No abstract

Aspirin and clopidogrel are key anti-thrombotic therapies. Results from platelet reactivity testing during therapy, have been shown to correlate with future events and would allow for the optimisation of therapy. However, there is little agreement among current tests and there remains a clear clinical need for a universal standardised test. It was the objective of this study to explore the potential of 96-well plate aggregometry as a definitive clinical test of platelet reactivity with respect to aspirin and clopidogrel. A small non-blinded trial of 16 healthy male volunteers assigned to seven days of aspirin (75mg/day) or clopidogrel (75mg/day) therapy. Blood was collected before and on day 7 of treatment. Platelet aggregation was measured using a 96-well plate based aggregation method, and thrombi adhesion measured by colourimetric assay. Platelet agonists used were ADP (0.1-30microM), arachidonic acid (0.03-1.3mM), collagen (0.1-30microg/ml), adrenaline (0.001-100microM), ristocetin (0.2-3mg/ml), TRAP6 amide (0.130microM) and U46619 (0.130microM). Concentration response curves were constructed to each agonist under the various conditions and used to extract data such as log EC(50), Hill slope, and area under the curve. These demonstrated low intra- and inter-assay variability and strong discrimination of drug effects. This study demonstrates the ability of the 96-well plate based aggregation and adhesion method to detect and differentiate between stable aspirin and clopidogrel treatment in healthy volunteers. Moreover, this assay marries the ability to test subjects or patients using a range of platelet agonists with more rapidity and ease than the current gold standard platelet assay, traditional light transmission aggregometry, making it a serious alternative assay for use in clinical settings.

SummaryBackground: Strong P2Y12 blockade, as can be achieved with novel anti-platelet agents such as prasugrel, has been shown in vitro to inhibit both ADP and thromboxane A2-mediated pathways of platelet aggregation, calling into question the need for the concomitant use of aspirin. Objective: The present study investigated the hypothesis that aspirin provides little additional anti-aggregatory effect in a group of healthy volunteers taking prasugrel. Study participants/methods: In all, 9 males, aged 18 to 40 years, enrolled into the 21-day study. Prasugrel was loaded at 60 mg on day 1 and maintained at 10 mg until day 21. At day 8, aspirin 75 mg was introduced and the dose increased to 300 mg on day 15. On days 0, 7, 14 and 21, platelet function was assessed by aggregometry, response to treatments was determined by VerifyNow™ and urine samples were collected for quantification of prostanoid metabolites. Results: At day 7, aggregation responses to a range of platelet agonists were reduced and there was only a small further inhibition of aggregation to TRAP-6, collagen and epinephrine at days 14 and 21, when aspirin was included with prasugrel. Urinary prostanoid metabolites were unaffected by prasugrel, and were reduced by the addition of aspirin, independent of dose. Conclusions: In healthy volunteers, prasugrel produces a strong anti-aggregatory effect, which is little enhanced by the addition of aspirin. The addition of aspirin as a dual-therapy with potent P2Y12 receptor inhibitors warrants further investigation.

This study evaluated the effect of prasugrel alone, and in combination with low and high dose aspirin, on urinary metabolites of thromboxane A2 (TXA2) and prostaglandin I2 (PGI2), TX-M and PGI-M, respectively. 9 healthy males, aged 18–40, were enrolled in the 21-day study. Prasugrel was loaded at 60 mg on day 1 and maintained at 10 mg once daily until day 21. At day 8 aspirin 75 mg o.d. was introduced and the dose increased to 300 mg o.d. on day 15. On days 0, 7, 14 and 21 urine samples were obtained and TX-M and PGI-M assayed by mass spectrometry. Platelet aggregation to a TXA2-mimetic (U46619) was also determined at each time point. Data are presented as ng of urinary metabolite per mg creatine (mean±SEM). Data were analysed by one-way ANOVA with Bonferroni's post-test. Prasugrel alone did not reduce either TX-M (day 7, 0.24±0.02 ng per mg creatine; day 0, 0.28±0.04) or PGI-M (day 7, 0.15±0.03; day 0, 0.15±0.04), although both metabolites were reduced by the addition of low dose aspirin (day 14: TX-M, 0.09±0.01; PGI-M, 0.07±0.01; p<0.01 vs day 0). There were no further effects with the higher aspirin dose (p>0.05). Platelet aggregation to U46619 was largely inhibited by prasugrel, with no further effect of aspirin. Prasugrel alone preserves urinary PGI2 metabolites while inhibiting TXA2-driven platelet aggregation. Addition of aspirin reduces the excreted levels of metabolites of both TXA2 and vasoprotective PGI2.