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Vol 9, Issue 1, 2016 ISSN - 0974-2441
HEMOSTASIS PROFILE AND CLINICAL OUTCOME OF ACUTE ISCHEMIC STROKE PATIENTS
TREATED WITH ORAL LUMBROKINASE DLBS1033: A COMPARATIVE STUDY VERSUS ASPIRIN
AND CLOPIDOGREL
ISMAIL SETYOPRANOTO1, SAMEKTO WIBOWO1, RAYMOND R TJANDRAWINATA2*
1Department of Neurology, Faculty of Medicine, University of Gadjah Mada/Dr. Sardjito Hospital, Yogyakarta, Indonesia. 2Dexa
Laboratories of Biomolecular Sciences, Dexa Medica Group, Indonesia. Email: raymond@dexa-medica.com
Received: 20 August 2015, Revised and Accepted: 30 September 2015
ABSTRACT
Objectives: This clinical study was conducted to determine the hemostasis profile and clinical outcome of acute ischemic stroke patients treated with
DLBS1033 in comparison with aspirin or clopidogrel. DLBS1033 is a proprietary bioactive protein fraction derived from the earthworms (Lumbricus
rubellus) that possesses both fibrinolytic and antithrombotic properties.
Methods: This was a 3-arm, parallel, randomized, controlled, open-label, blinded-evaluation study involving 126 acute ischemic stroke patients. Each
subject received any of the following study medication within 96 hrs after the stroke onset: Aspirin 80 mg daily (Group 1), or clopidogrel 75 mg daily
(Group 2), DLBS1033 490 mg 3 times daily (Group 3), for 90 days. Hemostasis parameters evaluated were prothrombin time (PT), activated partial
thromboplastin time (aPTT), and international normalized ratio (INR), while the clinical outcomes were measured using Gadjah Mada Stroke Scale
(SSGM) and Barthel index (BI).
Results: Baseline characteristics, including the hemostasis and clinical profiles, were comparable between groups. At the end of the study, PT, aPTT,
and INR values were not significantly different between groups, which were all within the normal ranges. There was a significant improvement of BI
as well as SSGM from baseline in each group. The improvement size of BI was found similar between groups (p=0.098). However, a significantly better
improvement of SSGM was observed in those received DLBS1033 (6.98±4.90; p=0.001 vs. aspirin [3.74±3.66], p=0.006 vs. clopidogrel [4.26±4.21]).
Conclusion: It was concluded that DLBS1033 provided a safe hemostasis profile (PT, aPTT, and INR) comparable to that of aspirin or clopidogrel in
ischemic stroke patients. Treatment with DLBS1033 improved clinical outcomes indicated by the BI and SSGM, and the improvement size of SSGM was
even better than that of aspirin or clopidogrel treatment.
Keywords: Acute ischemic stroke, Barthel index, DLBS1033, Hemostasis, Gadjah Mada Stroke Scale, Oral lumbrokinase.
INTRODUCTION
Thrombosis remains involved in the pathological course of some most
common vascular diseases, including ischemic stroke [1,2]. Substantial
progress has been made in understanding the biology of thrombus
formation and the pathophysiology of thrombosis. Several more established
pharmacologic agents, including thrombolytic therapy, antiplatelet agents,
and anticoagulants, have been recommended for the early management
of acute ischemic stroke [3]. However, all the recommended, as well as
neuro-protective agents available for prevention or treatment that have
been in use for decades, have currently been replaced with newer variants
that offer a modest incremental improvement [4-10]. Yet, the ideal drug
for prophylaxis and treatment of thrombotic disease that will inhibit
the thrombosis but not the hemostasis remains scarce. This situation is
complicated further by the emerging resistance to therapy with the most
established antiplatelets, aspirin, and clopidogrel that brings potentially
severe consequences such as recurrent myocardial infarction, stroke, or
death [11-13]. Such a reduced sensitivity to antiplatelet drugs was even
reported to be more remarkable in diabetic as compared to non-diabetic
patients [14]. Thus, an expedite translation of new knowledge from “test-
tube,” and animal studies to bedside pharmaceutical development should
be pursued for new and more strategic advances in the prevention of
thrombotic diseases.
For thousands of years, earthworms have widely been used in Indonesia,
China, Japan, and the Far East to treat various chronic diseases.
A group of serine protease enzymes collectively called lumbrokinase
extracted from the earthworms of Lumbricidae family could directly
dissolve fibrin and activate plasminogen [15,16]. Lumbrokinase
possesses strong fibrinolytic and fibrinogenolytic properties, lowers
blood viscosity, markedly inhibits platelet aggregation, and promotes
thrombus degradation without causing excessive bleeding [17-19].
Lumbrokinase is stable over a wide range of pH and temperature; thus,
it can be administered orally [20].
DLBS1033 investigated in this clinical study is a lumbrokinase
fractionated from the earthworms, Lumbricus rubellus, through a
proprietary technology of extraction. DLBS1033 possesses eight
major proteins, each with a molecular weight below 100 kDa, named
as Lumbricus low-molecular-weight proteins [21]. This specific
pattern of proteins confers a unique characteristic of DLBS1033
with its mechanism of action as an antithrombotic and thrombolytic
agent. The antithrombotic and thrombolytic activities of the bioactive
protein fraction have been demonstrated in vitro and ex vivo [21].
Furthermore, DLBS1033 has also been proven for its safety profile,
through toxicological studies in animal [22], and safety studies in
human [23,24]. To date, DLBS1033 has been approved by National
Agency of Drug and Food Control, Republic of Indonesia, to be marketed
as Indonesian standardized herbal medicine. Since then, no clinically
significant adverse drug reactions have been reported.
This current study was conducted to investigate whether the virtues
of DLBS1033 that have been demonstrated preclinically would also
be translated into clinical benefits for ischemic stroke patients. In this
study, we evaluated the hemostasis profile as well as functional and
neurological outcomes of ischemic stroke patients receiving DLBS1033,
in comparison with the more established antiplatelet agents used in
such cases, aspirin and clopidogrel.
Research Article
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Setyopranoto et al.
172
METHODS
Study design
This study was conducted in compliance with the Declaration of
Helsinki and the Good Clinical Practice. The study protocol was
reviewed and approved by the Independent Ethics Committee of
Gadjah Mada University, Yogyakarta, Indonesia, prior to trial initiation.
This was a 3-arm, parallel, randomized, controlled, open-label,
blinded-evaluation (PROBE-designed) clinical study, over 90 days of
treatment, to determine the hemostasis profile as well as functional
and neurological outcomes of ischemic stroke patients. Eligible subjects
were randomly allocated to receive any of the following regimens:
Aspirin 80 mg once daily (Group 1, Aspirin), clopidogrel 75 mg once
daily (Group 2, Clopidogrel), or DLBS1033 490 mg three times daily
(Group 3, DLBS1033). In this study, both aspirin and clopidogrel served
as the positive controls to examine the efficacy and safety of the new
treatment with DLBS1033.
Subjects were acute ischemic stroke patients admitted in the Stroke
Unit or Neurology-Ward of the Central General Hospital Dr. Sardjito
Yogyakarta, Indonesia. The inclusion criteria included: (1) adult male
or female; (2) acute ischemic stroke diagnosed by cranial CT-scan;
(3) having admitted in the hospital within less than 96 hrs after the
onset of stroke; and (4) willingness to participate in the study and give
subject’s written informed consent. The patients were excluded if any
of the following criteria was met: (1) recurrent stroke; (2) transient
ischemic attack; (3) intra-cerebral or subarachnoid hemorrhagic stroke;
(4) undefinable stroke onset; (5) regular therapy with antiplatelets,
anticoagulants; (6) hemostasis or coagulation disorders; (7) major
surgery within the last 6 months or having to have a surgery within
the next 3 months; (8) renal impairment defined as serum creatinine
level >3× upper limit of normal or history of hemodialysis; (9) systemic
inflammatory response syndrome; (10) unconsciousness; or
(11) uncontrolled hypertension (systolic blood pressure > 185 mmHg
or diastolic blood pressure > 110 mmHg).
Subjects were withdrawn from the study if any of the following
conditions applied during the course of the subject’s participation:
(1) Worsened prognosis, unconsciousness, and neurological deficits;
(2) hypersensitive to any of the study medication; (3) bleeding adverse
events, including nausea and vomiting, severe headache, gastrointestinal
pain, hematoma, and any of bleeding signs; and (3) subjects were
suffering from any disease, including physical accidents that would
interfere with the evaluation of the hemostasis profile.
Study medication
The management of acute ischemic stroke in the Unit Stroke of
Dr. Sardjito Yogyakarta Hospital was in accordance with the acquired
hemophilia A (AHA)/ASA Guidelines for the Early The Management
of Patients with acute ischemic stroke.3 Aspirin (Thrombo Aspilets®
enteric-coated tablets, Medifarma Laboratories Inc., Jakarta, Indonesia)
at the dose of 80 mg once daily, clopidogrel (Vaclo® film-coated
tablets, Dexa Medica, Palembang, Indonesia) 75 mg [25], once daily,
or DLBS1033 490 mg (Disolf® enteric-coated tablets, Dexa Medica,
Palembang, Indonesia) three times daily was initially administered to
the eligible subjects within 96 hrs after the stroke onset, to be taken
for the next 90 days. Concomitant medications taken by study subjects
along the study were valsartan 80/160 mg daily and/or amlodipin
5/10 mg daily (by hypertensive subjects); (2) insulin therapy (by
diabetic subjects); (3) simvastatin 20 mg daily (all subjects); and (4) oral
citicholine 1000 mg twice daily (all subjects). After study completion,
subjects continued their therapy in accordance with the management
for ischemic stroke applicable in the hospital.
Characterization of DLBS1033 was as previously described by Trisina
et al. [21]. A pharmaceutical study has also been conducted to support
its oral enteric-coated formulation and dosing regimen [26].
Randomization code was prepared using the permuted 3-block-
allocation and random numbers generator. In order to keep the
blinded-evaluation procedures, the study products were administered
by a particularly designated nurse, and the Investigator who evaluated
the outcomes was blind of the subject allocation. Blinding codes
were disclosed when the study had been completed, and trial-related
database had been frozen.
Hemostasis profile and clinical outcomes
Hemostasis parameters evaluated in this study were prothrombin time
(PT), activated partial thromboplastin time (aPTT), and international
normalized ratio (INR). Adverse events were observed and recorded
during the study conduct. The aPTT is an appropriate parameter to
evaluate the intrinsic and common pathways of the coagulation cascade.
The PT (or can be expressed as INR) evaluates the extrinsic and common
pathways [27]. Therefore, measurement of PT (or INR), or aPTT can be
beneficial to detect the abnormal steps in the coagulation cascade, for the
diagnosis of coagulation disorders [28]. Since we expected that therapy
with DLBS1033 as an antithrombotic/thrombolytic agent should not
impair the hemostasis profile, we hypothesized that the bioactive fraction
would not significantly interfere with those coagulation parameters.
The functional outcomes evaluated in this study were Barthel index
(BI) [29-33]. All variables were measured at baseline (day 0, before
study medication) and day 90 (end of study). The BI is based on a rating
scale that is completed by an observer, and it has a good inter-rater
reliability [30]. The items can be divided into a group that is related to
self-care (feeding, grooming, bathing, dressing, bowel and bladder care,
and toilet use) and a group related to mobility (ambulation, transfers,
and stair climbing). A total of 10 activities are scored, and the values are
then added to give a total score ranging from 0 (totally dependent) to 100
(completely independent). Lower scores indicate greater dependency.
Scoring on the BI can be interpreted as follows: Independent (BI score
of 80-100), needs minimal help (60-79), partially dependent (40-59),
very dependent (20-39), and totally dependent (<20). The BI measures
what the patient actually does rather than what they can do [30-32].
Neurological outcome in this study was measured by the validated
Gadjah Mada Stroke Scale (SSGM) [34]. The SSGM consists of 14 items,
including level of consciousness, orientation, language, visual field, eye
movement, facial movement, motoric function of the arms and limbs
both the passive and active (affected) sides, each of which is scored 0
(severe impairment) to 2 or 3 (normal, without impairment), resulting
in a total score ranging from 0 (the most severe neurological deficit)
to 38 (complete recovery) as shown in Table 1. The interpretation of
the SSGM is that a score of ≥23 means mild to moderate neurological
deficits, and a score of <23 means severe neurological deficits [34].
Statistical analysis
Demography and baseline characteristics were tabulated and
summarized by the group. Analyzes on hemostatic parameters as well as
clinical outcomes were carried out on intent-to-treat (ITT) population,
consisting of all patients who were randomized, exposed to at least one
dose of the study product, and then returned for at least one visit after
treatment-initiation. This would include data from withdrawn subjects.
All of the data are expressed as mean±standard deviation unless
otherwise is specified. Comparability of baseline characteristics
between groups was statistically analyzed by Kruskal–Wallis and Chi-
square, for continuous and categorical data, respectively. Hemostasis
and efficacy parameters (PT, aPTT, INR, SSGM, and BI) were statistically
analyzed between groups using Kruskal–Wallis and within-group using
Wilcoxon signed-rank. All statistical tests were at 5% significance level.
The SPSS® version 14.0 was used for the analyzes.
RESULTS
The clinical study had been conducted in the Stroke Unit or Neurology-
Ward of the Central General Hospital Dr. Sardjito Yogyakarta since May
2012 until December 2013. A total of 129 subjects were enrolled and
randomly allocated into any of the 3 arms, each of which consisted of
43 subjects. Of 129 subjects, only 126 were available for ITT analysis.
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Three subjects (one subject in each group) moved out of town and were
lost to follow-up. They did not have any post-treatment data, thus were
not evaluable.
Baseline characteristics of the study subjects were comparable between
groups as shown in Table 2. In all groups, subjects were aged between
50 and 65 years old, with male predominance (around 70% of subjects
in each group).
Onset of stroke, BI, PT, aPTT, and INR, as well as other risk factors, such
as plasma glucose, lipid profile, blood pressure, were also comparable
between groups at baseline. In terms of SSGM, at baseline, subjects in
DLBS1033 Group showed a slightly lower (worse) score (p=0.030) than
those in the aspirin group. Therefore, the real between-group difference
in neurological outcome was also evaluated by comparing the size of
improvement of SSGM from baseline, in addition to the final score of
SSGM at the end of study.
Table 1: SSGM [34]
Tested item Responses and scores
Level of consciousness 3 - Alert
2 - Drowsy, somnolent
1 - Obtunded, stupor
0 - Coma/unresponsive
Orientation (time, space, people) questions 3 - Answers each of all three questions correctly
2 - Answers two of three questions correctly
1 - Answers one of three questions correctly
0 - Answers neither correctly
Articulation 3 - Normal
2 - Mild dysarthria
1 - Severe dysarthria
0 - Mute or global aphasia
Gaze 3 - Normal horizontal movements
2 - Eyeball in medial position, able to deviate to either side
1 - Eyeball in lateral position, able to return to medial position
0 - Complete gaze palsy (conjugate deviation)
Facial movement 2 - Normal
1 - Partial facial weakness (paresis)
0 - Complete unilateral palsy (paralysis)
Visual fields 2 - Normal
1 - Partial hemanopia
0 - Complete hemanopia
Arm’s motoric function - Passive (unaffected side) 3 - No drift
Examiner raises both subject’s arms to a position of 45°
(if the subject is in a supine position), or 90° (if seated).
Subject is asked to hold such a position for 10 seconds
2 - Falls before10 seconds
1 - Unable to hold, but performs noticeable effort against gravity
0 - No effort against gravity
Arm’s motor function - Active (affected side) 3 - Able to raise perfectly
Examiner raises both subject’s arms to a position of 45°
(if the subject is in a supine position), or 90° (if seated).
Subject is asked to hold such a position for 10 seconds
2 - Able to raise
1 - Able to raise with flexed arm
0 - No movement
Extension of wrist 3 - Full extension, full strength
2 - Full extension, less strength
1 - Partial extension
0 - Unable to extend
Fingers strength 2 - Balanced strength of both hands
Subject is asked to perform pinch (squeeze between
the finger and thumb) with both hands, and the
examiner has to release the squeeze with one finger
1- Less strength on the affected hand
0 - Unable to perform pinching
Limb’s motoric function - Passive (unaffected side) 3 - No drift
Examiner raises both subject’s limbs to a position of 30°
(with the subject in a supine position). Subject is asked
to hold such a position for 5 seconds
2 - Falls before 5 seconds
1 - Unable to hold, but performs noticeable effort against gravity
0 - No effort against gravity
Limb’s motoric function - Active (affected side) 2 - Full flexion
Subject is asked to perform thigh and knee flexion 1 - Partial flexion
0 - No flexion
Dorsiflexion 2 - Normal strength to perform dorsiflexion
1 - Less strength to perform dorsiflexion
0 - Unable to perform dorsiflexion
Gait 4 - Walks 5 m long without aid or other person’s assistance
3 - Walks with aid (no assistance from other person)
2 - Walks with assistance from other person
1 - Unable to walk, but able to stand with aid
0 - Neither walks or stands
SSGM: Gadjah Mada Stroke Scale
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After 90 days of treatment, we found no difference in hemostasis
profile between groups as shown by PT, aPTT, and INR values (Table 3).
Within-group analysis showed that there was a bit of shortened PT
with DLBS1033, but of no clinical importance. The aPTT and INR in
each group did not change from their respective baseline values. All
measured hemostasis parameters in each group remained within their
respective normal ranges.
Hemostasis profile at the end of the study demonstrated that
administration of DLBS1033 at the dose of 3 × 490 mg daily for 90 days
in ischemic stroke subjects was safe and comparable to aspirin 80 mg
daily or clopidogrel 75 mg daily. There was no significant prolongation
of hemostasis parameters found. Neither were there bleeding adverse
events in any groups observed during the study conduct.
All subjects in all groups received simvastatin 20 mg daily and
citicholine 1000 mg twice daily during the study participation. For the
antihypertensive agents, subjects might receive amlodipine, valsartan
or a combination of both, depending on their individual condition. All
diabetic subjects received insulin therapy.
In terms of clinical outcome, each group demonstrated a significant
improvement of BI (p<0.001) from baseline to day 90, with a
mean score of >85 in all groups at the end of study. The greatest
improvement was observed in DLBS1033 Group, with the size of
improvement of 23.09±19.16 from baseline, but it was not significantly
different (p=0.098) with that of aspirin (15.12±15.71) or clopidogrel
(17.98±19.03), as shown in Fig. 1.
In line with the improvement of BI, DLBS1033 Group demonstrated
the greatest improvement of SSGM (6.98±4.90) from baseline that was
also significantly greater (p=0.002) than that of aspirin (3.74±3.66) or
Clopidogrel (4.26±4.21) (Fig. 2).
We also found there were similar percentages of subject in DLBS1033
Group (83.3% and 100%) to those in aspirin (92.9% and 100%) who
achieved BI of ≥85 and SSGM of ≥23, respectively, at the end of study;
both of which were slightly higher than those in clopidogrel (73.8% and
97.6%, respectively), even though they were not statistically different
(Fig. 3). Compared to aspirin, treatment with DLBS1033 for 90 days
seemed to be effectively comparable in achieving BI ≥85 (odds ratio
Table 2: Baseline characteristics
Variable Group 1 Aspirin (n=42) Group 2 Clopidogrel (n=42) Group 3 DLBS1033 (n=42) p‡
Age (year) 58.81±9.40 62.45±10.54 61.29±8.73 0.210
Gender (%)
Male 31 (73.8) 29 (69.0) 31 (73.%) 0.854
Female 11 (26.2) 13 (31.0) 11 (26.2)
Onset of stroke (hours before
initial dose of study medication)
25.26±32.85 23.24±26.75 33.52±47.41 0.399
Blood pressure (mmHg)
Systolic 166.90±35.97 162.74±26.09 168.45±29.39 0.570
Diastolic 98.69±18.45 93.93±11.77 95.60±13.58 0.356
Concomitant diseases (%)
Hypertension 22 (52.4) 15 (35.7) 21 (50.0) 0.253
Diabetes mellitus 18 (42.9) 14 (33.3) 19 (45.2) 0.501
Dyslipidemia 32 (76.2) 24 (57.1) 20 (47.6) 0.024*
Concomitant medication (%)
Amlodipine 15 (35.7) 10 (23.8) 13 (31.0) 0.489
Valsartan 11 (26.2) 6 (14.3) 11 (26.2) 0.317
Insulin 18 (42.9) 14 (33.3) 19 (45.2) 0.501
Clinical outcomes
Barthel index 81.43±23.07 73.57±28.14 71.55±27.42 0.262
SSGM 32.05±5.66 29.64±7.37 28.52±6.67 0.030*
Laboratoric parameters
FPG (mg/dl) 143.36±68.44 144.36±80.23 141.74±69.03 0.927
Hemoglobin (g/dl) 14.58±1.52 13.88±1.76 14.28±1.57 0.365
Total cholesterol (mg/dl) 226.21±43.84 217.69±48.02 214.28±45.61 0.222
HDL (mg/dl) 42.81±9.36 43.71±11.96 44.41±9.99 0.876
LDL (mg/dl) 160.08±36.74 140.79±35.63 142.24±32.03 0.044*
Triglyceride (mg/dl) 173.40±90.83 154.17±80.92 147.27±61.66 0.559
BUN (mg/dl) 13.10±3.93 12.55±3.18 12.80±3.07 0.908
Serum creatinin (mg/dl) 1.23±0.70 1.17±0.41 1.09±0.51 0.766
Uric acid (mg/dl) 6.48±1.42 6.20±1.72 6.50±1.50 0.748
PT (seconds) 12.98±1.22 13.07±1.15 13.09±1.29 0.866
aPTT (seconds) 29.66±1.22 29.39±3.39 29.65±4.73 0.978
INR 0.94±0.16 0.93±0.13 0.97±0.15 0.545
Continuous data are expressed in mean±SD. Categorical data are expressed in a number of subjects (n) and percentage (%). ‡Between-group analysis: Categorical
variables were analyzed using Chi-square; while the continuous ones using Kruskal–Wallis. aPTT: Activated partial thromboplastin time, BUN: Blood urea nitrogen,
FPG: Fasting plasma glucose, HDL: High-density lipoprotein cholesterol, INR: International normalized ratio, LDL: Low-density lipoprotein cholesterol, PT: Prothrombin
time, SSGM: Gadjah Mada Stroke Scale, SD: Standard deviation
Table 3: Hemostasis profile between groups at baseline and day 90
Hemostasis profile Baseline p‡Day 90 p‡p†
PT (second)
Aspirin (n=42) 12.98±1.22 0.866 12.36±1.39 0.788 0.080
Clopidogrel (n=42) 13.07±1.15 12.53±1.19 0.076
DLBS1033 (n=42) 13.09±1.29 12.67±1.55 0.039*
aPTT (second)
Aspirin (n=42) 29.66±5.49 0.978 29.59±4.25 0.619 0.523
Clopidogrel (n=42) 29.39±3.39 29.96±4.16 0.302
DLBS1033 (n=42) 29.65±4.73 30.54±4.41 0.096
INR
Aspirin (n=42) 0.94±0.16 0.545 0.94±0.13 0.154 0.318
Clopidogrel (n=42) 0.93±0.13 0.96±0.11 0.051
DLBS1033 (n=42) 0.97±0.15 0.99±0.13 0.132
Data are expressed in mean±SD. †Within-group analysis using Wilcoxon
Signed-Rank. ‡Between-group analysis using Kruskal–Wallis. *Statistically
significant (p<0.05). aPTT: Activated partial thromboplastin time,
INR: International normalized ratio, PT: Prothrombin time, SSGM: Gadjah
Mada Stroke Scale, SD: Standard deviation
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[OR]=0.38; 95% confidence interval [CI], 0.09-1.60; p=0.189). That
was also the case with DLBS1033 in comparison with clopidogrel
(OR= 1.77; 95% CI, 0.61-5.14; p=0.291). DLBS1033 treatment was
also effectively comparable in achieving SSGM ≥23, either compared
to aspirin (OR=1.00; 95% CI, 0.019-51.57; p=1.000) or clopidogrel
(OR=3.07; 95% CI, 0.12-77.60; p=0.496).
DISCUSSION
This study demonstrated that DLBS1033 therapy in ischemic stroke
patients provided a safe hemostatic profile, which was comparable
to that of aspirin or clopidogrel therapy (Table 3). All measured
parameters, i.e., PT, aPTT, and INR, were within the normal range at the
Fig. 1: Effect of aspirin, clopidogrel, and DLBS1033 treatment on the functional outcome measured by Barthel index. The error bars
represent the standard errors; Wilcoxon signed-rank was used in within-group analysis; Kruskal–Wallis was used in between-group
analysis
Fig. 2. Effect of aspirin, clopidogrel, and DLBS1033 treatment on the neurological outcome measured by Gadjah Mada Stroke Scale. The
error bars represent the standard errors; Wilcoxon signed-rank was used in within-group analysis; Kruskal–Wallis was used in between-
group analysis
Fig.3:ProportionofsubjectswithBathelindex≥85(left)andGadjahMadaStrokeScale≥23(right)
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end of the study, suggesting that there were no deficient coagulation
factors observed during therapy with DLBS1033, aspirin, and
clopidogrel. Furthermore, the result of this study also indicates that the
risk for bleeding due to the administration of DLBS1033 was evidently
low, similar to that of aspirin or clopidogrel at their respective usual
dose regimen. This was also clinically proven by zero bleeding event
in all groups observed during the study conduct. The safety profile of
DLBS1033 was aligned to that reported by Zhang in a former study with
a similar protease enzymes, lumbrokinase [20], demonstrating that the
proteases did not affect hemostasis profile measured by PT or aPTT,
thus did not interfere with the INR.
With respect to aspirin and clopidogrel treatment, our current
3-month-study showed no prolongation of aPTT. Neither did it show the
increase of INR nor any bleeding events. This was somewhat different
with a former report by Tamura et al, in which aspirin and clopidogrel
treatment were reported associated with bleeding complications
(hemorrhage, melaena, and hematochezia). Aspirin was also reported
associated with prolongation of aPTT, but not with increased INR.
While clopidogrel treatment was associated neither with prolongation
of aPTT nor increased INR [35]. However, in the report, aspirin might be
used concomitantly with warfarin and/or clopidgrel and for a long-term
therapy as well, while the observation in our study was limited only
to the first 3 months of therapy. That may explain the difference with
the former report. In this study, we found neither clinically significant
bleeding events nor adverse changes of the laboratory hemostatic
parameters. A case study also reported that clopidogrel treatment was
associated with prolonged aPTT and induced AHA in a patient with
cerebellar infarction, but the mechanism of how the drug could induce
the event is unknown [36], and thus, the adverse effect was likely to be
an anecdotal case.
In addition to its promisingly safe hemostatic profile, DLBS1033 also
improved the clinical outcomes. A significant improvement of BI from
baseline occurred in each treatment group. The improvement was
similar across all groups (Fig. 2). This result suggested that DLBS1033
was as efficacious as aspirin or clopidogrel in improving functional
outcome of patients with ischemic stroke. This study also demonstrated
that DLBS1033 treatment was effectively comparable to aspirin or
clopidogrel in achieving BI of ≥85 that means DLBS1033 treatment
effectively improved subjects’ daily performance from disability or
necessity for other people’s assistance to becoming independence.
A score of 85 of BI usually corresponded to independence with minimal
assistance that the majority of patients were able to get dressed and to
move from armchair to bed without assistance [37]. Score of <85 also
corresponded to a dependent state in which patients reported needing
assistance in performing daily living activities, with a sensitivity of 94-
95% and a specificity of 80-86% [38-40].
Further, even though that the efficacy of DLBS1033 treatment seemed
to be comparable to aspirin or clopidogrel in achieving SSGM of ≥23,
the size of improvement of the neurological function was significantly
greater with DLBS1033 than that with aspirin or clopidogrel (Fig. 2).
In this study, the improvement of both functional and neurological
outcome with DLBS1033 treatment was likely due to DLBS1033
activities as both the antithrombotic and thrombolytic agent [21,41].
In vitro studies demonstrated reduced expression of several genes
involved in inflammatory and atherogenic reaction by DLBS1033,
such as nuclear factor kappa B (NF-κB), tumor necrosis factor-alpha
(TNF-α), vascular cell adhesion molecule-1 (VCAM-1), and P-selectin.
The down-regulation of NF-κB has a link to the inhibition of atheroma
formation, suggesting that DLBS1033 works as an anti-atherogenesis
agent. DLBS1033 can inhibit the progression of other cytokines that
are activated by TNF-α and adherence of leukocytes to endothelium.
Activity of DLBS1033 in decreasing the expression of P-selectin may be
related to the suppression of de novo synthesis of P-selectin mediated by
cytokine. DLBS1033 suppressed the expression of VCAM-1, a member
of the immunoglobulin gene superfamily that mediates leukocyte
binding to the endothelial cell. In the same study, it was shown that
DLBS1033 suppressed the expression of MMP-9 gene, a marker of
plaque instability, suggesting that this bioactive protein fraction has
the ability to control plaque stabilization. MMP-9 is a protease that
degrades extracellular matrix proteins including gelatin, collagen,
elastin, and laminin that are important in tissue destruction; and
also in tissue remodeling and inflammation. This study indicated that
DLBS1033 could regulate the uncontrolled event of plaque rupture by
inhibiting the expression of MMP-9 [21,41]. Our current clinical findings
indicate that DLBS1033 is promising in accommodating the necessity
of treatment for acute ischemic diseases, due to its dual features Asan
antiplatelet and thrombolytic agent and the oral formulation offers
more practical administration.
Due to the limited scope of our study, we did not discuss to what extent
other variables such as age, NIHSS score at admission, cardio-/cerebro-
vascular history (such as myocardial infarction and stroke), dementia,
socio-economic status, presence of fever, undernutrition, as were also
reported in former studies [42-47], might influence the outcomes.
However, random allocation applied in this study allowed a comparable
distribution of those confounding factors between groups; therefore,
a valid and reliable interpretation could still be made as previously
discussed.
CONCLUSIONS
The study concluded that treatment with DLBS1033 at the dose
of 490 mg 3 times daily for 90 days in ischemic stroke subjects
demonstrated a safe hemostatic profile, which was comparable to that
of aspirin 80 mg once daily or clopidogrel 75 mg once daily. In terms
of functional and neurological outcomes, the study indicated that
DLBS1033 clinically benefitted the ischemic stroke subjects.
ACKNOWLEDGMENTS
The study was supported by Dexa Medica, Indonesia. The technical
assistance of Liana W. Susanto, MBiomed., in the statistical analysis of
the data as well as manuscript preparation is gratefully appreciated.
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