Coagulation assays for assessing the bypassing of factor Xa direct oral anticoagulants using VMX-C001

Figures

• 

  Figure 1. Interindividual variation and population averages for dPT clotting times measured in (A) undiluted and (B) 2.5× diluted individual plasma samples from healthy volunteers and for dRVVT clotting times in (C) 3× diluted and (D) 4× diluted individual plasma samples from healthy volunteers in samples without FXa DOAC (gray box plots) and spiked in vitro with apixaban 200 ng/mL (blue box plots). One subject with a clotting time above the limit of 300 seconds was removed from (B) as an outlier. ∗∗∗∗P < .0001 Apixaban vs no FXa DOAC (Wilcoxon test). DOAC, direct oral anticoagulant; dPT, dilute prothrombin time; dRVVT, dilute Russell viper venom time; FXa, factor Xa.

• 

  Figure 2. Effect of apixaban on dPT and dRVVT assays. Clotting times were obtained in (A) undiluted and (B) 2.5× diluted plasma for dPT and in (C) 3× and (D) 4× diluted plasma for dRVVT in individual plasma samples from 22 healthy volunteers spiked in vitro with apixaban (0-400 ng/mL). The dotted lines represent the minimum and maximum values of clotting times in the absence of apixaban (control). In all panels, the clotting times for each concentration of apixaban were compared with those for the samples without apixaban using the paired Friedman test for multiple comparisons. All comparisons were significant (P < .05, not indicated in the figure) except for those indicated in the figure. dPT, dilute prothrombin time; dRVVT, dilute Russell viper venom time.

• 

  Figure 3. Evaluation of the (A) dPT and (B) dRVVT assays measured in 2.5× and 4× diluted plasma, respectively, in individual plasma samples from 22 healthy volunteers spiked in vitro with apixaban (0-400 ng/mL) without (control) and with VMX-C001 (30 μg/mL). The clotting time upper limit was 300 seconds. The dotted lines represent the minimum and maximum values for the control samples with 0-ng/mL apixaban. Comparisons between samples with and without VMX-C001 at each concentration of apixaban were made using a 2-way anova for multiple comparisons: ∗P < .05; ∗∗P < .01; ∗∗∗P < .001; ∗∗∗∗P < .0001. dPT, dilute prothrombin time; dRVVT, dilute Russell viper venom time; ns, not significant.

• 

  Figure 4. Dose-dependent reversal by VMX-C001 (15, 30, and 60 μg/mL) (A, C, E, and G) and andexanet (50, 100, and 200 μg/mL) (B, D, F, and H) of the dose-dependent prolongation of the dPT (A–D) and dRVVT (E–H) clotting times measured in NPP spiked in vitro with apixaban (0-1600 ng/mL). Dotted lines represent maximal readout times for both assays. dPT, dilute prothrombin time; dRVVT, dilute Russell viper venom time; NPP, normal pooled platelet-poor plasma.

• 

  Figure 5. Effect of 4F-PCC on apixaban-induced prolongation of dPT (A, B) and dRVVT (C, D) clotting times in normal pooled plasma samples spiked in vitro with 4F-PCC (0.25 and 0.5 IU/mL) and increasing concentrations of apixaban (0-1600 ng/mL). (E‒H) The dose-dependent reversal by VMX-C001 of the prolonged clotting times in the presence of 4F-PCC (0.30 IU/mL). 4F-PCC, 4-factor prothrombin complex concentrates; dPT, dilute prothrombin time; dRVVT, dilute Russell viper venom time.

• 

  Figure 6. Time course of dPT assay results in healthy older subjects who received apixaban (A) or rivaroxaban (B) and time course of dRVVT assay results in subjects who received apixaban (C) or rivaroxaban (D). dPT and dRVVT were measured in 2.5× and 4× diluted plasma, respectively, in samples taken from healthy subjects who received apixaban (5 mg twice daily) or rivaroxaban (20 mg once daily) for 3.5 days, followed by a single intravenous infusion of placebo for 5 minutes, started 5 minutes after the last dose of FXa DOAC on day 4. Citrated platelet-poor plasma samples were obtained immediately before the final dose of apixaban or rivaroxaban (trough), at the end of placebo infusion (00:05 [EOI]), and at various times up to 144 hours after the start of infusion. The dotted line represents the median level at 144 hours when the effects of apixaban or rivaroxaban had worn off. The data at each time point were compared with those at 144 hours using the paired Friedman test; all comparisons were significant (P < .05, not indicated in the figure) except for those indicated in the figure. DOAC, direct oral anticoagulant; dPT, dilute prothrombin time; dRVVT, dilute Russell viper venom time;…

• 

  Figure 7. Correlation between dPT and dRVVT clotting times in healthy older subjects who received apixaban (top panels; n = 8 with 49 points) or rivaroxaban (bottom panels; n = 5 with 30 points) and apixaban or rivaroxaban plasma levels measured using LC-MS/MS. Individual subjects are represented by different colored points. dPT, dilute prothrombin time; dRVVT, dilute Russell viper venom time; LC-MS/MS, liquid chromatography-tandem mass spectrometry.

• 

  Figure 8. Correlation between dPT and dRVVT clotting times in nursing home residents, who were receiving rivaroxaban (top panels; n = 17), apixaban (middle panels; n = 30), or edoxaban (bottom panels; n = 8). The gray areas in each panel indicate the values obtained from healthy volunteers. Plasma concentrations of rivaroxaban, apixaban, and edoxaban were measured with the Ratachrom/STA-Liquid anti-Xa reagent on a Siemens CS-2500 coagulation analyzer. dPT, dilute prothrombin time; dRVVT, dilute Russell viper venom time.