Petra Jilma-Stohlawetz

Pharmacokinetics of anthocyanins and antioxidant effects after the consumption of anthocyanin-rich acai juice and pulp (Euterpe oleracea Mart.) in human healthy volunteers.

The acai berry is the fruit of the acai palm and is traditionally consumed in Brazil but has gained popularity abroad as a food and functional ingredient, yet little information exists on its health effect in humans. This study was performed as an acute four-way crossover clinical trial with acai pulp and clarified acai juice compared to applesauce and a non-antioxidant beverage as controls. Healthy volunteers (12) were dosed at 7 mL/kg of body weight after a washout phase and overnight fast, and plasma was repeatedly sampled over 12 h and urine over 24 h after consumption. Noncompartmental pharmacokinetic analysis of total anthocyanins quantified as cyanidin-3-O-glucoside showed Cmax values of 2321 and 1138 ng/L at t max times of 2.2 and 2.0 h, and AUC last values of 8568 and 3314 ng h L(-1) for pulp and juice, respectively. Nonlinear mixed effect modeling identified dose volume as a significant predictor of relative oral bioavailability in a negative nonlinear relationship for acai pulp and juice. Plasma antioxidant capacity was significantly increased by the acai pulp and applesauce. Individual increases in plasma antioxidant capacity of up to 2.3- and 3-fold for acai juice and pulp, respectively were observed. The antioxidant capacity in urine, generation of reactive oxygen species, and uric acid concentrations in plasma were not significantly altered by the treatments. Results demonstrate the absorption and antioxidant effects of anthocyanins in acai in plasma in an acute human consumption trial.

Morphine Decreases Clopidogrel Concentrations and Effects A Randomized, Double-Blind, Placebo-Controlled Trial

OBJECTIVES This study sought to examine the possible drug-drug interactions between clopidogrel and morphine. BACKGROUND Because morphine-the recommended treatment for pain of myocardial infarction-is associated with poor clinical outcome, we hypothesized that morphine lowers the plasma levels of clopidogrel active metabolite as well as its effects on platelets. METHODS Twenty-four healthy subjects received a loading dose of 600 mg clopidogrel together with placebo or 5 mg morphine intravenously in a randomized, double-blind, placebo-controlled, cross-over trial. Pharmacokinetics was determined by liquid chromatography tandem mass spectrometry, and clopidogrel effects were measured by platelet function tests. RESULTS Morphine injection delayed clopidogrel absorption (p = 0.025) and reduced the area under the curve levels of its active metabolite by 34% (p = 0.001). Morphine delayed the maximal inhibition of platelet aggregation on average by 2 h (n = 24; p < 0.001). Residual platelet aggregation was higher 1 to 4 h after morphine injection (n = 24; p < 0.005). Furthermore, morphine delayed the inhibition of platelet plug formation under high shear rates (P2Y-Innovance; n = 21; p < 0.004) and abolished the 3-fold prolongation in collagen adenosine diphosphate-induced closure times seen in extensive and rapid metabolizers (n = 16; p = 0.001). CONCLUSIONS Morphine delays clopidogrel absorption, decreases plasma levels of clopidogrel active metabolite, and retards and diminishes its effects, which can lead to treatment failure in susceptible individuals. (Drug/Drug Interactions of Aspirin and P2Y12-inhibitors; NCT01369186).

Clinical ResearchAntithrombotic TherapyMorphine Decreases Clopidogrel Concentrations and Effects: A Randomized, Double-Blind, Placebo-Controlled Trial

OBJECTIVES This study sought to examine the possible drug-drug interactions between clopidogrel and morphine. BACKGROUND Because morphine-the recommended treatment for pain of myocardial infarction-is associated with poor clinical outcome, we hypothesized that morphine lowers the plasma levels of clopidogrel active metabolite as well as its effects on platelets. METHODS Twenty-four healthy subjects received a loading dose of 600 mg clopidogrel together with placebo or 5 mg morphine intravenously in a randomized, double-blind, placebo-controlled, cross-over trial. Pharmacokinetics was determined by liquid chromatography tandem mass spectrometry, and clopidogrel effects were measured by platelet function tests. RESULTS Morphine injection delayed clopidogrel absorption (p = 0.025) and reduced the area under the curve levels of its active metabolite by 34% (p = 0.001). Morphine delayed the maximal inhibition of platelet aggregation on average by 2 h (n = 24; p < 0.001). Residual platelet aggregation was higher 1 to 4 h after morphine injection (n = 24; p < 0.005). Furthermore, morphine delayed the inhibition of platelet plug formation under high shear rates (P2Y-Innovance; n = 21; p < 0.004) and abolished the 3-fold prolongation in collagen adenosine diphosphate-induced closure times seen in extensive and rapid metabolizers (n = 16; p = 0.001). CONCLUSIONS Morphine delays clopidogrel absorption, decreases plasma levels of clopidogrel active metabolite, and retards and diminishes its effects, which can lead to treatment failure in susceptible individuals. (Drug/Drug Interactions of Aspirin and P2Y12-inhibitors; NCT01369186).

Inhibition of von Willebrand factor by ARC1779 in patients with acute thrombotic thrombocytopenic purpura

Thrombotic thrombocytopenic purpura (TTP) can cause severe organ damage due to enhanced platelet aggregation by ultra-large von Willebrand factor (VWF) multimers. Thus inhibition of VWF by the anti-VWF ARC1779 might potentially be beneficial for TTP patients. This prospective trial tested the safety, pharmacokinetics and pharmacodynamics of the anti-VWF aptamer ARC1779 added to plasma exchange therapy (PEX) in patients with acute TTP. Seven patients received bolus primed continuous i.v. infusions of ARC1779 (1-2 μg/kg/min) in addition to PEX until remission of TTP was induced or for 14 days. Mean steady state ARC1779 plasma concentrations of 9.9 μg/ml reduced VWF activity to 5% (mean baseline activity was 125% in TTP patients compared to a reference plasma). PEX reduced ARC1779 levels by 50%, but steady state concentrations were restored rapidly with a mini-bolus. After discontinuation of PEX, ARC1779 alone further increased platelet counts in one patient. Stopping ARC1779 was associated with an immediate drop of platelet counts in this patient. This suggests that ARC1779 can block the progression of TTP in patients with severe ADAMTS13 is deficiency. ARC1779 was generally well tolerated without any signs of bleeding. Pharmacokinetics and pharmacodynamics of ARC1779 were well predictable and in agreement with those observed in a previous trial with healthy volunteers. Based on its mechanism of action and the observed effect on platelet counts, ARC1779 used as an adjunctive to PEX may help accelerate recovery from organ dysfunction.

Duffy antigen modifies the chemokine response in human endotoxemia

Objective:The Duffy receptor is a promiscuous receptor for chemokines and selectively binds CXC and CC chemokines with high affinity. Preclinical data show that presence of the Duffy receptor on red blood cells may influence plasma levels of proinflammatory cytokines and chemokines and be protective during inflammation. This trial was designed to investigate the influence of the Duffy antigen on human inflammation in vivo. Design:Prospective, analyst-blinded clinical trial. Patients:A total of 32 healthy male volunteers: 16 Duffy-positive white subjects and 16 Duffy-negative subjects of African descent. Measurements and Main Results:All subjects received an intravenous bolus of 2 ng/kg endotoxin (lipopolysaccharide). Cytokines, chemokines, and their receptors were quantified by enzyme immunoassay, reverse transcriptase–polymerase chain reaction, and flow cytometry. Results:Plasma levels of tumor necrosis factor, interleukin-6, interleukin-10, and whole blood growth-related oncogen-α, monocyte chemoattractant protein-1, and interleukin-8 messenger RNA increased similarly in both groups after lipopolysaccharide infusion. Monocyte chemoattractant protein-1 peak plasma levels were roughly two-fold higher in Duffy-positive subjects compared with Duffy-negative subjects (16 ng/mL vs. 7 ng/mL, p < .0001). Similarly, growth-related oncogen-α levels were 2.5-fold higher in Duffy-positive subjects 2 hrs after lipopolysaccharide infusion (210 pg/mL vs. 85 pg/mL; p < .001). Erythrocyte-bound monocyte chemoattractant protein-1, growth-related oncogen-α, and interleukin-8 increased 20- to 50-fold in Duffy-positive subjects (p < .00001 vs. baseline). Conclusion:The Duffy antigen substantially alters chemokine concentrations in blood, but it does not have a protective effect during human endotoxemia.

A dose ranging phase I/II trial of the von Willebrand factor inhibiting aptamer ARC1779 in patients with congenital thrombotic thrombocytopenic purpura

Congenital thrombotic thrombocytopenic purpura (TTP) is a very rare but potentially life-threatening disorder. This phase I/II trial compared the pharmacokinetics and pharmacodynamics and safety of three different administration modes of the anti-von Willebrand factor (VWF) aptamer ARC1779. This was a prospective clinical trial with a partial cross-over design: three periods comprised subcutaneous injections of 50 mg of ARC1779 on seven subsequent days, a low-dose infusion of ARC1779 (0.002 mg/kg/min) for 24-72 hours and a high-dose infusion (0.004-0.006 mg/kg/min) up to 72 hours. ARC1779 concentrations were determined with high performance liquid chromatography, VWF inhibition was measured with enzyme immunoassay and platelet function was determined with the platelet function analyser (PFA-100) and impedance aggregometry. ARC1779 was well tolerated without any bleeding at concentrations spanning over three orders of magnitude. The daily s.c. injection yielded plasma levels (0.5 μg/ml) of the drug that were too low to sufficiently suppress VWF. The low-dose i.v. infusion increased platelet counts in one patient, whereas the high i.v. dose increased plasma concentrations up to 69 μg/ml, completely blocked free A1 domains, VWF-dependent platelet plug formation and enhanced platelet counts in 2/3 patients. In conclusion, infusion of ARC1779 dose-dependently inhibits VWF-dependent platelet function and during infusion ARC1779 increases or stabilises platelet counts in congenital TTP. However, the tested doses, particularly the daily s.c. injections, did not correct all clinical or laboratory features of TTP.

A randomised pilot trial of the anti-von Willebrand factor aptamer ARC1779 in patients with type 2b von Willebrand disease

Desmopressin aggravates thrombocytopenia in type 2B von Willebrand disease (VWF type 2B) by release of large and hyper-adhesive von Willebrand Factor (VWF) multimers. This pilot study investigated whether the anti-VWF aptamer ARC1779 can prevent desmopressin-induced thrombocytopenia and interferes with the excessive VWF turnover in patients with VWF type 2B. Concentration effect curves of ARC1779 were established for five patients in vitro and two patients with VWF type 2B were treated by infusion of ARC1779, desmopressin, or their combination in a randomised, controlled, double-blind design. ARC1779 concentrations in the range of 1-3 microg/ml blocked free A1 domain binding sites by 90% in vitro. In vivo, desmopressin alone induced a profound (-90%) drop in platelet counts in one of the patients. ARC1779 (4-5 microg/ml) completely inhibited VWF A1 domains and prevented this desmopressin-induced platelet drop. Desmopressin alone increased VWF antigen two- to three-fold, accompanied by concordant changes in VWF Ristocetin cofactor activity (RCo) and coagulation factor VIII activity. ARC1779 substantially enhanced the desmopressin-induced maximal increase in these parameters, and improved multimer patterns. No treatment related adverse events were observed and no bleeding occurred despite marked thrombocytopenia. These data provide first proof of concept in humans and evidence that ARC1779 is a potent inhibitor of VWF. ARC1779 prevented the rapid consumption of VWF multimers together with agglutinated platelets that occurred in response to desmopressin challenge in patients with VWD type 2B.

Anti–von Willebrand factor aptamer ARC1779 for refractory thrombotic thrombocytopenic purpura

BACKGROUND: Plasma exchange is the main therapy for thrombotic thrombocytopenic purpura (TTP). No treatments other than plasma exchange have been documented to be effective nor are approved for treatment of TTP. The anti–von Willebrand factor (VWF) aptamer ARC1779 effectively inhibits VWF activity in plasma samples of TTP patients and thus shear‐dependent platelet (PLT) function as measured by the PLT function analyzer PFA‐100 (Dade Behring). It was hypothesized that ARC1779 would offer a potentially effective treatment option for a critically ill patient, refractory to standard care.

Changes in thrombopoiesis and platelet reactivity in extremely low birth weight infants undergoing erythropoietin therapy for treatment of anaemia of prematurity.

Erythropoietin (Epo) is frequently administered to premature infants to stimulate erythropoiesis. There is evidence from studies in animals and healthy adults that Epo also interacts with thrombopoiesis and platelet function. This study investigates the effect of Epo therapy on platelet reactivity, peripheral platelet counts and thiazole orange-positive (TO+) platelets in extremely low birth weight (ELBW) infants. In a randomised-controlled trial, ELBW infants with a birth weight < or =800 g and a gestational age < or =32 weeks were either randomised to a group receiving Epo during the first weeks of life or to a control group. Our results show that thrombin receptor-activating peptide (TRAP-6) -induced expression of P-selectin increased significantly during the first two weeks of Epo treatment. With the exception of week five, the number of TO+ platelets was significantly higher during the first eight weeks in Epo-treated infants compared to controls. The increase of TO+ platelets was not paralleled by an increase in total platelet count. We can conclude that Epo therapy has a short-lasting effect on platelet reactivity toTRAP-6 in ELBW infants during the first two weeks of life. Furthermore, Epo therapy is associated with an increase in the number of TO+ platelets compared to controls.

The anti-von Willebrand factor aptamer ARC1779 increases von Willebrand factor levels and platelet counts in patients with type 2B von Willebrand disease

Blockade of hyperactive von Willebrand factor (VWF) by ARC1779 blunted the platelet drop induced by desmopressin in patients with type 2B von Willebrand disease (VWD). Thus, we hypothesised that ARC1779 may increase VWF levels and correct thrombocytopenia. Three thrombocytopenic patients suffering from type 2B VWD received a loading dose of 0.23 mg/kg ARC1779 followed by 4 μg/kg/min intravenously for 72 hours in a prospective clinical trial. ARC1779 was well tolerated and safe. Plasma concentrations of ARC1779 increased to 76 μg/ml (59-130) leading to an immediate decrease of free VWF A1 domains. VWF/FVIII levels increased as early as 12 h after start of infusion, peaked near the end of infusion, and returned to baseline at follow-up. VWF ristocetin cofactor activity (VWF:RCo) showed a median 10-fold increase 8 hours after end of infusion, while the median VWF-antigen and FVIII increase was less (5-fold and 4-fold, respectively). Most importantly inhibition of hyperactive VWF rapidly increased platelet counts from 40 x 10(9)/l (38-58 x 10(9)//l) to a maximum of 146 x 10(9)//l (107-248 x 10(9)//l). In conclusion, ARC1779 markedly increases VWF/FVIII levels and most importantly improves or even corrects thrombocytopenia in VWD type 2B patients. This underscores the in vivo potency of ARC1779.