aPTT Measurements Research Articles

Prothrombin and activated partial thromboplastin times, thromboelastography, hematocrit, and platelet count in a feline hemorrhage/over-resuscitation model using lactated Ringer's solution or 6% tetrastarch 130/0.4.

To describe and compare prothrombin time (PT), activated partial thromboplastin time (aPTT), thromboelastography (TEG), HCT, and platelet count measurements in a hemorrhage/over-resuscitation model. Randomized crossover study. University teaching hospital. Six cats. Anesthetized cats underwent 3 treatments at 2-month intervals. The treatments were as follows: NHR-no controlled hemorrhage and sham resuscitation; LRS-controlled hemorrhage and lactated Ringer's solution (LRS) for resuscitation; and Voluven-controlled hemorrhage and 6% tetrastarch 130/0.4 for resuscitation. The LRS and Voluven were administered at 60 and 20mL/kg/h, respectively, for 120minutes. Blood samples were drawn for PT, aPTT, TEG, HCT, and platelet count measurements at a healthy check (T - 7d), after controlled hemorrhage (T0), at 60 and 120minutes of resuscitation (T60 and T120), and at 24hours after completion of resuscitation (T24h). Data were analyzed using a general linear mixed model approach (significance was P<0.05). Total median blood loss (controlled hemorrhage and blood sampling from T0 to T120) at T120 was 11.4, 31.0, and 30.8mL/kg for NHR, LRS, and Voluven, respectively. PT and aPTT during LRS and Voluven were prolonged at T60 and T120 compared to NHR (P<0.001). On TEG, the reaction time, kinetic time, and alpha-angle were within reference intervals for cats at all time points in all treatments, while maximum amplitude was less than the reference interval (40mm) at T0, T60, and T120 during Voluven and at T60 and T120 during LRS compared to NHR (both P<0.001). The HCT and platelet count were significantly lower at T60 and T120 during LRS and Voluven compared to NHR (P<0.001). Hypocoagulopathy was observed during hemorrhage and liberal fluid resuscitation. Prolongation of PT and aPPT and decreased clot strength may have been caused by hemodilution and platelet loss.

Are shortened aPTT values always to be attributed only to preanalytical problems?

It has been recognized that shortened activated partial thromboplastin time (aPTT) may be caused by various preanalytical conditions. As coagulation Factor VIII is included in the invitro intrinsic coagulation cascade measured by aPTT, we hypothesized that the shortened aPTT could be a result of elevated FVIII activity. We aimed to inspect the connection of elevated FVIII with shortened aPTT, and the possible effect inflammation has on routine laboratory parameters. 40 patients from various hospital departments with aPTT measurement below the lower limit of the reference interval (<23.0 s) were included in the study. To compare the obtained results with aPTT measurements in the non-inflammatory state, samples from 25 volunteers (laboratory personnel) were collected. White blood cell count, C-reactive protein, aPTT, and FVIII values were measured in the control group. Only two samples among 40 patients with shortened aPTT (5 %) were clotted. Out of the remaining 38, 26 had FVIII activity above 150 % (upper limit of a reference interval), median value of 194 % (IQR: 143-243 %). Seven samples in the control group had shortened aPTT results (36 %). However, all coagulation samples were clot and hemolysis-free. Multiple regression identified only FVIII activity as an independent variable in predicting aPTT values (p=0.001). Our results support the thesis that shortened aPTT is rarely a consequence of preanalytical problems. Elevated FVIII activity causes shortened aPTT, not only in theinflammatory state but also in individuals with concentration of inflammatory markers within reference intervals.

Applications of rotational thromboelastometry in heparin monitoring in critical COVID-19 disease: Observations in the Maastricht Intensive Care COVID cohort

BackgroundCritically ill COVID-19 patients are at risk for venous thromboembolism (VTE). Therefore, they receive thromboprophylaxis and, when appropriate, therapeutic unfractionated heparin (UFH) or low-molecular-weight heparin (LMWH). To monitor heparins in COVID-19 disease, whole-blood rotational thromboelastometry (ROTEM) may be a promising alternative to the aPTT and anti-Xa assays. ObjectiveTo evaluate the ROTEM INTEM/HEPTEM ratios in mechanically ventilated COVID-19 patients treated with UFH and therapeutic LMWH. Material and methodsA subcohort of mechanically ventilated COVID-19 patients of the prospective Maastricht Intensive Care Covid (MaastrICCht) cohort was studied. Anti-Xa, aPTT, and ROTEM measurements following treatment with UFH or therapeutic dose of LMWH (nadroparin) were evaluated using uni- and multivariable linear regression analysis and receiver operating characteristics. ResultsA total of 98 patients were included, of which 82 were treated with UFH and 16 with therapeutic LMWH. ROTEM-measured INTEM/HEPTEM CT ratio was higher in patients using UFH (1.4 [1.3–1.4]) compared to patients treated with LMWH (1.0 [1.0–1.1], p < 0.001). Both the aPTT and anti-Xa were associated with the CT ratio. However, the β-regression coefficient (95%CI) was significantly higher in patients on UFH (0.31 (0.001–0.62)) compared to therapeutic LMWH (0.09 (0.05–0.13)) for comparison with the anti-Xa assay. Furthermore, ROC analysis demonstrated an area under the curve for detecting UFH of 0.936(0.849–1.00), 0.851(0.702–1.000), and 0.645(0.465–0.826) for the CT ratio, aPTT, and anti-Xa, respectively. ConclusionThe ROTEM INTEM/HEPTEM CT ratio appears a promising tool to guide anticoagulant therapy in ICU patients with COVID-19 disease, but associations with clinical endpoints are currently lacking.

Monitoring heparin therapy: stability of two different anti-Xa assays using blood samples collected in citrate-containing and CTAD tubes

BackgroundAnti-factor Xa assays and activated partial thromboplastin time (aPTT) are mainly employed to monitor patients treated with heparins. According to the Clinical and Laboratory Standards Institute and the French Working Group on Haemostasis and Thrombosis, anti-factor Xa activity and aPTT should be tested within 2 h of blood sampling for unfractionated heparin (UFH) monitoring. However, discrepancies exist depending on the used reagents and collecting tubes. The study aim was to determine the stability of aPTT and anti-factor Xa measurements using blood samples collected in citrate-containing or citrate-theophylline-adenosine-dipyridamole (CTAD) tubes and stored for up to 6 h.MethodsPatients receiving UFH or low molecular weight heparin (LMWH) were enrolled; aPTT and anti-factor Xa activity were tested using two different analyser/reagent pairs (Stago and reagent without dextran sulfate; Siemens and reagent with dextran sulfate) after 1, 4 and 6 h of sample storage as whole blood or as plasma.ResultsFor UFH monitoring, comparable anti-factor Xa activity and aPTT results were obtained with both analyser/reagent pairs when samples were stored as whole blood before plasma isolation. With samples stored as plasma, anti-factor Xa activity and aPTT were not affected up to 6 h after sampling when using the Stago/no-dextran sulfate reagent pair. With the Siemens/dextran sulfate-containing reagent, aPTT was significantly altered after 4 h of storage. For LMWH monitoring, anti-factor Xa activity remained stable (whole blood and plasma) for at least 6 h. Results were comparable with citrate-containing and CTAD tubes.ConclusionsAnti-factor Xa activity in samples stored as whole blood or plasma was stable for up to 6 h, regardless of the reagent (with/without dextran sulfate)/collection tube. Conversely, aPTT was more variable because other plasma parameters can influence its measure and complicate the interpretation of its variations after 4 h.

Comparison of a Fully Weight-Based Protocol with a Non-Weight-Based Dosage Titration Protocol for IV Unfractionated Heparin: A Before-and-After Study.

Unfractionated heparin (UFH) is used for the prevention and treatment of arterial or venous thromboembolism. The dosage for IV infusion of UFH is generally based on the patient's weight, with adjustment to a specific target for activated partial thromboplastin time (aPTT). In May 2019, the UFH protocols at the study institution were changed from being fully weight-based (i.e., for both initial dosing and subsequent dosage titrations) to weight-based initial dosing and non-weight-based dosage titrations, but the relative effectiveness of these 2 approaches was not known. The primary objective was to compare the effectiveness in achieving therapeutic aPTT with the fully weight-based and non-weight-based dosage titration protocols. The secondary objective was to compare the effectiveness of the non-weight-based dosage titration protocol with that of the previous fully weight-based one for patients with low-target aPTT. A single-centre, retrospective, observational before-and-after study was conducted for patients receiving therapeutic UFH for any indication. Patients in the "before" group (fully weight-based protocol) were treated from January 2015 to October 2016, and those in the "after" group (non-weight-based titration) from January to October 2020. From a total of 1969 charts screened, 137 patients treated according to the fully weight-based protocols and 130 patients treated according to the non-weight-based titration protocols were included. In terms of the co-primary objective, the median number of dosage adjustments to achieve therapeutic anticoagulation was 1 in both groups (p = 0.48), and the proportion of patients with therapeutic anticoagulation at 24 h was similar (96.2% [125/130] with the non-weight-based titration protocols versus 99.3% [136/137] with the fully weight-based protocols; p = 0.09). Among patients treated according to the low-target UFH protocols, those with the non-weight-based titration protocol were less likely to have therapeutic anticoagulation at first measurement of aPTT than those with the fully weight-based protocol (37.9% [25/66] versus 44.6% [41/92], p = 0.033). This retrospective, observational, before-and-after study showed that the effectiveness of the non-weight-based dosage titration protocols in achieving therapeutic aPTT was similar to that of fully weight-based UFH protocols.

Prevalence and Impact of Lupus Anticoagulant in Patients Receiving Extracorporeal Membrane Oxygenation.

Monitoring of blood coagulation is essential in ECMO patients. We investigated the prevalence of lupus anticoagulant (LA) and its association with coagulation testing and hemostaseologic complications in patients treated with ECMO. This is a retrospective analysis including adult patients who received ECMO at a medical intensive care unit at the Medical University of Vienna. The primary outcome was the prevalence of LA. Secondary outcomes included conditions associated with LA positivity, rates of bleeding and thromboembolic events, as well as the proportions of aPTT and antiXa measurements within the target range. Between 2013 and 2021 193 patients received ECMO, in 62 (32%) of whom LA diagnostics were performed. Twenty-two (35%) patients tested positive. LA positive patients had more frequently received VV ECMO (77.3% vs 34.3%; p = 0.002), were more frequently diagnosed with viral respiratory infections (SARS-CoV2: 45.5% vs 20%; p = 0.041, influenza virus: 22.7% vs 0%; p = 0.003), had a longer ECMO treatment duration (25 vs 10 days; p = 0.011) and a longer ICU stay (48 vs 25 days; p = 0.022), but similar rates of bleeding and thromboembolic events.

Risk for excessive anticoagulation during hemodialysis is associated with type of vascular access and bedside coagulation testing: Results of a cross-sectional study.

Recommendations and practice patterns for heparin dosing during hemodialysis show substantial heterogeneity and are scantly supported by evidence. This study assessed the variability in unfractionated heparin (UFH) dosing during hemodialysis and its clinical and biological anticoagulatory effects, and identified explanatory factors of heparin dosing. Cross-sectional study assessing UFH dosing, coagulation tests - activated partial thromboplastin time (aPTT) and activated clotting time (ACT) before dialysis start, 1 h after start and at treatment end (4 h) - and measurement of residual blood compartment volume of used dialyzers. 101 patients, 58% male, with a median dialysis vintage of 33 (6-71) months received hemodialysis using a total UFH dose of 9,306 ± 4,079 (range 3,000-23,050) IU/session. Use of a dialysis catheter (n = 56, 55%) was associated with a 1.4 times higher UFH dose (p < 0.001) irrespective of prior access function. aPTT increased significantly more than ACT both 1 h and 4 h after dialysis start, independent of the dialysis access used. 53% of patients with catheter access and ACT ratio < 1.5, 1 h after dialysis start had simultaneous aPTT ratios > 2.5. Similar findings were present at 1 h for patients with AVF/AVG and at dialysis end for catheter use. No clinically significant clotting of the extracorporeal circuit was noted during the studied sessions. Dialyzer's blood compartment volume was reduced with a median of 9% (6-20%) without significant effect of UFH dose, aPTT or ACT measurements and vascular access type. UFH dose adaptations based on ACT measurements frequently result in excessive anticoagulation according to aPTT results. Higher doses of UFH are used in patients with hemodialysis catheters without evidence that this reduces dialyzer clotting.

Evaluation of Platelet Indices, PT and PTT in Sudanese Chronic Renal Failure Patients at Khartoum state, 2022

Background: Chronic renal failure is a global public health problem; Kidney transplantation and dialysis are possible solutions of kidney failure. The aim of our study was to assess the platelet count, platelet indices, prothrombin time (PT) and partial thromboplastin time (PTT) in Sudanese chronic renal failure patients at Khartoum sate, 2022.&#x0D; Materials and Method: This was analytical case control study conducted at Khartoum state. A total of 200 blood samples were collected (100 samples from chronic renal failure patients and 100 healthy individuals from control group). A total of five ml of whole blood was collected from each participant and divided into EDTA anticoagulated tube for platelet count and platelet indices, and into sodium citrate anticoagulated tube. for PT and PTT measurement. Examination of platelet count and platelet indices was done by using Haematology analyser (Sysmex-XP-300), where PT and APTT measurement was done by using coagulation analyser (Coatron-M-1). &#x0D; Results: The study revealed that in the case group; the mean age was (48.19± 15.53), 42.1% of them were female and 57.1% were male, where in the control group; the mean age was (45.54 ± 16.82), 57.9% were female and 42.9% were male. Regarding to the haematological results; the mean platelet count and PCT of the case group was significantly decreased when compared with the control group with P-value of 0.000. where the mean average of MPV, PDW and PLCR showed no statistically significant differences between cases and control with P-value of &gt; 0.05. According to the PT and PTT; they were increased in the case group when compared with the control group with P-value of 0.000.&#x0D; Conclusion: The study concluded that platelet count and plateletcrit was decreased, where mean platelet volume and platelet distribution width in addition to PLCR, showed no statstically significant difference. PT and PTT were increased in chronic renal failure patients.&#x0D; Keywords: Platelet count, Platelet indices, PT, PTT, chronic renal failure, haemodialysis, bleeding

A Comparison of Activated Partial Thromboplastin Time and Activated Coagulation Time for Anticoagulation Monitoring during Extracorporeal Membrane Oxygenation Therapy.

Unfractionated heparin is used to prevent coagulation activation in patients undergoing extracorporeal membrane oxygenation (ECMO) support. We designed this study to determine the preferable indicator for anticoagulation monitoring. We conducted a retrospective study and divided the patients into an activated coagulation time (ACT)-target group and an activated partial thromboplastin time (aPTT)-target group. The correlations between ACT, aPTT, and the heparin dose were explored. Thirty-six patients were included (19 aPTT-target and 17 ACT-target patients); a total of 555 matched pairs of ACT/aPTT results were obtained. The correlation between the ACT and aPTT measurements was Spearman's Rank Correlation Coefficient (rs) = 0.518 in all 555 pairs. The Bland-Altman plot showed data points outside the displayed range (51.2-127.7), suggesting that the agreement between ACT and aPTT was poor. The aPTT group had fewer heparin dose changes (2.12 ± 0.68 vs. 2.57 ± 0.64, p = 0.05) and a lower cumulative heparin dose (317.6 ± 108.5 vs. 396.3 ± 144.3, p = 0.00) per day than the ACT group. There was no difference in serious bleeding (9 vs. 5; p = 0.171) or embolism events (3 vs. 3; p = 1.0) or in the red blood cell and fresh frozen plasma transfusion volumes between the ACT- and aPTT-target groups. Similarly, there was no significant difference in the ECMO duration (9 [4-15] days vs. 4 [3-14] days; p = 0.124) or length of ICU hospitalization (17 [5-32] days vs. 13 [4-21] days; p = 0.451) between the groups. The correlation between ACT and aPTT and the heparin dose was poor. The aPTT group had fewer daily heparin dose changes and a lower cumulative heparin dose per day than the ACT group, with no more bleeding and thrombotic events. Therefore, we recommend aPTT rather than ACT to adjust heparin dose in the absence of better monitoring indicators.

Monitoring of Unfractionated Heparin Therapy in the Intensive Care Unit Using a Point-of-Care aPTT: A Comparative, Longitudinal Observational Study with Laboratory-Based aPTT and Anti-Xa Activity Measurement.

Continuous intravenous unfractionated heparin (UFH) is administered routinely in the intensive care unit (ICU) for the anticoagulation of patients, and monitoring is performed by the activated partial thromboplastin time (APTT) or anti-Xa activity. However, these strategies are associated with potentially large time intervals before dose adjustments, which could be detrimental to the patient. The aim of the study was to compare a point-of-care (POCT) version of the APTT to (i) laboratory-based APTT and (ii) measurements of anti-Xa activity in terms of correlation, agreement and turnaround time (TAT). Thirty-five ICU patients requiring UFH therapy were prospectively included and followed longitudinally for a maximum duration of 15 days. UFH was administered according to a local adaptation of Raschke and Amanzadeh’s aPTT nomograms. Simultaneous measurements of POCT-APTT (CoaguCheck® aPTT Test, Roche Diagnostics) on a drop of fresh whole blood, laboratory-based APTT (C.K. Prest®, Stago) and anti-Xa activity (STA®Liquid anti-Xa, Stago) were systematically performed two to six times a day. Antithrombin, C-reactive protein, fibrinogen, factor VIII and lupus anticoagulant were measured. The time tracking of sampling and analysis was recorded. The overall correlation between POCT-APTT and laboratory APTT (n = 795 pairs) was strongly positive (rs = 0.77, p < 0.0001), and between POCT-APTT and anti-Xa activity (n = 729 pairs) was weakly positive (rs = 0.46, p < 0.0001). Inter-method agreement (Cohen’s kappa (k)) between POCT and laboratory APTT was 0.27, and between POCT and anti-Xa activity was 0.30. The median TATs from sample collection to the lab delivery of results for lab-APTT and anti-Xa were 50.9 min (interquartile range (IQR), 38.4–69.1) and 66.3 min (IQR, 49.0–91.8), respectively, while the POCT delivered results in less than 5 min (p < 0.0001). Although the use of the POCT-APTT device significantly reduced the time to results, the results obtained were poorly consistent with those obtained by lab-APTT or anti-Xa activity, and therefore it should not be used with the nomograms developed for lab-APTT.

Hemostatic management of hemophilia

Replacement therapy is the basic treatment for hemophilia by infusing deficient clotting factors, including replacement therapy for prophylaxis (i.e., prevention of breakthrough bleeding for physical activity), episodic replacement therapy, replacement therapy during and after procedures and surgery, and replacement therapy for physical activity, each administered at doses and intervals appropriate for the purpose and the product used. Although emicizumab is increasingly used for prophylaxis in severe hemophilia A, the combination of replacement therapy is necessary, especially during severe bleeding, highly invasive procedures, and major surgery. Furthermore, the usual APTT measurements cannot be used for monitoring the replacement therapy and detecting the presence of inhibitors while patients are receiving emicizumab. Hemostatic management of patients with inhibitors should be implemented based on the purpose of the therapy, the latest inhibitor titer, and the clinical response to the product, with the choice of inhibitor neutralization or bypass agent therapy. When bypassing agents are used in patients with hemophilia A with emicizumab inhibitor during and after bleeding, procedures, and surgery, the choice of agent and dose adjustment should be made with attention to thrombotic complications.

Measurement of Plasma Fibrinogen Levels, PT and APTT among HIV Infected Patients: A Critical Bio-Marker for Coagulation Dysfunctions

Background: The literature stated that Human immunodeficiency virus (HIV) infection led to activation of coagulation, and habitually linked with an augmented risk of venous and arterial thrombosis. So the purpose of the study was to determine the plasma fibrinogen level in Sudanese HIV-infected patients.&#x0D; Materials and Methods: A total of one hundred participants were recruited, and classified into two groups; the case group include (50) HIV patients, and the control group enrolled (50) healthy individuals. Three ml of blood was collected. Fresh Poor Plasma was prepared from citrated venous blood by centrifuged for 15 minutes at 3000 pm. Fibrinogen levels were measured by an automated coagulation analyzer (Thrombolyzer XRC Germany). Data were collected using a directly structured questionnaire. Data were analyzed using SPSS Version 21. &#x0D; Results: The present study showed that the mean of plasma fibrinogen levels was statistically significantly higher in HIV infection in comparison with those normal healthy control (470.50 ±67.75 vs 214.75±21.25 with P-value 0.00). There was a significantly decreased level of PT, and PTT among the HIV group comparing with the control (9.575±0.64, and 22.39±4.94) VS (12.483±0.72, and 30.78±3.55) consequently, (P-value ≤0.001). Fibrinogen levels were significantly increased with the progression of HIV disease (469.84 ±67.15, 472.74 ±87.75, 478.47 ±61.92) in stage I, stage II, and stage III respectively.&#x0D; Conclusions: An HIV-infected patient had elevated plasma fibrinogen levels, as well as other coagulation dysfunctions.

An ex vivo comparison of partial thromboplastin time and activated clotting time for heparin anticoagulation in an ovine model.

Sheep are a primary model of mechanical circulatory support (MCS) with heparin anticoagulation therapy frequently being monitored by activated clotting time (ACT) due to ease and cost. In patients undergoing long-term heparin therapy, other anticoagulation monitoring strategies, such as activated partial thromboplastin time (aPTT), have proven to be more reliable indicators for the adequacy of anticoagulation, frequently determined by heparin concentration. As there is a paucity of similar studies in sheep, we sought to investigate the correlation between heparin concentration and ACT and aPTT using whole sheep blood in an ex vivo model. Fresh whole blood was serially drawn from an adult female Dorset-hybrid sheep and aliquots were placed into tubes containing heparin saline solutions with concentrations ranging from 0 to 7.81 U heparin per mL of whole blood. ACT and aPTT values were measured on each of the samples. The experiment was performed four times with the same animal. A simple linear regression was performed to determine correlation, and subgroup analysis was performed on low versus high heparin concentrations typically seen in human patients on long-term MCS, such as extracorporeal membrane oxygenation (ECMO), versus cardiopulmonary bypass, respectively. aPTT measurements versus the heparin concentration had an R2 =0.7295. ACT measurements versus the heparin concentration had a R2 =0.4628. aPTT measurements versus the ACT measurements had a R2 =0.2974. The strength of the correlation between aPTT and heparin concentration increased at low heparin concentrations (R2 =0.8392). aPTT had a more reliable correlation to heparin concentration and thus anticoagulation level than ACT. This was particularly true at lower heparin concentrations, similar to ranges seen for patients on ECMO. The correlation between aPTT and ACT values was poor. Further in vivo studies should be performed to confirm our results.

Comparison of the sensitivity of commercial aPTT tests in measurement of the pharmacodynamic response of milvexian, a novel FXIa inhibitor

Abstract Introduction The activated partial thromboplastin time (aPTT) assay is a test routinely used to evaluate abnormalities or deficiencies in coagulation factors of the intrinsic and common pathways. The composition of the surface activator and phospholipids in the aPTT reagent is known to influence the response of heparin and direct FXa or thrombin inhibitors. Milvexian (formerly referred to as BMS-986177/JNJ-70033093) is an investigational small-molecule Factor XIa (FXIa) inhibitor being studied for the prevention and treatment of major thrombotic conditions. Clinical pharmacology evaluation of milvexian includes aPTT as the primary pharmacodynamic assay, thus underscoring the need for a sensitive reagent to evaluate its anticoagulant activity. Purpose This study evaluated the sensitivity of six commercially available aPTT reagents in plasma from individual donors spiked with milvexian, at concentrations spanning the anticipated clinically relevant exposures. Methods Platelet-poor plasma (PPP) prepared from citrated whole blood collected from consenting, healthy adult volunteers (n=12) was spiked with vehicle (DMSO, 0.5% v/v) or 0.1–10 μM milvexian. The aPTT was measured using six commercially available aPTT reagent kits which included silica; kaolin, or ellagic acid as the contact activator, in combination with natural or synthetic phospholipids. The coagulation automated analysers used for testing were matched to the kit's manufacturer recommendations. All reagents and respective normal or abnormal controls were prepared as instructed by the manufacturer. Results Milvexian exhibited dose-dependent prolongation of aPTT with all reagents tested. Assays performed with aPTT reagents containing kaolin or ellagic acid demonstrated the highest sensitivity, as measured by the concentration that achieved a 2-fold aPTT prolongation (EC2x), and showed the widest dynamic range of response. Coefficient of variability of aPTT measurements in plasma from 12 individual donors was between 5.6–7.9% for Dade® Actin® FS, and 7.1–8% for STA®-C.K. Prest® 5. Conclusions Prolongation of aPTT in PPP spiked with milvexian exhibited a dose-dependent relationship, with statistically significant differences observed among reagents at milvexian concentrations above 0.3 μM. The highest sensitivity measured by changes in the ratio to baseline was obtained with aPTT reagents containing an activator/procoagulant phospholipid combination of ellagic acid + purified soy phosphatides or kaolin + cephalin, which performed similarly. Identification of the reagents with the best combination of sensitivity, precision, and dynamic range may help guide the selection of reagent for assessing milvexian activity. Funding Acknowledgement Type of funding sources: Private company. Main funding source(s): Janssen Research &amp; Development, LLC

Agreement between Anti-Factor Xa and Activated Partial Thromboplastin Time Measurements in Syncardia Total Artificial Heart Recipients Receiving Postoperative Anticoagulation with Unfractionated Heparin

<h3>Purpose</h3> The Activated Partial Thromboplastin Time (aPTT) has been the historical method of choice for anticoagulation monitoring in patients undergoing mechanical circulatory support with intravenous unfractionated heparin (IVUFH). However, the assay is susceptible to interference effects and is being progressively superseded by the anti-Factor Xa (anti-Xa) method. The study aim was to investigate the agreement between aPTT and anti-Xa measurements in patients receiving post-operative anticoagulation with IVUFH following Syncardia Inc. Total Artificial Heart (TAH) implantation. <h3>Methods</h3> A retrospective single arm, single centre analysis of 20 patients who underwent TAH implantation from July 2014 to October 2018 entailed simultaneous determinations of aPTT and anti-Xa. Agreement between these parameters was assessed using a novel statistical approach based on the Bland-Altman method. <h3>Results</h3> In spite of a positive correlation between aPPT and anti-Xa measurements, normal target ranges were poorly aligned: from 5^th to 30^th post-operative day, for anti-Xa values of 0.2 and 0.4 U/mL corresponding APTT values were 51.1 and 64.3 seconds, respectively. Moreover, white blood cell count was associated with a trend towards lower aPTT levels with respect to anti-Xa. Conversely, bilirubin level was significantly associated with an increase in aPTT levels with respect to anti-Xa values. <h3>Conclusion</h3> A novel method developed to investigate the agreement between aPTT and anti-Xa in the peri-operative phase of TAH therapy showed comparatively low levels of anticoagulation (with respect to historical target aPTT range) were not associated with thromboembolic sequalae. Methodology developed for this study is generalisable to other forms of mechanical circulatory support.

Real-World Clinical and Laboratory Characteristics in Severe Hemophilia a Patients Commencing Emicizumab

Introduction Emicizumab (a bispecific antibody mimicking Factor VIIIa) has been authorised for use in the United Kingdom National Health Service (NHS) in severe hemophilia A patients with and without inhibitors since August 2019. The Summary of Product Characteristics estimates a steady state of emicizumab achieved after four loading doses for patients &amp;gt;12 years of age at between 50-60ug/mL. Real world data outwith trial conditions are lacking to evaluate clinical efficacy, and which assays may be appropriate to determine emicizumab levels in vivo. Methods A prospective analysis of patients switching from FVIII concentrate to emicizumab was conducted at Guy's and St Thomas' Hospital Haemophilia Centre between September 2019 and July 2020 inclusive. Patients' age and weight were recorded and doses of emicizumab prescribed according to United Kingdom Haemophilia Centres' Doctors Organisation guidelines. Post emicizumab initiation, samples for coagulation testing were taken one week, four weeks and eight weeks after commencing emicizumab, with some pre-emicizumab initiation samples obtained. Three assays were used (human chromogenic FVIII assay (CSA) (Hyphen Biomed); bovine chromogenic FVIII assay (BCA) (Siemens); and a modified one-stage emicizumab assay (OSEA) with emicizumab calibrators to derive drug levels). Where possible with sufficient sample volume all three assays were run simultaneously. Results Within the cohort of all severe hemophilia A patients (n=118), 9 patients with inhibitors had already been previously switched from FVIII concentrate. 21 patients switched to emicizumab between September 2019 and July 2020, of which 2 patients had active FVIII inhibitors, with emicizumab prophylaxis patients comprising 25.4% (30/118) of the hospital cohort. Mean age of switchers was 39.3 years (standard deviation ± 13.9), with a mean dose of 116 ± 20.7mg on a weekly basis. 13/21 were on 1.5mg/kg weekly dosing and the remainder on fortnightly dosing at 3mg/kg. OSEA levels at 1 week (n=15), 4 weeks (n=15) and 8 weeks (n=7) were 19.7 ± 14.8ug/mL, 54.8 ± 11.7ug/mL and 61.3 ± 12.2ug/mL respectively. CSA results were 17 ± 9.0 IU/dL (n=9), 27.7 ± 6.7 IU/dL (n=10), and 26.7 ± 9.6 IU/dL (n=5) respectively. Simultaneous 1-stage based OSEA and CSA assays were performed on 20 samples, with high correlation between the two types of assay (Pearson correlation coefficient = 0.909 (CI 0.780-0.964, p &amp;lt;0.0001)). Six pre-emicizumab initiation samples were tested for both OSEA and CSA, which suggested low-level cross-reactivity of OSEA with FVIII concentrate in vivo. 4/6 were non-washout samples, which showed apparent positive OSEA levels (range 2.3-7.9ug/mL) with corresponding CSA ranges from 17.6-86.6 IU/dL. BCA in 3 patients at 1 week showed a low level of residual FVIII (all &amp;lt;8 IU/dL) consistent with prior FVIII infusions. All samples at 4 weeks and 8 weeks had undetectable FVIII concentrate levels consistent with the cessation of FVIII administration (bovine FVIII &amp;lt;1 IU/dL) and correspondingly positive OSEA levels, confirming the lack of cross-reactivity between the two assays. No patients developed anti-emicizumab antibodies as determined by clinical efficacy or by APTT measurement. No patients required any surgical or interventional procedures during this period. One spontaneous bleed and one traumatic bleed were recorded in two separate patients. Conclusion Emicizumab in a real-world setting is confirmed in our experience to have excellent clinical efficacy with infrequent episodes of spontaneous bleeding. 18% of patients switched to emicizumab within a year of its availability on the NHS. It has been well-tolerated with no clinical or laboratory evidence of anti-drug antibodies as measured by APTT. The OSEA correlated well with levels predicted by the emicizumab SPC at each time point. At all concentrations an OSEA and CSA are highly correlated, which may suggest that only one of the two assays is routinely required in laboratory use to assay emicizumab. Caution is however advised in assessing OSEA on its own when FVIII concentrate has recently been administered as this may result in some low level, false positive results. Figure Disclosures Austin: Chugai: Honoraria. Dolan:Pfizer: Consultancy; Roche: Honoraria.

The Influence of Haemostatic System Maturation on the Dose-Response Relationship of Unfractionated Heparin.

Unfractionated heparin (UFH) dosing and monitoring guidelines for children are often extrapolated from adult data. This practice is suboptimal given the inherent differences in haemostatic maturation and drug handling in children compared with adults. The aim of this work was to investigate the impact of haemostatic system maturation on the dose-response relationship of UFH in children. A quantitative model for haemostasis in adults was adapted to account for maturation in UFH pharmacokinetic (PK) parameters with and without age-related changes in coagulation factor concentrations. The adult and adapted models were used to predict the time courses of anti-factor Xa activity (aXa) and activated partial thromboplastin time (aPTT) in patients receiving UFH infusion. Predictions from both models were compared with observed aXa and aPTT measurements from 31 paediatric patients receiving UFH during extracorporeal membrane oxygenation (ECMO). The model with maturation for both UFH PK and the haemostatic system had an improved aXa and aPTT predictive performance compared with maturation in UFH PK only and the original adult model. Despite the minor effect of haemostatic system maturation on baseline aPTT, it led to substantial changes in the time course of aPTT sensitivity to UFH. This finding suggests that between-subject variability in clotting factors concentrations is potentially a major contributor to the overall variability of aPTT response to UFH. In addition, time-varying clotting factors concentrations may explain within-subject changes in aPTT sensitivity to UFH. We developed the first quantitative systems pharmacology (QSP) model that provides a mechanistic and quantitative basis for linking physiological and pharmacological maturation to UFH effect and response biomarkers. After appropriate clinical validation, the model could be useful for the development of paediatric-specific individualised UFH dosing recommendations.

C-reactive protein-induced activated partial thromboplastin time prolongation in heparinized samples is attenuated by elevated factor VIII.

Activated partial thromboplastin time (aPTT) and antifactor Xa (anti-Xa) activity are used to monitor unfractionated heparin therapy in children on extracorporeal membrane oxygenation (ECMO). Elevated C-reactive protein (CRP) can prolong aPTT and cause discrepancy between these two assays. We aimed to evaluate CRP effect on aPTT and anti-Xa assays in the presence of heparin and to determine whether elevated CRP affects laboratory monitoring in pediatric ECMO patients. Citrated normal specimens were spiked with CRP, heparin, and recombinant factor VIII (FVIII) and followed by measurement of aPTT and anti-Xa activity. Additionally, aPTT, anti-Xa activity, FVIII, fibrinogen, and CRP were measured in 18 ECMO specimens. Elevated CRP prolonged aPTT in normal specimens with or without heparin, but did not affect anti-Xa assay. In contrast, ECMO specimens showed similar aPTT and anti-Xa values regardless of CRP level. Elevated CRP in specimens was accompanied by increased fibrinogen and FVIII activity. Additional in vitro experiments confirmed that FVIII spiked simultaneously with CRP attenuated CRP-induced aPTT prolongation in heparinized specimens. In vitro CRP-induced aPTT prolongation is not observed in pediatric ECMO samples due to concomitant FVIII increase. Discordant changes of CRP and FVIII in plasma could contribute to aPTT/anti-Xa discrepancies observed during heparin therapy in the pediatric population. The anti-Xa assay is preferable for heparin monitoring in pediatric ECMO settings.

Determining the maximal storage time of centrifuged citrated samples for performing add-on routine coagulation tests

BackgroundSeveral routine coagulation tests have been developed to give insight in the coagulation pathway. The laboratory diagnostical process consists of 3 phases, the pre-analytical, analytical and post-analytical phase; however, the pre-analytical phase is most sensitive to errors. The amount of time blood is stored, can affect the measurements of these coagulation tests and result in an incorrect conclusion. Therefore, we performed experiments to determine the maximal storage time, centrifuged blood samples can be reliably measured. MethodsCitrated whole blood from hospital patients, who were tested for routine coagulation, was collected in 2.7 mL citrate tubes. These whole blood samples were centrifuged and the plasma was stored on top of the cell pellet at room temperature. After 2 h, 4 h, 6 h, 12 h and 24 h, the prothrombin time (PT), international normalized ratio (INR), activated partial thromboplastin time (aPTT), fibrinogen concentration, antithrombin activity, D-dimer concentration and thrombin time were measured using Sysmex CS2100 coagulation analysers. Results and conclusionAnalytical evaluation of routine coagulation tests resulted in various significant differences and large variations between the various time intervals. Our results indicated that the PT and INR can be measured up till 24 h of storage. Centrifuged blood for measuring the fibrinogen concentration, antithrombin activity, D-dimer concentration and thrombin time can be stored up to 4 h, while 2 h of storage might already be too long for obtaining reliable aPTT measurements.

Quantitative Systems Pharmacology Model-Based Predictions of Clinical Endpoints to Optimize Warfarin and Rivaroxaban Anti-Thrombosis Therapy.

BackgroundTight monitoring of efficacy and safety of anticoagulants such as warfarin is imperative to optimize the benefit-risk ratio of anticoagulants in patients. The standard tests used are measurements of prothrombin time (PT), usually expressed as international normalized ratio (INR), and activated partial thromboplastin time (aPTT).ObjectiveTo leverage a previously developed quantitative systems pharmacology (QSP) model of the human coagulation network to predict INR and aPTT for warfarin and rivaroxaban, respectively.MethodsA modeling and simulation approach was used to predict INR and aPTT measurements of patients receiving steady-state anticoagulation therapy. A previously developed QSP model was leveraged for the present analysis. The effect of genetic polymorphisms known to influence dose response of warfarin (CYP2C9, VKORC1) were implemented into the model by modifying warfarin clearance (CYP2C9 *1: 0.2 L/h; *2: 0.14 L/h, *3: 0.04 L/h) and the concentration of available vitamin K (VKORC1 GA: −22% from normal vitamin K concentration; AA: −44% from normal vitamin K concentration). Virtual patient populations were used to assess the ability of the model to accurately predict routine INR and aPTT measurements from patients under long-term anticoagulant therapy.ResultsThe introduced model accurately described the observed INR of patients receiving long-term warfarin treatment. The model was able to demonstrate the influence of genetic polymorphisms of CYP2C9 and VKORC1 on the INR measurements. Additionally, the model was successfully used to predict aPTT measurements for patients receiving long-term rivaroxaban therapy.ConclusionThe QSP model accurately predicted INR and aPTT measurements observed during routine therapeutic drug monitoring. This is an exemplar of how a QSP model can be adapted and used as a model-based precision dosing tool during clinical practice and drug development to predict efficacy and safety of anticoagulants to ultimately help optimize anti-thrombotic therapy.