Synergistic inhibitory effects of clopidogrel and rivaroxaban on platelet function and platelet-dependent thrombin generation in cats

Figures

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  FIGURE 1. Flow diagram illustrating drug administration and blood sampling timeline. Clop, Clopidogrel; DAT, dual antithrombotic treatment; Riva, Rivaroxaban.

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  FIGURE 2. Scatter dot plots demonstrating platelet inhibition, measured as percent (%) inhibition, in 9 cats after 7 days of rivaroxaban (RIVA), clopidogrel, or dual antithrombotic treatment (DAT) with rivaroxaban and clopidogrel. ADP‐induced whole blood impedance aggregometry was measured as maximum aggregation (Ag max), velocity and area under the cure (AUC). Percent inhibition >0% indicates platelet inhibition (up arrows) and inhibition (%) < 0% indicates platelet activation (down arrows) after 7 days of treatment. Clopidogrel alone and DAT resulted in significant inhibition compared to Riva alone. Riva alone resulted in platelet activation in 7 of the 9 cats. Line represents median and error bars represent interquartile range. *P < .05; **P < .005; ***P < .001.

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  FIGURE 3. Scatter dot pots demonstrating platelet activation measured as number of P‐selectin positive platelets (%) or response to agonists, ADP or thrombin, measured as fold change in median fluorescence intensity (MFI FC log 10), by flow cytometry in 9 cats treated with 7 days of rivaroxaban (RIVA), clopidogrel (CLOP) or dual antithrombotic treatment (DAT). (A) Only cats treated with DAT had significantly lower number of P‐selectin‐positive (%) resting (unstimulated) platelets compared to single agent treatment with RIVA or CLOP. (B) When platelets were treated with 20 μM ADP, CLOP or DAT significantly modulated response to ADP, while 7 days of RIVA treatment resulted in an increase response to ADP, which were significantly higher on D7 compared to CLOP or DAT. (C) Only DAT modulated platelet response to 0.005 U/mL thrombin, as shown by decreased percentage of P‐selectin‐positive platelets, and MFI FC from D0 to D7. Response to thrombin was significantly higher on D7 in cats treated with RIVA alone. *P < .05; **P < .005; ***P < .001; ****P < .0005.

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  FIGURE 4. Representative thrombograms demonstrating continuous thrombin generation, measured as fluorescent signal in arbitrary unit (AU), in platelet rich plasma in a cat before and after 3 days of rivaroxaban treatment (RIVA) (A) and 7 days of dual antithrombotic treatment (DAT) with rivaroxaban and clopidogrel (B). Data points were derived from means of duplicate measurements at a single time point for 60 minutes. Note the flattening of thrombogram following DAT compared to rivaroxaban treatment alone.

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  FIGURE 5. Scatter dot plots demonstrating platelet dependent thrombin generation over time in 9 cats, measured as (A) time (seconds) to maximum thrombin generation (T max), (B) peak thrombin generation (Relative Fluorescence Unit [RFU]), (C) thrombin generation potential (Area Under the Curve [AUC]) over 1 hour and (D) maximum velocity (V max, unit/s). After 7 days of treatment with rivaroxaban (RIVA), T max and peak thrombin generation were significantly modulated from D0 to D7. In addition to T max and peak thrombin generation, dual agent treatment (DAT) modulated thrombin generation potential and maximum velocity of thrombin generation from D0 to D7. Compared to RIVA, DAT also significantly prolonged T max compared to clopidogrel (CLOP) alone on D7. *P < .05; **P < .005; ***P < .001.