Various laboratory protocols for measuring thromboxane A2 generation to detect the effectiveness of acetylsalicylic acid therapy (original) (raw)
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Journal of Thrombosis and Thrombolysis, 2023
Despite decades of investigations, the optimal assessment of the "therapeutic response" to early after loading dose of acetylsalicylic acid (ASA) remains unclear. Limited information is available on the relation between pharmacodynamic (PD) and pharmacokinetic (PK) measurements assessed immediately after ASA administration. Serial PD and PK analyses were performed immediately after a single 162 or 650 mg dose of chewed and swallowed ASA in ten healthy adults. ASA response was defined as > 95% inhibition of serum thromboxane (Tx)B 2, < 550 aspirin reaction units (ARU) by VerifyNow Aspirin (VN) test, and ≤ 20% arachidonic acid (AA)-induced platelet aggregation (PA). Correlation analyses between PK and PD measurements and receiver operating characteristic (ROC) curve analyses were performed. ASA response measured by VN test and AA-induced PA was achieved within 30 min of ASA administration. A correlation was observed between ARU and AA-induced maximum PA (r = 0.69, p < 0.001), serum TxB 2 (r = 0.74 and p < 0.001), and serum TxB 2 inhibition (r = 0.79, p < 0.001). In ROC curve analyses, ≤ 558 ARU and ≤ 7% AA-induced PA were associated with > 95% inhibition of TxB 2. 686 ng/ml plasma ASA cutoff point was associated with > 95% inhibition of serum TxB 2 , ≤ 7% 1 mM AA-induced PA, and ≤ 585 ARU. A modest ~ 50% inhibition of TxB 2 inhibition was associated with marked inhibition of 1 mM AAinduced platelet aggregation by LTA. Our analyses demonstrated important relationships between pharmacodynamic, and pharmacokinetic parameters measured immediately following oral ASA and cutoff values for ARU and AA-induced PA that is associated with > 95% inhibition of serum TxB 2 .
Journal of Thrombosis and Thrombolysis, 2020
Arachidonic acid (AA)-induced platelet aggregation (PA) and serum thromboxane B 2 (TxB 2) inhibition are widely used to indicate cyclooxygenase-1 activity and the antiplatelet effect of acetylsalicylic acid (ASA). Despite decades of investigations, the relation between these measurements remains unclear. We sought to evaluate the relation between AA-PA and serum TxB 2 inhibition. We serially measured AA-PA (conventional aggregation), serum TxB 2, plasma ASA and salicylic acid (SA) (liquid chromatography-mass spectrometry), and urinary 11-dehydro thromboxane B 2 (u11-dh TxB 2) (enzymelinked immunosorbent assay) levels at 10 times over 24 hours in seventeen healthy volunteers receiving a single dose of 162 mg chewed and swallowed ASA (n = 6), 50 mg inhaled ASA (n = 6), or 100 mg inhaled ASA (n = 5) (ClinicalTrials. gov Identifier: NCT04328883, April 1, 2020). Baseline variability was more pronounced with serum TxB 2 (31-680 ng/mL) as compared to maximal AA-PA (65-81%) and u11-dh TxB 2 (1556-4440 pg/mg creatinine). The relation between serum TxB 2 inhibition and AA-PA was stepwise; after 30-40% inhibition of serum TxB 2 , AA-PA fell to < 5%. By receiver operating characteristic curve analysis using AA-PA < 5% to define aspirin responsiveness, serum TxB2 inhibition > 49% and u11-dh TxB2 < 1520 pg/mg creatinine met the definition. Our study demonstrates a non-linear relation between serum TxB 2 inhibition and AA-PA. Aggregation was nil once TxB 2 inhibition reached > 49%. Moreover, these results suggest that the definition of > 95% inhibition of serum TxB 2 to indicate the level of platelet COX-1 inhibition needed for clinical efficacy may be overestimated and should be reconsidered in future translational research investigations that attempt to link the clinical efficacy of ASA with a laboratory measurement cutoff.
11-Dehydrothromboxane B2: A Quantitative Index of Thromboxane A2 Formation in the Human Circulation
Proceedings of The National Academy of Sciences, 1986
In human plasma, 11-dehydrothromboxane (TX) B2 is a major long lived metabolite (t1/2 45 min) formed from infused TXB2, the hydration product of biologically active TXA2. Plasma concentrations of TXB2 itself are readily confounded by ex vivo platelet activation and, theoretically, an enzymatic derivative of this compound, not subject to formation in whole blood, would more accurately reflect TXA2 formation in vivo. To address this hypothesis, we developed a sensitive assay for both 11-dehydro-TXB2 and TXB2, using capillary gas chromatography/negative-ion chemical ionization mass spectrometry. We established that whole blood possesses a minor capacity to form 11-dehydro-TXB2, attributable to nonenzymatic formation in erythrocytes. However, the nonenzymatic formation of 11-dehydro-TXB2 was not a practical limitation to its use as an index of TX biosynthesis. Blood was drawn from healthy volunteers (i) via an indwelling catheter at the time of insertion and at 30, 60, 90, 180, and 240 min thereafter and (ii) via separate venipunctures at 0 time and at 90 and 240 min thereafter. Plasma TXB2 drawn via the catheter at baseline (66 ± 63 pg/ml) was substantially greater than the maximal estimate of endogenous TXB2 (1-2 pg/ml) in plasma [Patrono, C., Ciabattoni, G., Pugliese, F., Perruci, A., Blair, I. A. & FitzGerald, G. A. (1986) J. Clin. Invest. 77, 590-594] and increased in magnitude and variance over time (339 ± 247 pg/ml at 240 min). By contrast, 11-dehydro-TXB2 did not change significantly in the sequential catheter samples or in the samples drawn by separate venipuncture. Basal plasma concentrations in volunteers were depressed by pretreatment with 325 mg of aspirin. Furthermore, the range of. concentrations in patients with severe atherosclerosis in whom urinary 2,3-dinor-TXB2 was increased was significantly higher (5-50 pg/ml, P < 0.01) than in healthy subjects (0.9-1.8 pg/ml). Concentrations of 11-dehydro-TXB2 were increased in patients who had recently suffered a pulmonary embolism to a greater extent than either the 11-dehydro-13,14-dihydro-15keto-TXB2 or the 2,3-dinor-TXB2 metabolites in plasma. These results indicate that plasma TXB2 is readily confounded by platelet activation ex vivo. Measurement of enzymatic metabolites of TXB2 minimizes this problem. The 11-dehydro metabolite is the most appropriate analytic target to detect phasic release of TXA2 in the human circulation, such as might occur in human syndromes of platelet activation.