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Antiplatelet effects of aspirin with phytosterols: Comparison with non-enteric coated aspirin alone
Thrombosis Research 126 (2010) 384–385
Contents lists available at ScienceDirect
Thrombosis Research j o u r n a l h o m e p a g e : w w w. e l s ev i e r. c o m / l o c a t e / t h r o m r e s
Brief Communication
Antiplatelet effects of aspirin with phytosterols: Comparison with non-enteric coated aspirin alone Mark J. Antonino a, Rosa Coppolecchia b, Elisabeth Mahla a, Kevin P. Bliden a, Udaya S. Tantry a, Paul A. Gurbel a,⁎ a b
Sinai Center for Thrombosis Research, Baltimore Maryland Bayer HealthCare LLC, Morristown, NJ
a r t i c l e
i n f o
Article history: Received 12 February 2009 Received in revised form 24 April 2009 Accepted 1 May 2009 Available online 15 May 2009 Keywords: Aspirin Phytosterols Platelets Thromboxane Aggregation
a b s t r a c t The novel combination of aspirin and phytosterols may be a potential strategy to treat patients with cardiovascular disease. We sought to determine if the antiplatelet effects of a combination caplet of 81 mg aspirin with 400 mg phytosterols differed from the antiplatelet effects of non-enteric coated aspirin. The first five days of aspirin therapy alone (T1) produced marked reductions in collagen-induced, ADP-induced, and archidonic acid- induced platelet aggregation, and in serum and urine TxB2 compared to baseline. Five days after randomization to aspirin alone versus aspirin + phytosterols (T2), there were no differences in any measurement of platelet function within each group compared to T1 or between groups. The present study suggests that the antiplatelet effect of non-enteric coated 81 mg twice-daily aspirin therapy alone is not affected by the addition of phytosterols in a combination product. © 2009 Elsevier Ltd. All rights reserved.
Introduction Aspirin is one of the most widely used and effective antiplatelet agents for primary and secondary prevention of cardiovascular disease. As demonstrated by numerous studies and one meta-analysis aspirin treatment is associated with a 25% reduction in adverse cardiovascular events [1,2]. Aspirin irreversibly inhibits platelet cyclooxygenase thus attenuating the formation of thromboxane (Tx)A2 and TxA2-induced platelet aggregation [1]. Phytosterols are naturally occurring compounds found in plants that are chemically related to cholesterol. Phytosterols reduce biliary cholesterol absorption by displacing cholesterol from micelles resulting in decreased intestinal cholesterol solubility. Clinical trials have demonstrated that dietary intake of phytosterols is effective in lowering total and low-density lipoprotein (LDL) cholesterol with a low incidence of side effects. Phytosterol therapy has also been investigated in patients with hypercholesterolemia [3]. Intestinal absorption may play an important role in the efficacy of aspirin. In a recent study, incomplete suppression of TxB2 generation was found to occur in 8% of the aspirin group and 54.3% of the enteric-coated aspirin group [4]. Since the novel combination of aspirin and phytosterols may be a potential strategy to treat patients with cardiovascular disease, it is important to exclude any effect on cyclooxygenase inhibition. The current study was conducted to determine if the antiplatelet effects of “Aspirin with Heart Advantage” (Bayer HealthCare LLC, Morristown, NJ), a combination caplet of 81 mg aspirin with 400 mg ⁎ Corresponding author. Sinai Center for Thrombosis Research, Sinai Hospital, 2401 W. Belvedere Ave, Baltimore, MD 21215. Tel.: +1 410 601 9600; fax: +1 410 601 9601. E-mail address: [email protected] (P.A. Gurbel). 0049-3848/$ – see front matter © 2009 Elsevier Ltd. All rights reserved. doi:10.1016/j.thromres.2009.05.003
phytosterols, differed from the antiplatelet effects of non-enteric coated (chewable) aspirin tablet (Bayer HealthCare LLC, Morristown, NJ). Methods This study was a randomized, controlled, open label, parallel group, single center study. Thirty healthy male and female volunteers, 18-55 years of age with a body mass index (BMI) of 18 - 30 kg/m2, were enrolled. Subjects underwent a screening visit 7-14 days before enrollment. Exclusion criteria were aspirin allergy, pregnancy or lactation, use of aspirin or aspirin containing products or NSAIDS for two weeks prior to and during the treatment period, history of gastrointestinal ulcers, bleeding, perforation or melena, clinical evidence of any significant cardiovascular, renal, hepatic, gastrointestinal, neurological, endocrine, metabolic, hematologic, pulmonary, or psychiatric disease as determined by medical history, previous or current history of smoking or tobacco use, positive breath alcohol level or positive urine for illicit drugs, and history of surgery within 30 days. There were 2 treatment periods. Treatment Period 1 (T1) consisted of 5 days (day 1 to 5) when subjects were administered 81 mg aspirin twice daily and Treatment Period 2 (T2) consisted of day 6 to 10. On the first day of T2 (day 6), subjects were administered in a random fashion either 81 mg twice daily aspirin tablet (Group 1) or twice daily aspirin 81 mg plus phytosterols (400 mg) caplet (Group 2). Blood Sampling Blood was collected from the antecubital vein into Vacutainer tubes containing 3.2% final concentration citrate (Becton-Dickinson,
M.J. Antonino et al. / Thrombosis Research 126 (2010) 384–385
385
Table 1 Antiplatelet Effects of Aspirin and Aspirin Plus Phytosterols in Healthy Volunteers. Baseline
T1
p-value Baseline versus T1
T2
p-value T1 versus T2
(Group 1, n = 15) 2mmM AA- aggregation (%) 4ug/ml Collagen- aggregation (%) 5uM ADP- aggregation (%) Serum TxB2 (pg/ml) Urinary Thromboxane (pg 11-dh-TxB2/mg creatinine)
83 ± 6 87 ± 7 80 ± 18 5320 ± 6207 3301 ± 1601
3±1 14 ± 15 63 ± 17 33 + 35 542 ± 299
p b 0.01 p b 0.01 p b 0.01 p b 0.01 p b 0.01
3±1 17 ± 23 68 ± 9 40+48 498 ± 148
NS NS NS NS NS
(Group 2, n = 15) 2mmM AA- aggregation (%) 4ug/ml Collagen- aggregation (%) 5uM ADP- aggregation (%) Serum TxB2 (pg/ml) Urinary Thromboxane (pg 11-dh-TxB2/mg creatinine)
86 ± 5 90 ± 6 77 ± 16 4425 ± 3183 3085 ± 1307
4±1 20 ± 24 70 ± 16 18 ± 17 546 ± 242
p b 0.01 p b 0.01 p b 0.01 p b 0.01 p b 0.01
4±1 17 ± 24 62 ± 19 17 ± 19 512 ± 209
NS NS NS NS NS
Franklin Lakes, NJ) between 7-9AM at baseline on day 1 before initial drug administration, on day 6 before randomization and day 11. After the first 2 to 3 mL of free-flowing blood was discarded, the tubes were filled to capacity and gently inverted 3 to 5 times to ensure complete mixture of the anticoagulant.
ADP-induced, and archidonic acid- induced platelet aggregation, and in serum and urine TxB2 compared to baseline. Five days after randomization to aspirin alone versus aspirin+ phytosterols (end of T2) there were no differences in any measurement of platelet function within each group compared to T1 or between groups.
Light Transmittance Aggregometry
Discussion
Two and 4 μg/mL collagen-induced, 5 μM adenosine diphosphate (ADP)-induced, 2 mM arachidonic acid-induced aggregation were performed as described previously [5].
The present study suggests that the antiplatelet effect of nonenteric coated 81 mg twice-daily aspirin therapy alone is not affected by the addition of phytosterols in a combination product. Aspirin therapy in both preparations markedly inhibited aggregation and TxB2 production, consistent with the results of previous investigations [2,5]. The influence on platelet function of the new aspirin preparation with phytosterols would most likely have been observed within 5 days since approximately 10% of new platelets are generated each day [6]. The results of our study may be useful in addressing the antiplatelet effects of co-administered phytosterols and aspirin.
Serum Thromboxane B2 Serum TxB2 levels were assayed by the Serum Thrombaxane B2 EIA Kit (Cayman Chemicals Inc., Ann Arbor, Michigan). Unanticoagulated samples were allowed to clot for 30 minutes at 37°C, and then centrifuged for 15 minutes at 2000 g. Indomethacin (final concentration 10 µM, Sigma Chemicals, St. Louis, Missouri) was then added and the serum samples were stored at -70 °C until analysis. Urinary 11-dehydro- Thromboxane B2 Urinary 11-dehydro (dh)-TxB2 was determined by the AspirinWorks® (Corgenix; Broomfield, Colorado) enzyme-linked immunosorbent assay (ELISA). Briefly, 100 μl of urine in assay buffer was incubated with a monoclonal antibody followed by the addition of 11-dh-Tx B2alkaline phosphate tracer. Urinary 11-dh-TxB2 concentrations were determined by measuring color development at 405 nm using an ELISA reader and expressed as pg/mg creatinine.
Conflicts of Interest Statement Dr. Paul A Gurbel has received research grants and honoraria from Schering Plough, Haemoscope, Astra Zeneca, Medtronic, Lilly/Daiichi Sankyo, Sanofi-Aventis, Boston-Scientific and Bayer Healthcare, Portola Pharmaceuticals, and Pozen. Dr. Rosa Coppolecchia is an employee of Bayer HealthCare LLC, Morristown, NJ. and contributed to the study design. Dr. Coppolecchia had no role in the writing of this brief communication and in the collection, analysis or interpretation of the data. The remaining authors report no conflicts. Source of Funding
Statistical Analysis Responsiveness to aspirin is defined as a N95% inhibition of serum thromboxane. Non inferiority between the two treatment strategies was considered if there was a ≤5% difference in platelet inhibition between the groups. Based on a power of 0.9 and an alpha value of 0.05 a sample size of 15 in each group was calculated. Data were analyzed by one-way analyis of variance with posthoc Dunn's test correcting for multiple comparisions. P b 0.05 was considered significant (SigmaStat, Systat software, Inc., San Jose, CA). Results Results are shown in the Table 1. There was no difference in demographic variables, hematologic data, renal function, baseline platelet aggregation or baseline TxB2 (serum and urine) levels between treatment groups. The first five days of aspirin therapy alone (end of T1) produced marked reductions in collagen-induced,
Bayer HealthCare LLC, Morristown, NJ. References [1] Collaborative meta-analysis of randomised trials of antiplatelet therapy for prevention of death, myocardial infarction, and stroke in high risk patients. BMJ 2002;324:71–86. [2] Patrono C, Baigent C, Hirsh J, Roth G. Antiplatelet drugs: American College of Chest Physicians Evidence-Based Clinical Practice Guidelines (8th Edition). Chest 2008;133:199S–233S. [3] Van Horn L, McCoin M, Kris-Etherton PM, Burke F, Carson JA, Champagne CM, et al. The evidence for dietary prevention and treatment of cardiovascular disease. J Am Diet Assoc 2008;108:287–331. [4] Cox D, Maree AO, Dooley M, Conroy R, Byrne MF, Fitzgerald DJ. Effect of enteric coating on antiplatelet activity of low-dose aspirin in healthy volunteers. Stroke 2006;37:2153–8. [5] Gurbel PA, Bliden KP, DiChiara J, Newcomer J, Weng W, Neerchal NK, et al. Evaluation of dose-related effects of aspirin on platelet function: results from the Aspirin-Induced Platelet Effect (ASPECT) study. Circulation 2007;115:3156–64. [6] George JN. Platelets. Lancet 2000;355:1531–9.
Contents lists available at ScienceDirect
Thrombosis Research j o u r n a l h o m e p a g e : w w w. e l s ev i e r. c o m / l o c a t e / t h r o m r e s
Brief Communication
Antiplatelet effects of aspirin with phytosterols: Comparison with non-enteric coated aspirin alone Mark J. Antonino a, Rosa Coppolecchia b, Elisabeth Mahla a, Kevin P. Bliden a, Udaya S. Tantry a, Paul A. Gurbel a,⁎ a b
Sinai Center for Thrombosis Research, Baltimore Maryland Bayer HealthCare LLC, Morristown, NJ
a r t i c l e
i n f o
Article history: Received 12 February 2009 Received in revised form 24 April 2009 Accepted 1 May 2009 Available online 15 May 2009 Keywords: Aspirin Phytosterols Platelets Thromboxane Aggregation
a b s t r a c t The novel combination of aspirin and phytosterols may be a potential strategy to treat patients with cardiovascular disease. We sought to determine if the antiplatelet effects of a combination caplet of 81 mg aspirin with 400 mg phytosterols differed from the antiplatelet effects of non-enteric coated aspirin. The first five days of aspirin therapy alone (T1) produced marked reductions in collagen-induced, ADP-induced, and archidonic acid- induced platelet aggregation, and in serum and urine TxB2 compared to baseline. Five days after randomization to aspirin alone versus aspirin + phytosterols (T2), there were no differences in any measurement of platelet function within each group compared to T1 or between groups. The present study suggests that the antiplatelet effect of non-enteric coated 81 mg twice-daily aspirin therapy alone is not affected by the addition of phytosterols in a combination product. © 2009 Elsevier Ltd. All rights reserved.
Introduction Aspirin is one of the most widely used and effective antiplatelet agents for primary and secondary prevention of cardiovascular disease. As demonstrated by numerous studies and one meta-analysis aspirin treatment is associated with a 25% reduction in adverse cardiovascular events [1,2]. Aspirin irreversibly inhibits platelet cyclooxygenase thus attenuating the formation of thromboxane (Tx)A2 and TxA2-induced platelet aggregation [1]. Phytosterols are naturally occurring compounds found in plants that are chemically related to cholesterol. Phytosterols reduce biliary cholesterol absorption by displacing cholesterol from micelles resulting in decreased intestinal cholesterol solubility. Clinical trials have demonstrated that dietary intake of phytosterols is effective in lowering total and low-density lipoprotein (LDL) cholesterol with a low incidence of side effects. Phytosterol therapy has also been investigated in patients with hypercholesterolemia [3]. Intestinal absorption may play an important role in the efficacy of aspirin. In a recent study, incomplete suppression of TxB2 generation was found to occur in 8% of the aspirin group and 54.3% of the enteric-coated aspirin group [4]. Since the novel combination of aspirin and phytosterols may be a potential strategy to treat patients with cardiovascular disease, it is important to exclude any effect on cyclooxygenase inhibition. The current study was conducted to determine if the antiplatelet effects of “Aspirin with Heart Advantage” (Bayer HealthCare LLC, Morristown, NJ), a combination caplet of 81 mg aspirin with 400 mg ⁎ Corresponding author. Sinai Center for Thrombosis Research, Sinai Hospital, 2401 W. Belvedere Ave, Baltimore, MD 21215. Tel.: +1 410 601 9600; fax: +1 410 601 9601. E-mail address: [email protected] (P.A. Gurbel). 0049-3848/$ – see front matter © 2009 Elsevier Ltd. All rights reserved. doi:10.1016/j.thromres.2009.05.003
phytosterols, differed from the antiplatelet effects of non-enteric coated (chewable) aspirin tablet (Bayer HealthCare LLC, Morristown, NJ). Methods This study was a randomized, controlled, open label, parallel group, single center study. Thirty healthy male and female volunteers, 18-55 years of age with a body mass index (BMI) of 18 - 30 kg/m2, were enrolled. Subjects underwent a screening visit 7-14 days before enrollment. Exclusion criteria were aspirin allergy, pregnancy or lactation, use of aspirin or aspirin containing products or NSAIDS for two weeks prior to and during the treatment period, history of gastrointestinal ulcers, bleeding, perforation or melena, clinical evidence of any significant cardiovascular, renal, hepatic, gastrointestinal, neurological, endocrine, metabolic, hematologic, pulmonary, or psychiatric disease as determined by medical history, previous or current history of smoking or tobacco use, positive breath alcohol level or positive urine for illicit drugs, and history of surgery within 30 days. There were 2 treatment periods. Treatment Period 1 (T1) consisted of 5 days (day 1 to 5) when subjects were administered 81 mg aspirin twice daily and Treatment Period 2 (T2) consisted of day 6 to 10. On the first day of T2 (day 6), subjects were administered in a random fashion either 81 mg twice daily aspirin tablet (Group 1) or twice daily aspirin 81 mg plus phytosterols (400 mg) caplet (Group 2). Blood Sampling Blood was collected from the antecubital vein into Vacutainer tubes containing 3.2% final concentration citrate (Becton-Dickinson,
M.J. Antonino et al. / Thrombosis Research 126 (2010) 384–385
385
Table 1 Antiplatelet Effects of Aspirin and Aspirin Plus Phytosterols in Healthy Volunteers. Baseline
T1
p-value Baseline versus T1
T2
p-value T1 versus T2
(Group 1, n = 15) 2mmM AA- aggregation (%) 4ug/ml Collagen- aggregation (%) 5uM ADP- aggregation (%) Serum TxB2 (pg/ml) Urinary Thromboxane (pg 11-dh-TxB2/mg creatinine)
83 ± 6 87 ± 7 80 ± 18 5320 ± 6207 3301 ± 1601
3±1 14 ± 15 63 ± 17 33 + 35 542 ± 299
p b 0.01 p b 0.01 p b 0.01 p b 0.01 p b 0.01
3±1 17 ± 23 68 ± 9 40+48 498 ± 148
NS NS NS NS NS
(Group 2, n = 15) 2mmM AA- aggregation (%) 4ug/ml Collagen- aggregation (%) 5uM ADP- aggregation (%) Serum TxB2 (pg/ml) Urinary Thromboxane (pg 11-dh-TxB2/mg creatinine)
86 ± 5 90 ± 6 77 ± 16 4425 ± 3183 3085 ± 1307
4±1 20 ± 24 70 ± 16 18 ± 17 546 ± 242
p b 0.01 p b 0.01 p b 0.01 p b 0.01 p b 0.01
4±1 17 ± 24 62 ± 19 17 ± 19 512 ± 209
NS NS NS NS NS
Franklin Lakes, NJ) between 7-9AM at baseline on day 1 before initial drug administration, on day 6 before randomization and day 11. After the first 2 to 3 mL of free-flowing blood was discarded, the tubes were filled to capacity and gently inverted 3 to 5 times to ensure complete mixture of the anticoagulant.
ADP-induced, and archidonic acid- induced platelet aggregation, and in serum and urine TxB2 compared to baseline. Five days after randomization to aspirin alone versus aspirin+ phytosterols (end of T2) there were no differences in any measurement of platelet function within each group compared to T1 or between groups.
Light Transmittance Aggregometry
Discussion
Two and 4 μg/mL collagen-induced, 5 μM adenosine diphosphate (ADP)-induced, 2 mM arachidonic acid-induced aggregation were performed as described previously [5].
The present study suggests that the antiplatelet effect of nonenteric coated 81 mg twice-daily aspirin therapy alone is not affected by the addition of phytosterols in a combination product. Aspirin therapy in both preparations markedly inhibited aggregation and TxB2 production, consistent with the results of previous investigations [2,5]. The influence on platelet function of the new aspirin preparation with phytosterols would most likely have been observed within 5 days since approximately 10% of new platelets are generated each day [6]. The results of our study may be useful in addressing the antiplatelet effects of co-administered phytosterols and aspirin.
Serum Thromboxane B2 Serum TxB2 levels were assayed by the Serum Thrombaxane B2 EIA Kit (Cayman Chemicals Inc., Ann Arbor, Michigan). Unanticoagulated samples were allowed to clot for 30 minutes at 37°C, and then centrifuged for 15 minutes at 2000 g. Indomethacin (final concentration 10 µM, Sigma Chemicals, St. Louis, Missouri) was then added and the serum samples were stored at -70 °C until analysis. Urinary 11-dehydro- Thromboxane B2 Urinary 11-dehydro (dh)-TxB2 was determined by the AspirinWorks® (Corgenix; Broomfield, Colorado) enzyme-linked immunosorbent assay (ELISA). Briefly, 100 μl of urine in assay buffer was incubated with a monoclonal antibody followed by the addition of 11-dh-Tx B2alkaline phosphate tracer. Urinary 11-dh-TxB2 concentrations were determined by measuring color development at 405 nm using an ELISA reader and expressed as pg/mg creatinine.
Conflicts of Interest Statement Dr. Paul A Gurbel has received research grants and honoraria from Schering Plough, Haemoscope, Astra Zeneca, Medtronic, Lilly/Daiichi Sankyo, Sanofi-Aventis, Boston-Scientific and Bayer Healthcare, Portola Pharmaceuticals, and Pozen. Dr. Rosa Coppolecchia is an employee of Bayer HealthCare LLC, Morristown, NJ. and contributed to the study design. Dr. Coppolecchia had no role in the writing of this brief communication and in the collection, analysis or interpretation of the data. The remaining authors report no conflicts. Source of Funding
Statistical Analysis Responsiveness to aspirin is defined as a N95% inhibition of serum thromboxane. Non inferiority between the two treatment strategies was considered if there was a ≤5% difference in platelet inhibition between the groups. Based on a power of 0.9 and an alpha value of 0.05 a sample size of 15 in each group was calculated. Data were analyzed by one-way analyis of variance with posthoc Dunn's test correcting for multiple comparisions. P b 0.05 was considered significant (SigmaStat, Systat software, Inc., San Jose, CA). Results Results are shown in the Table 1. There was no difference in demographic variables, hematologic data, renal function, baseline platelet aggregation or baseline TxB2 (serum and urine) levels between treatment groups. The first five days of aspirin therapy alone (end of T1) produced marked reductions in collagen-induced,
Bayer HealthCare LLC, Morristown, NJ. References [1] Collaborative meta-analysis of randomised trials of antiplatelet therapy for prevention of death, myocardial infarction, and stroke in high risk patients. BMJ 2002;324:71–86. [2] Patrono C, Baigent C, Hirsh J, Roth G. Antiplatelet drugs: American College of Chest Physicians Evidence-Based Clinical Practice Guidelines (8th Edition). Chest 2008;133:199S–233S. [3] Van Horn L, McCoin M, Kris-Etherton PM, Burke F, Carson JA, Champagne CM, et al. The evidence for dietary prevention and treatment of cardiovascular disease. J Am Diet Assoc 2008;108:287–331. [4] Cox D, Maree AO, Dooley M, Conroy R, Byrne MF, Fitzgerald DJ. Effect of enteric coating on antiplatelet activity of low-dose aspirin in healthy volunteers. Stroke 2006;37:2153–8. [5] Gurbel PA, Bliden KP, DiChiara J, Newcomer J, Weng W, Neerchal NK, et al. Evaluation of dose-related effects of aspirin on platelet function: results from the Aspirin-Induced Platelet Effect (ASPECT) study. Circulation 2007;115:3156–64. [6] George JN. Platelets. Lancet 2000;355:1531–9.