maintenance dose to
150–300 mg/day or switching to ticlopidine
(500–1,000 mg/day) [32]. Aradi et al. used MEA to tailor
antiplatelet therapy in ACS patients treated with PCI.
Patients with HTPR were assigned to either prasugrel or
high-dose clopidogrel. After one year, patients on prasugrel
had similar outcome as those without initial HTPR, while
patients on high-dose clopidogrel experienced more ische-
mic and bleeding events [34]. Design of our investigation
was based on the assumption that determining platelet
reactivity at more occasions would lead to a better insight of
platelet response during DAPT, better patient control, better
compliance, optimal treatment and better outcome. We
believe this approach to be reasonable as it is known that
platelet reactivity is a dynamic variable with inter- and in-
traindividial variations [35]. It is also known that HTPR is
associated with clopidogrel dose [36] and that the exact
timing to perform PFT has not been established [11, 12].
This could explain why the patients on high-dose clopido-
grel in study by Aradi et al. [34] experienced more ischemic
and bleeding events as some of those patients might have
been over or undertreated with clopidogrel. Analysis of the
ISAR-HPR registry performed by Mayer et al. with almost
one thousand patients treated with PCI and with determined
HTPR phenotype using MEA showed that patients who
recieved P2Y12 inhibitor dose adjustment based on MEA
had significantly lower incidence of ST and composite of
death without statistically significant increase in major
bleeding events in comparison to control cohort group after
30 days [37]. Siller-Matula et al. conducted a non-ran-
domized prospective study evaluating antiplatelet therapy
adjustment based on PFT using MEA. The authors enrolled
798 patients presenting mostly with stable CAD (63 %).
Patients in the guided group recieved up to four 600 mg
clopidogrel loading doses and prasugrel 60 mg loading dose
to overcome HTPR. After 30 days a significantly lower rate
of ST and MI was observed in the guided group without
significant differences in hemorrhagic events between study
groups [38]. The longest PFT study conducted was TRIL-
OGY-ACS Platelet Function Substudy which found no as-
socciation between PFT and clinical outcome throughout
30 months in patients with ACS initially treated without
revascularization [39]. This, however, can not yet be applied
to ACS patients treated with PCI as HTPR was shown to be
an independent risk factor for early and late ST [10].
0
20
40
60
80
100
120
140
160
0
20
40
60
80
100
120
140
160
Pla
tele
t rea
ctiv
ity -
AD
P (
U)
Pla
tele
t rea
ctiv
ity -
AD
P (
U)
PatientsPatients
a b
Fig. 6 Intra-individual platelet reactivity range during follow up. a Control group. b Interventional group
Serial clopidogrel dose adjustment
123
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Table 3 Intra-individual responder/non responder crossovers in the interventional group
Patient Day 1 Day 2 Day 3 Day 7 Mo. 1 Mo. 2 Mo. 3 Mo. 6 Mo. 9 Mo. 12 Responder/
nonresponder
crossover (n)
1 48 U B46 U HTPR HTPR B46 U B46 U B46 U B46 U B46 U B46 U 3
2 48 U B46 U B46 U B46 U HTPR B46 U B46 U B46 U B46 U HTPR 4
3 93 U HTPR HTPR HTPR B46 U B46 U HTPR HTPR B46 U B46 U 3
4 53 U B46 U B46 U B46 U B46 U B46 U B46 U B46 U B46 U B46 U 1
5 51 U B46 U B46 U HTPR B46 U HTPR HTPR B46 U HTPR HTPR 6
6 53 U B46 U B46 U B46 U B46 U B46 U B46 U B46 U B46 U HTPR 2
7 57 U B46 U B46 U B46 U ND ND ND ND ND ND 1
8 76 U B46 U B46 U HTPR B46 U B46 U B46 U B46 U B46 U B46 U 3
9 53 U B46 U B46 U B46 U B46 U HTPR B46 U HTPR B46 U HTPR 6
10 94 U B46 U B46 U HTPR B46 U B46 U B46 U B46 U B46 U B46 U 3
11 47 U B46 U B46 U HTPR B46 U B46 U B46 U B46 U B46 U B46 U 3
12 55 U B46 U B46 U B46 U B46 U HTPR B46 U B46 U B46 U B46 U 3
13 89 U HTPR HTPR HTPR ND ND ND ND ND ND 0
14 47 U HTPR B46 U B46 U B46 U B46 U B46 U B46 U B46 U B46 U 1
15 50 U B46 U B46 U B46 U HTPR HTPR ND ND ND ND 2
16 74 U HTPR HTPR HTPR HTPR B46 U B46 U HTPR B46 U B46 U 3
17 76 U B46 U B46 U HTPR B46 U HTPR B46 U B46 U B46 U B46 U 5
18 66 U B46 U B46 U B46 U B46 U B46 U ND ND ND ND 1
19 96 U HTPR HTPR HTPR HTPR HTPR HTPR HTPR HTPR HTPR 0
20 54 U B46 U B46 U B46 U B46 U B46 U B46 U ND ND ND 1
21 70 U B46 U B46 U B46 U B46 U B46 U B46 U B46 U B46 U B46 U 1
22 60 U HTPR HTPR HTPR HTPR HTPR B46 U B46 U B46 U B46 U 1
23 52 U B46 U HTPR B46 U HTPR HTPR B46 U HTPR HTPR HTPR 6
24 51 U B46 U HTPR HTPR B46 U ND ND ND ND ND 3
25 57 U HTPR HTPR HTPR B46 U B46 U B46 U HTPR HTPR HTPR 2
26 62 U B46 U B46 U HTPR B46 U B46 U B46 U B46 U B46 U B46 U 3
27 77 U HTPR HTPR HTPR HTPR HTPR B46 U B46 U HTPR HTPR 2
28 59 U B46 U B46 U HTPR HTPR B46 U B46 U B46 U HTPR B46 U 4
29 49 U B46 U B46 U HTPR HTPR HTPR HTPR B46 U B46 U HTPR 4
30 75 U B46 U B46 U HTPR B46 U HTPR HTPR HTPR HTPR HTPR 4
31 68 U HTPR B46 U HTPR B46 U B46 U B46 U HTPR B46 U B46 U 5
32 63 U HTPR HTPR HTPR HTPR B46 U B46 U B46 U B46 U B46 U 1
33 49 U B46 U B46 U B46 U B46 U B46 U B46 U B46 U B46 U B46 U 1
34 79 U HTPR B46 U B46 U HTPR HTPR HTPR HTPR HTPR HTPR 2
35 48 U B46 U HTPR HTPR B46 U B46 U B46 U B46 U HTPR B46 U 5
36 48 U B46 U B46 U B46 U B46 U B46 U B46 U B46 U HTPR B46 U 3
37 49 U B46 U HTPR B46 U B46 U B46 U B46 U HTPR B46 U B46 U 5
38 48 U B46 U B46 U B46 U HTPR B46 U B46 U HTPR B46 U HTPR 6
39 48 U HTPR B46 U B46 U HTPR B46 U B46 U HTPR B46 U HTPR 6
40 48 U B46 U B46 U B46 U B46 U B46 U B46 U B46 U B46 U B46 U 1
41 63 U B46 U B46 U B46 U B46 U HTPR HTPR HTPR HTPR HTPR 2
42 64 U B46 U B46 U B46 U B46 U B46 U HTPR B46 U B46 U B46 U 3
43 71 U B46 U B46 U B46 U HTPR HTPR HTPR HTPR HTPR HTPR 2
HTPR high on-treatment platelet reactivity, ND not done
S. Jure et al.
123
Page 9
Table 4 Intra-individual responder/non responder crossovers in the control group
Patient Day 1 Day 2 Day 3 Day 7 Mo. 1 Mo. 2 Mo. 3 Mo. 6 Mo. 9 Mo. 12 Responder/
nonresponder
crossover (n)
1 98 U HTPR B46 U HTPR B46 U B46 U B46 U B46 U B46 U B46 U 3
2 66 U HTPR HTPR HTPR HTPR HTPR HTPR HTPR HTPR HTPR 0
3 60 U B46 U B46 U HTPR B46 U B46 U B46 U B46 U B46 U B46 U 3
4 62 U HTPR B46 U HTPR B46 U HTPR HTPR B46 U HTPR HTPR 6
5 49 U B46 U B46 U B46 U B46 U B46 U HTPR HTPR HTPR HTPR 2
6 48 U HTPR HTPR B46 U B46 U B46 U B46 U B46 U B46 U B46 U 1
7 56 U B46 U B46 U HTPR B46 U HTPR ND ND ND ND 4
8 48 U B46 U B46 U B46 U B46 U B46 U HTPR ND ND ND 2
9 70 U HTPR B46 U HTPR HTPR HTPR HTPR HTPR HTPR HTPR 2
10 49 U B46 U B46 U B46 U B46 U HTPR B46 U B46 U B46 U B46 U 3
11 58 U B46 U B46 U B46 U B46 U B46 U HTPR B46 U HTPR HTPR 4
12 61 U HTPR B46 U HTPR HTPR HTPR HTPR HTPR HTPR HTPR 2
13 51 U B46 U HTPR B46 U B46 U B46 U B46 U B46 U B46 U B46 U 3
14 80 U B46 U B46 U HTPR B46 U B46 U HTPR HTPR HTPR HTPR 4
15 58 U HTPR HTPR HTPR HTPR HTPR HTPR HTPR HTPR HTPR 0
16 54 U B46 U HTPR B46 U HTPR ND ND ND ND ND 4
17 77 U HTPR HTPR HTPR B46 U B46 U HTPR B46 U HTPR B46 U 5
18 49 U HTPR HTPR HTPR HTPR HTPR HTPR HTPR HTPR HTPR 0
19 47 U HTPR HTPR HTPR HTPR HTPR B46 U B46 U HTPR B46 U 3
20 47 U HTPR B46 U HTPR B46 U B46 U HTPR HTPR HTPR HTPR 4
21 48 U B46 U HTPR HTPR B46 U HTPR HTPR B46 U B46 U HTPR 6
22 65 U HTPR B46 U B46 U B46 U B46 U B46 U B46 U B46 U HTPR 2
23 68 U HTPR HTPR HTPR B46 U B46 U B46 U B46 U B46 U B46 U 1
24 56 U B46 U B46 U HTPR HTPR B46 U B46 U B46 U B46 U B46 U 3
25 53 U HTPR HTPR HTPR HTPR HTPR HTPR HTPR HTPR HTPR 0
26 48 U HTPR HTPR HTPR HTPR HTPR HTPR HTPR HTPR HTPR 0
27 50 U B46 U B46 U HTPR B46 U B46 U B46 U B46 U HTPR HTPR 4
28 64 U HTPR HTPR HTPR B46 U HTPR HTPR HTPR HTPR HTPR 2
29 70 U HTPR HTPR B46 U B46 U B46 U B46 U B46 U B46 U B46 U 1
30 47 U HTPR HTPR B46 U HTPR HTPR HTPR B46 U HTPR HTPR 4
31 92 U HTPR HTPR HTPR HTPR B46 U HTPR B46 U B46 U HTPR 4
32 52 U B46 U B46 U B46 U B46 U B46 U B46 U B46 U B46 U B46 U 1
33 74 U HTPR HTPR HTPR HTPR HTPR HTPR HTPR HTPR HTPR 0
34 53 U HTPR HTPR HTPR B46 U HTPR HTPR HTPR HTPR HTPR 2
35 72 U HTPR HTPR HTPR HTPR HTPR HTPR HTPR HTPR HTPR 0
36 61 U HTPR B46 U B46 U B46 U B46 U B46 U B46 U B46 U B46 U 1
37 82 U HTPR HTPR HTPR HTPR HTPR HTPR HTPR HTPR HTPR 0
38 49 U B46 U HTPR HTPR HTPR HTPR HTPR HTPR B46 U HTPR 4
39 89 U HTPR HTPR HTPR HTPR HTPR HTPR HTPR HTPR HTPR 0
40 51 U HTPR HTPR HTPR HTPR HTPR HTPR HTPR HTPR HTPR 0
41 62 U HTPR HTPR HTPR B46 U B46 U B46 U B46 U B46 U B46 U 1
42 47 U HTPR HTPR B46 U B46 U B46 U HTPR B46 U B46 U B46 U 3
43 47 U ND ND ND ND ND ND ND ND ND NA
44 48 U B46 U B46 U ND ND ND ND ND ND ND 1
HTPR high on-treatment platelet reactivity, NA not applicable, ND not done
Serial clopidogrel dose adjustment
123
Page 10
Using vasodilator-stimulated phosphoprotein (VASP)
Bonello et al. showed that repeated clopidogrel loading
doses to achieve optimal platelet reactivity before stent
placement decreased the rate of ischemic events after one
month without increasing the risk of bleeding in stable
CAD patients [40]. We used a similar protocol of repeated
clopidogrel LD for persistent low responders in interven-
tional group in the first three days after PCI. We observed
that maintaining platelet reactivity in therapeutic window
range by tailoring clopidogrel dose according to MEA
during 12 months of follow-up in ACS patients with initial
HTPR reduced the incidence of adverse events, and thus
adequately protected the patients from future ischemic
events without increasing the risk of bleeding. Therefore,
our results support the hypothesis that HTPR might be a
modifiable risk factor and that serial use of PFT to optimize
platelet reactivity below HTPR threshold to a P2Y12
inhibitor might reduce adverse outcomes in these patients
without inceasing the risk of bleeding. We believe that
optimization of antithrombotic regimens might be impor-
tant both during hospitalization and after discharge in these
high risk patients.
There are several limitations to this study. First, there
was a significant difference in the extension of CAD
(number of diseased vessels) between interventional and
control group that might have had notable influence on
patient outcomes. Chirumamilla et al. showed that greater
extension of CAD is associated with increased platelet
reactivity and could actually be a cause to HTPR pheno-
type [41]. Furthermore, there was a statistically significant
correlation between the complexity of CAD and primary
endpoints in the total study population (p \ 0.05). This
indicates that significantly lesser extent of CAD in the
interventional group might have influenced better control
of platelet reactivity as well as better outcome. Exclusion
of the most sick patients in the study could be another
limitation. Finally, a relatively small patient sample cannot
guarantee significant power, especially for the safety
endpoint.
These results should be confirmed in large, similarly
designed, randomized trials with the use of new antiplatelet
agents as well. The clinical benefit of individualized
treatment based on PFT in ACS patients is still not ade-
quatelly investigated. Large expectations of implementing
personalized antiplatelet treatment based on PFT in ACS
patients are now focused on ongoing trials such as ANT-
ARTIC and TROPICAL ACS.
Acknowledgments This investigation was funded by the Croatian
Ministry of Science, Education and Sports.
Conflict of interest The authors declare that they have no conflict
of interest.
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