Antithrombin deficiency caused by SERPINC1 gene mutation in white matter lesions: A case report.

RATIONALE: White matter lesions (WMLs) are structural changes in the brain that manifest as demyelination in the central nervous system pathologically. Vasogenic WMLs are the most prevalent type, primarily associated with advanced age and cerebrovascular risk factors. Conversely, immunogenic WMLs, typified by multiple sclerosis (MS), are more frequently observed in younger patients. It is crucial to distinguish between these 2 etiologies. Furthermore, in cases where multiple individuals exhibit WMLs within 1 family, genetic testing may offer a significant diagnostic perspective. PATIENT CONCERNS: A 25-year-old male presented to the Department of Neurology with recurrent headaches. He was healthy previously and the neurological examination was negative. Brain magnetic resonance imaging (MRI) showed widespread white matter hyperintensity lesions surrounding the ventricles and subcortical regions on T2-weighted and T2 fluid-attenuated inversion recovery images, mimicking immunogenic disease-MS. DIAGNOSES: The patient was diagnosed with a patent foramen ovale, which could explain his headache syndrome. Genetic testing unveiled a previously unidentified missense mutation in the SERPINC1 gene in the patient and his father. The specific abnormal laboratory finding was a reduction in antithrombin III activity, and the decrease may serve as the underlying cause for the presence of multiple intracranial WMLs observed in both the patient and his father. INTERVENTIONS: The patient received percutaneous patent foramen ovale closure surgery and took antiplatelet drug recommended by cardiologists and was followed up for 1 month and 6 months after operation. OUTCOMES: While the lesions on MRI remain unchanging during follow-up, the patient reported a significant relief in headaches compared to the initial presentation. LESSONS: This case introduces a novel perspective on the etiology of cerebral WMLs, suggesting that hereditary antithrombin deficiency (ATD) could contribute to altered blood composition and may serve as an underlying cause in certain individuals with asymptomatic WMLs.

Clinical and Molecular Characterization of Nine Novel Antithrombin Mutations.

Antithrombin (AT) is the major plasma inhibitor of thrombin (FIIa) and activated factor X (FXa), and antithrombin deficiency (ATD) is one of the most severe thrombophilic disorders. In this study, we identified nine novel AT mutations and investigated their genotype-phenotype correlations. Clinical and laboratory data from patients were collected, and the nine mutant AT proteins (p.Arg14Lys, p.Cys32Tyr, p.Arg78Gly, p.Met121Arg, p.Leu245Pro, p.Leu270Argfs*14, p.Asn450Ile, p.Gly456delins_Ala_Thr and p.Pro461Thr) were expressed in HEK293 cells; then, Western blotting, N-Glycosidase F digestion, and ELISA were used to detect wild-type and mutant AT. RT-qPCR was performed to determine the expression of AT mRNA from the transfected cells. Functional studies (AT activity in the presence and in the absence of heparin and heparin-binding studies with the surface plasmon resonance method) were carried out. Mutations were also investigated by in silico methods. Type I ATD caused by altered protein synthesis (p.Cys32Tyr, p.Leu270Argfs*14, p.Asn450Ile) or secretion disorder (p.Met121Arg, p.Leu245Pro, p.Gly456delins_Ala_Thr) was proved in six mutants, while type II heparin-binding-site ATD (p.Arg78Gly) and pleiotropic-effect ATD (p.Pro461Thr) were suggested in two mutants. Finally, the pathogenic role of p.Arg14Lys was equivocal. We provided evidence to understand the pathogenic nature of novel SERPINC1 mutations through in vitro expression studies.

[Phenotypic and genetic analysis of a Chinese pedigree affected with Hereditary antithrombin deficiency due to a novel variant of SERPINC1 gene].

OBJECTIVE: To analyze the clinical phenotype and genetic characteristics of a Chinese pedigree affected with Hereditary antithrombin deficiency. METHODS: A pedigree diagnosed at the the Second Affiliated Hospital of Wenzhou Medical University, Yuying Children's Hospital in June, 2020 was selected as the study subject. Plasma prothrombin time (PT), activated partial thromboplastin time (APTT), fibrinogen (FIB), and thrombin time (TT) of the probands and their pedigree members were determined using a STA-R automatic coagulation analyzer. Antithrombin activity (AT: A) and antithrombin antigen (AT: Ag) in plasma were determined with chromogenic substrate and immunonephelometry assays. All exons and flanking sequences of the anticoagulant protein gene SERPINC1 were amplified by PCR and subjected to Sanger sequencing. Candidate variants were verified with bioinformatic tools (PolyPhen-2, SIFT, Mutation Taster and PYMOL) to explore their effect on the function and structural conformation of the protein. RESULTS: The probands (II-2, II-10), their brother (II-5) and sons (III-1, III-8) had shown normal PT, APTT, FIB, and TT, but significantly decreased AT: A and AT: Ag, with their levels being 34%, 57%, 56%, 48%, 53% and 13.51 mg/dL, 13.44 mg/dL, 18.39 mg/dL, 17.36 mg/dL, 17.71 mg/dL, respectively. The remaining pedigree members had normal values. Sanger sequencing revealed that the probands and all affected pedigree members had harbored a heterozygous c.851T>C (p.Met284Thr) missense variant in exon 5 of the SERPINC1 gene. Bioinformatic analysis and simulation suggested that the variant has resulted in alteration of hydrogen bonds at the c.851 position, which may affect the structure of the protein. Based on the guidelines from the American College of Medical Genetics and Genomics (ACMG), the variant was classified as pathogenic (PS1+PM1+PM5+PP1+PP4). CONCLUSION: The probands and other affected members were all diagnosed with type I hereditary AT deficiency, for which the c.851T>C (p.Met284Thr) variant of the SERPINC1 gene may be accountable.

Levels of procoagulant factors and peak thrombin generation in relation to dementia risk in older adults: The Cardiovascular Health Study.

INTRODUCTION: Markers of hemostasis such as procoagulant factors and peak thrombin generation are associated with cardiovascular outcomes, but their associations with dementia risk are unclear. We aimed to evaluate prospective associations of selected procoagulant factors and peak thrombin generation with dementia risk. METHODS: We measured levels of 7 hemostatic factors (fibrinogen, factor VII coagulant activity [FVIIc], activated factor VII [FVIIa], factor VIIa-antithrombin [FVIIa-AT], factor XI antigen [FXI], peak thrombin generation, and platelet count) among participants in the Cardiovascular Health Study, a cohort of older adults free of dementia in 1992/1993 (n = 3185). Dementia was adjudicated and classified by DSM-IV criteria through 1998/1999. Cox proportional hazards models estimated hazard ratios (HRs) for any dementia associated with 1-standard deviation (SD) differences, adjusting for sociodemographic and clinical factors and APOE genotype. Secondary analyses separately evaluated the risk of vascular dementia, Alzheimer's disease, and mixed dementia. RESULTS: At baseline, participants had a median age of 73 years. Over 5.4 years of follow-up, we identified 448 dementia cases. There was no evidence of linear associations between levels of these hemostatic factors with any dementia risk (HRs per 1-SD difference ranged from 1.0 to 1.1; 95 % confidence intervals included 1.0). Results of secondary analyses by dementia subtype were similar. CONCLUSIONS: In this prospective study, there was no strong evidence of linear associations between levels of fibrinogen, FVIIc, FVIIa, FVIIa-AT, FXI, peak thrombin generation, or platelet count with dementia risk. Despite their associations with cardiovascular disease, higher levels of these biomarkers measured among older adults may not reflect dementia risk.

Anticoagulant effects of protein C, protein S, and antithrombin levels on the protein C pathway in young children.

The protein C (PC) pathway involves physiological anticoagulant factors (PC, protein S [PS], and factor V) and performs major anticoagulant functions in adults. Variations in overall PC pathway function due to dynamic changes in PC and PS in early childhood are poorly understood. We aimed to evaluate the contributions of PC pathway function during early childhood by measuring changes in plasma thrombin generation (TG) after administration of the PC activator protac. We evaluated correlations between anticoagulant factors and percentage of protac-induced coagulation inhibition (PiCi%). Before protac addition, TG in newborns (n = 35), infants (n = 42), young children (n = 35), and adults (n = 20) were 525 ± 74, 720 ± 96, 785 ± 53, and 802 ± 64 mOD/min, and PiCi% were 42.1 ± 9.9, 69.8 ± 11.0, 82.9 ± 4.4, and 86.9 ± 3.4%, respectively. The distribution of PiCi% on the two axes of TG (with or without protac) changed continuously with age and differed from that of warfarin-treated plasma and adult PC- or PS-deficient plasma. PiCi% increased dynamically during infancy and correlated with PS levels in newborns and PC levels in young children. Addition of PC or fresh frozen plasma equivalent to approximately 25% PC to PC-deficient plasma improved PiCi%. This automatic measurement requires only a small sample volume and is useful for analysis of developmental hemostasis.

Exploring antithrombin: insights into its physiological features, clinical implications and analytical techniques.

Antithrombin is an essential protein that acts as a natural anticoagulant in the human body. It is synthesized by the liver and belongs to the serine protease inhibitors, which are commonly referred to as the SERPINS superfamily. The antithrombin molecule comprises 432 amino acids and has a molecular weight of approximately 58 200 D. It consists of three domains, including an amino-terminal domain, a carbohydrate-rich domain, and a carboxyl-terminal domain. The amino-terminal domain binds with heparin, whereas the carboxyl-terminal domain binds with serine protease. Antithrombin is a crucial natural anticoagulant that contributes approximately 60-80% of plasma anticoagulant activities in the human body. Moreover, antithrombin has anti-inflammatory effects that can be divided into coagulation-dependent and coagulation-independent effects. Furthermore, it exhibits antitumor activity and possesses a broad range of antiviral properties. Inherited type I antithrombin deficiency is a quantitative disorder that is characterized by low antithrombin activity due to low plasma levels. On the other hand, inherited type II antithrombin deficiency is a qualitative disorder that is characterized by defects in the antithrombin molecule. Acquired antithrombin deficiencies are more common than hereditary deficiencies and are associated with various clinical conditions due to reduced synthesis, increased loss, or enhanced consumption. The purpose of this review was to provide an update on the structure, functions, clinical implications, and methods of detection of antithrombin.

Discovering peptide inhibitors of thrombin as a strategy for anticoagulation.

Unusual blood clots can cause serious health problems, such as lung embolism, stroke, and heart attack. Inhibiting thrombin activity was adopted as an effective strategy for preventing blood clots. In this study, we explored computational-based method for designing peptide inhibitors of human thrombin therapeutic peptides to prevent platelet aggregation. The random peptides and their 3-dimentional structures were generated to build a virtual peptide library. The generated peptides were docked into the binding pocket of human thrombin. The designed strong binding peptides were aligned with the native binder by comparative study, and we showed the top 5 peptide binders display strong binding affinity against human thrombin. The 5 peptides were synthesized and validated their inhibitory activity. Our result showed the 5-mer peptide AEGYA, EVVNQ, and FASRW with inhibitory activity against thrombin, range from 0.53 to 4.35 µM. In vitro anti-platelet aggregation assay was carried out, suggesting the 3 peptides can inhibit the platelet aggregation induced by thrombin. This study showed computer-aided peptide inhibitor design can be a robust method for finding potential binders for thrombin, which provided solutions for anticoagulation.

Investigation of real-world heparin resistance and anticoagulation management prior to cardiopulmonary bypass: report from a nationwide survey by the Japanese Association for Thoracic Surgery heparin resistance working group.

OBJECTIVE: Heparin resistance is often encountered during cardiopulmonary bypass. Heparin dose and activated clotting time target values for the initiation of cardiopulmonary bypass are not yet universally standardized; further no consensus exists on the management of heparin resistance. This study aimed to investigate the current real-world practice on heparin management and anticoagulant treatment for heparin resistance in Japan. METHODS: A questionnaire survey was conducted at medical institutions nationwide with which The Japanese Society of Extra-Corporeal Technology in Medicine members are affiliated, targeting surgical cases with cardiopulmonary bypass performed from January 2019 through December 2019. RESULTS: Among 69% (230/332) of the participating institutions, the criterion for heparin resistance was defined as "the target activated clotting time value not reached even with an additional dose of heparin administration". Cases of heparin resistance were reported in 89.8% (202/225) of the responded institutions. Of note, 75% (106/141) of the responded institutions reported heparin resistance associated with antithrombin activity ≥ 80%. Antithrombin concentrate was used in 38.4% (238/619 responses) or third dose of heparin in 37.8% (234/619 responses) for advanced heparin resistance treatment. Antithrombin concentrate was found to be effective in resolving heparin resistance in patients having normal, as well as lower antithrombin activity. CONCLUSION: Heparin resistance has occurred in many cardiovascular centers, even among patients with normal antithrombin activities. Interestingly, the administration of antithrombin concentrate resolved heparin resistance, regardless of the baseline antithrombin activity value.

The interplay between tamoxifen and endoxifen plasma concentrations and coagulation parameters in patients with primary breast cancer.

BACKGROUND: Tamoxifen is an effective treatment for primary breast cancer but increases the risk for venous thromboembolism. Tamoxifen decreases anticoagulant proteins, including antithrombin (AT), protein C (PC) and tissue factor (TF) pathway inhibitor, and enhances thrombin generation (TG). However, the relation between plasma concentrations of both tamoxifen and its active metabolite endoxifen and coagulation remains unknown. METHODS: Tamoxifen and endoxifen were measured in 141 patients from the prospective open-label intervention TOTAM-study after 3 months (m) and 6 m of tamoxifen treatment. Levels of AT and PC, the procoagulant TF, and TG parameters were determined at both timepoints if samples were available (n = 53-135 per analysis). Levels of coagulation proteins and TG parameters were correlated and compared between: 1) quartiles of tamoxifen and endoxifen levels, and 2) 3 m and 6 m of treatment. RESULTS: At 3 m, levels of AT, PC, TF and TG parameters were not associated with tamoxifen nor endoxifen levels. At 6 m, median TF levels were lower in patients in the 3rd (56.6 [33] pg/mL), and 4th (50.1 [19] pg/mL) endoxifen quartiles compared to the 1st (lowest) quartile (76 [69] pg/mL) (P=0.027 and P=0.018, respectively), but no differences in anticoagulant proteins or TG parameters were observed. An increase in circulating TF levels (3 m: 46.0 [15] versus 6 m: 54.4 [39] pg/mL, P < 0.001) and TG parameters was observed at the 6 m treatment timepoint, while AT and PC levels remained stable. CONCLUSIONS: Our results indicate that higher tamoxifen and endoxifen levels are not correlated with an increased procoagulant state, suggesting tamoxifen dose escalation does not further promote hypercoagulability.

Increased contact activated endogenous thrombin potential in pregnant women with preeclampsia.

Preeclampsia is a worldwide contributor to maternal and fetal morbidity and mortality. Women with preeclampsia are in a hyper-coagulable state with increased risk of thromboembolic disease later in life compared with normal pregnant women. The contact system (CAS) in plasma can mediate thrombin generation and is an important contributor to thrombus growth, but the activation of CAS during pregnancy complicated by preeclampsia is not yet elucidated, and CAS may play a role in the pathophysiology of preeclampsia. Therefore, the aim of the study is to address thrombin generation, and in particular, the capacity of the CAS-mediated pathway in patients with preeclampsia compared with pregnant controls. One hundred and seventeen women with preeclampsia and matched controls were included. The project was registered at www.clinicaltrials.gov as NCT04825145. CAS and tissue factor induced thrombin generation, proteins C and S, antithrombin, and histidine-rich glycoprotein (HRG) were assessed. Women with preeclampsia had significantly increased CAS and tissue factor-induced endogenous thrombin potential (ETP), and HRG compared with controls, P  = 0.022, P  = 0.024, and P  = 0.02, respectively. The concentrations of protein C and antithrombin were significantly reduced in the preeclampsia group, P  = 0.024 and P  < 0.0001, respectively. No significant difference in the concentration of protein S was detected, P  = 0.06. This study demonstrates a significant increased CAS-induced ETP and an overall decrease of important regulators of coagulation in women with preeclampsia compared with controls. These aspects can contribute to the hyper-coagulable state characterizing preeclampsia.

Argatroban Use in Pediatric Patients Supported by Paracorporeal Ventricular Assist Devices.

Direct thrombin inhibitor (DTI) use has been associated with decreased stroke and death rates in children on ventricular assist devices (VADs). Most information about DTI use for children on VADs has focused on bivalirudin with limited data on argatroban. We hypothesized that, compared to unfractionated heparin (UFH), argatroban would be associated with decreased bleeding, stroke, and death rates in children on VADs. We retrospectively collected data from patients <18 years old on paracorporeal VADs at Children's Wisconsin between January 1, 2010 and July 1, 2021. We divided patients into cohorts based on anticoagulation strategy with heparin or argatroban. Definitions of bleeding and neurologic events were the same as in other published reports on this population. We compared categorical variables with the χ 2 or Fisher's exact test, and continuous variables with the Mann-Whitney U test. Nineteen children were anticoagulated with argatroban, and 16 with heparin. Demographics between groups were not significantly different. Stroke, bleeding, and death rates did not differ between patients treated with UFH versus argatroban. The study population was complex with a high rate of extracorporeal membrane oxygenation (ECMO) use before VAD support, which likely impacted our findings. Our study does not support argatroban as a superior alternative anticoagulant compared to UFH in children requiring VADs.

Comparisons between diluted thrombin time, ecarin chromogenic assays, and UPLC-MS for plasma level dabigatran quantification: Results from DRIVING study.

INTRODUCTION: The knowledge of dabigatran levels is helpful for decision-making in specific situations such as urgent surgery or when the question of reversal arises (uncontrolled bleeding, eligibility for thrombolysis). However, a limited number of observational studies are available regarding comparisons between quantification methods. The objective of the study was to compare dabigatran plasma levels using three assays including the reference method (high-performance liquid chromatography coupled with mass spectrometry), focusing on the agreement around the 30-50 ng/mL clinically relevant thresholds. METHODS: Sixty healthy volunteers from DRIVING trial (NCT01627665) were given a single 300-mg dabigatran etexilate dose. Serial blood samplings were performed at pre-defined time points (0 to 24 h). We analyzed plasma samples using ultra-performance-liquid chromatography coupled with tandem mass spectrometry (UPLC-MS) (dabigatran reference method); ii/diluted thrombin time (dTT) (Hemoclot-DTI-Hyphen-Biomed); iii/ecarin-based chromogenic assay (ECA-II-Stago). RESULTS: Nine hundred sixty samples were analyzed using the three assays (2759 values). dTT and ECA-II values were highly correlated with those of UPLC-MS (Deming regression). Most values >50 ng/mL were higher using dTT and ECA-II compared to UPLC-MS: biases were constant, +14% and +16% with dTT and ECA-II, respectively (Bland-Altman plots), suggesting that active metabolites accounted for ~15% of thrombin inhibition. Regarding values <30 ng/mL, 30-50 ng/mL, or ≥50 ng/mL, the agreement probability between dTT and ECA-II was of 90.6% [88.4-92.5] (Cohen's kappa coefficient 0.84). CONCLUSION: dTT and ECA-II assays rapidly provide accurate dabigatran-level results for clinical practice, both assays being suitable in emergency, taking into account the thrombin inhibitory effect of dabigatran metabolites.

Anti-thrombotic Effects Mediated by a Novel Dual-Target Peptide Inhibiting Both Platelet Aggregation and Thrombin Activity without Causing Bleeding.

BACKGROUND: Classical anticoagulants and antiplatelets are associated with high frequencies of bleeding complications or treatment failure when used as single agents. Thrombin plays an important role in the blood coagulation system. GP IIb/IIIa is the central receptor of platelets, which can recognize the Arg-Gly-Asp (RGD) sequence and activate platelets. MATERIAL AND METHODS: Molecular simulation and homology modeling were performed to design a novel dual-target anticoagulant short peptide (PTIP ). The activities of PTIP on coagulation and platelet in vitro were analyzed. The antithrombotic activity of PTIP was determined by pulmonary thromboembolism model, ferric chloride injury model and arteriovenous bypass thrombosis model. Bleeding effect and toxicity of PTIP were evaluated. RESULTS: We have constructed a novel dual-target peptide (PTIP) based on the direct thrombin inhibitor peptide (DTIP). PTIP was expressed at high levels in Pichia pastoris. PTIP interfered with thrombin-mediated coagulation and ADP-induced platelet aggregation in vitro. When injected intravenously or subcutaneously, PTIP showed potent and dose-dependent extension of aPTT and PT which were similar to DTIP; but only PTIP was capable of inhibiting platelet aggregation. PTIP (1.0 mg/kg) decelerated thrombosis formation in venous and arterial vessels induced by FeCl3 injury. PTIP (1.0 mg/kg) also prevented deep venous thrombosis and increased the survival rate associated with pulmonary thromboembolism. And PTIP effectively reduced thrombus length in arteriovenous bypass thrombosis model. Moreover, the antithrombotic dose of PTIP could not induce bleeding. CONCLUSION: These data establish that PTIP represents a novel antithrombotic agent whose effects involve both inhibition of platelet activation and reduction of fibrin generation. And PTIP not only can be used in venous thrombosis and arterial thrombosis, it can also replace the combined treatment of antiplatelet and anticoagulant drugs in thrombotic diseases.

Antithrombin activity and central venous catheter-associated thrombosis in critically ill children at high risk of bleeding.

BACKGROUND: Normalization of antithrombin activity may prevent catheter-associated thrombosis in critically ill children at high risk of bleeding. OBJECTIVES: To characterize the temporal pattern of antithrombin activity, assess its association with catheter-associated thrombosis and clinically relevant bleeding, and evaluate its relationship with thrombin generation in these children. METHODS: In this prospective cohort study, critically ill children <18 years old at high risk of bleeding with central venous catheter were eligible. Antithrombin activity and thrombin generation were measured from platelet-poor plasma and after in vitro antithrombin supplementation. Systematic surveillance ultrasound was performed to diagnose thrombosis. Children were followed for bleeding. RESULTS: We enrolled 8 infants (median age: 0.2 years, IQR: 0.2, 0.3 years) and 72 older children (median age: 14.3 years, IQR: 9.1, 16.1 years). Mean antithrombin on the day of catheter insertion was 64 IU/dL (SD: 32 IU/dL) in infants and 83 IU/dL (SD: 35 IU/dL) in older children. Antithrombin normalized by the day of catheter removal. Thrombosis developed in 27 children, while 31 children bled. Thrombosis (regression coefficient: 0.008, 95% CI: -0.01, 0.03) and bleeding (regression coefficient: -0.0007, 95% CI: -0.02, 0.02) were not associated with antithrombin. Antithrombin was not correlated with in vivo change in endogenous thrombin potential (correlation coefficient: -0.07, 95% CI: -0.21, 0.08). In vitro supplementation reduced endogenous thrombin potential (correlation coefficient: -0.78; 95% CI: -0.95, -0.23). CONCLUSION: These findings may not support normalization of antithrombin activity to prevent catheter-associated thrombosis in critically ill children at high risk of bleeding.

Monitoring of Argatroban in Critically Ill Patients: A Prospective Study Comparing Activated Partial Thromboplastin Time, Point-of-Care Viscoelastic Testing with Ecarin Clotting Time and Diluted Thrombin Time to Mass Spectrometry.

BACKGROUND: The direct thrombin inhibitor argatroban is indicated for the treatment of heparin-induced thrombocytopenia II, but it is also used off-label to treat critically ill patients presenting with heparin resistance, severe antithrombin deficiency, or hypercoagulability. Direct drug monitoring is not routinely available, and argatroban dosing is mainly based on global coagulation assays such as activated partial thromboplastin time (PTT) or diluted thrombin time (TT), both of which have limitations in patients with hypercoagulability. METHODS: Blood samples were obtained from critically ill patients treated with argatroban. Activated PTT and diluted TT were measured with a STA R Max3 analyzer (STAGO Deutschland GmbH, Germany) using an argatroban-calibrated kit. Ecarin clotting time was measured using a point-of-care viscoelastic test device. Liquid chromatography with tandem mass spectrometry was performed using a reversed-phase column, a solvent gradient, and an API4000 mass spectrometer with electrospray. Correlation was described using Pearson correlation coefficient r and Bayesian multilevel regression to estimate relationships between outcomes and covariates. RESULTS: From June 2021 to March 2022, 205 blood samples from 22 patients were analyzed, allowing for 195 activated PTT-liquid chromatography with tandem mass spectrometry comparisons, 153 ecarin clotting time-liquid chromatography with tandem mass spectrometry comparison, and 105 diluted TT-liquid chromatography with tandem mass spectrometry comparisons. Compared to liquid chromatography with tandem mass spectrometry, performance of argatroban quantification was best for diluted TT (r = 0.91), followed by ecarin clotting time (r = 0.58) and activated PTT (r = 0.48). Regression analysis revealed that patients with sepsis were more prone to argatroban overdosing (coefficient, 4.194; 95% credible interval, 2.220 to 6.792). CONCLUSIONS: Although activated PTT monitoring of argatroban is the most commonly used test, in critically ill patients, diluted TT provides more precise measurements. Alternately, point-of-care viscoelastic ecarin clotting time also provides guidance for argatroban dosing to identify overdosing if available. The data also suggested that patients with sepsis are at greater risk for argatroban overdosing.

The clinical and genetic landscape of early-onset thrombophilia in Japan.

OBJECTIVES: To determine the optimal management for early-onset thrombophilia (EOT), the genetic and clinical features of protein C (PC)-, protein S (PS)-, or antithrombin (AT)-deficient patients of ≤20 years of age were studied in Japan. METHODS/RESULTS: Clinical and genetic information of all genetically diagnosed cases was collected through the prospective, retrospective study, and literature review. One-hundred-one patients had PC (n = 55), PS (n = 29), or AT deficiency (n = 18). One overlapping case had PC- and PS-monoallelic variant. Fifty-five PC-deficient patients (54%) had 26 monoallelic or 29 biallelic variant(s), and 29 (29%) PS-deficient patients had 20 monoallelic or nine biallelic variant(s). None of the patients had AT-biallelic variants. The frequent low-risk allele p.K193del (PC-Tottori) was found in five patients with monoallelic (19%) but not 29 with biallelic variant(s). The most common low-risk allele p.K196E (PS-Tokushima) was found in five with monoallelic (25%) and six with biallelic variant(s) (67%). One exceptional de novo PC variant was found in 32 families with EOT. Only five parents had a history of thromboembolism. Thrombosis concurrently developed in three mother-newborn pairs (two PC deficiency and one AT deficiency). The prospective cohort revealed the outcomes of 35 patients: three deaths with PC deficiency and 20 complication-free survivors. Neurological complications were more frequently found in patients with PC-biallelic variants than those with PC-, PS-, or AT-monoallelic variants (73% vs. 24%, p = .019). CONCLUSIONS: We demonstrate the need for elective screening for EOT targeting PC deficiency in Japan. Early prenatal diagnosis of PC deficiency in mother-infant pairs may prevent perinatal thrombosis in them.

Emicizumab-mediated hemostatic function assessed by thrombin generation assay in an in vitro model of factor VIII-depleted thrombophilia plasma.

Patients with hemophilia A (PwHA) may have concurrent deficiency of representative anticoagulant proteins, protein (P)C, PS, and antithrombin (AT), which reduces bleeding frequency. However, emicizumab-driven hemostasis in PwHA with such thrombophilic potential remains unclarified. This study investigated the influence of natural anticoagulants on emicizumab-driven coagulation in HA model plasma. Various concentrations of PS and AT were added to PS-deficient plasma and AT-deficient plasma in the presence of anti-FVIII antibody (FVIIIAb; 10BU/mL). PC-deficient plasma was mixed with normal plasma at various concentrations in the presence of FVIIIAb. Emicizumab (50 µg/mL) was added to these thrombophilic HA model plasmas, prior to tissue factor/ellagic acid-triggered thrombin generation assays. Co-presence of emicizumab increased peak thrombin values (PeakTh) dependent on PS, AT, and PC concentrations. Maximum coagulation potentials in the PS-reduced HA model plasmas remained normal in the presence of emicizumab. PeakTh were close to normal in the presence of 50%AT irrespective of emicizumab, but were higher than normal in the presence of 25%AT. Addition of recombinant FVIIa (corresponding to an administered dose of 90 µg/kg) enhanced coagulation potential to normal levels. Our findings provide novel information on hemostatic regulation in emicizumab-treated PwHA with a possible thrombophilic disposition.

Direct thrombin inhibitors fail to reverse the negative effects of heparin on lung growth and function after murine left pneumonectomy.

Neonates with congenital diaphragmatic hernia (CDH) frequently require cardiopulmonary bypass and systemic anticoagulation. We previously demonstrated that even subtherapeutic heparin impairs lung growth and function in a murine model of compensatory lung growth (CLG). The direct thrombin inhibitors (DTIs) bivalirudin and argatroban preserved growth in this model. Although DTIs are increasingly used for systemic anticoagulation clinically, patients with CDH may still receive heparin. In this experiment, lung endothelial cell proliferation was assessed following treatment with heparin-alone or mixed with increasing concentrations of bivalirudin or argatroban. The effects of subtherapeutic heparin with or without DTIs in the CLG model were also investigated. C57BL/6J mice underwent left pneumonectomy and subcutaneous implantation of osmotic pumps. Pumps were preloaded with normal saline, bivalirudin, or argatroban; treated animals received daily intraperitoneal low-dose heparin. In vitro, heparin-alone decreased endothelial cell proliferation and increased apoptosis. The effect of heparin on proliferation, but not apoptosis, was reversed by the addition of bivalirudin and argatroban. In vivo, low-dose heparin decreased lung volume compared with saline-treated controls. All three groups that received heparin demonstrated decreased lung function on pulmonary function testing and impaired exercise performance on treadmill tolerance testing. These findings correlated with decreases in alveolarization, vascularization, angiogenic signaling, and gene expression in the heparin-exposed groups. Together, these data suggest that bivalirudin and argatroban fail to reverse the inhibitory effects of subtherapeutic heparin on lung growth and function. Clinical studies on the impact of low-dose heparin with DTIs on CDH outcomes are warranted.NEW & NOTEWORTHY Infants with pulmonary hypoplasia frequently require cardiopulmonary bypass and systemic anticoagulation. We investigate the effects of simultaneous exposure to heparin and direct thrombin inhibitors (DTIs) on lung growth and pulmonary function in a murine model of compensatory lung growth (CGL). Our data suggest that DTIs fail to reverse the inhibitory effects of subtherapeutic heparin on lung growth and function. Clinical studies on the impact of heparin with DTIs on clinical outcomes are thus warranted.

Prognostic Value of Antithrombin Activity Levels in the Early Phase of Intensive Care: A 2-Center Retrospective Cohort Study.

To investigate the relationship between antithrombin (AT) activity level and prognosis in patients requiring intensive care. Patients whose AT activity was measured within 24 h of intensive care unit (ICU) admission were enrolled for analysis. The primary endpoint was mortality at discharge. Prognostic accuracy was examined using receiver operating characteristic (ROC) curves and cox hazard regression analysis. Patients were divided into 6 groups based on predicted mortality, and a χ2 independence test was performed on the prognostic value of AT activity for each predicted mortality; P < .05 was considered significant. A total of 281 cases were analyzed. AT activity was associated with mortality at discharge (AT% [interquartile range, IQR]): survivor group, 69 (56-86) versus nonsurvivor group, 56 (44-73), P = .0003). We found an increasing risk for mortality in both the lowest level of AT activity (<50%; hazard ratio [HR] 2.43, 95% confidence interval [CI] 1.20-4.89, P = .01) and the middle-low level of AT activity (≥ 50% and < 70%; HR 2.06, 95% CI 1.06-4.02, P = .03), compared with the normal AT activity level (≥ 70%). ROC curve analysis showed that the prediction accuracy of AT was an area under the curve (AUC) of 0.66 (cutoff 58%, sensitivity 61.4%, specificity 68.2%, P = .0003). AT activity was significantly prognostic in the group with 20% to 50% predicted mortality (AUC 0.74, sensitivity: 24.0%-55.5%, specificity: 83.3%-93.0%). An early decrease in AT activity level in ICU patients may be a predictor of mortality at discharge.