Reduced circulating FABP2 in patients with moderate to severe COVID-19 may indicate enterocyte functional change rather than cell death.

The gut is of importance in the pathology of COVID-19 both as a route of infection, and gut dysfunction influencing the severity of disease. Systemic changes caused by SARS-CoV-2 gut infection include alterations in circulating levels of metabolites, nutrients and microbial products which alter immune and inflammatory responses. Circulating plasma markers for gut inflammation and damage such as zonulin, lipopolysaccharide and ß-glycan increase in plasma along with severity of disease. However, Intestinal Fatty Acid Binding Protein / Fatty Acid Binding Protein 2 (I-FABP/FABP2), a widely used biomarker for gut cell death, has paradoxically been shown to be reduced in moderate to severe COVID-19. We also found this pattern in a pilot cohort of mild (n = 18) and moderately severe (n = 19) COVID-19 patients in Milan from March to June 2020. These patients were part of the first phase of COVID-19 in Europe and were therefore all unvaccinated. After exclusion of outliers, patients with more severe vs milder disease showed reduced FABP2 levels (median [IQR]) (124 [368] vs. 274 [558] pg/mL, P < 0.01). A reduction in NMR measured plasma relative lipid-CH3 levels approached significance (median [IQR]) (0.081 [0.011] vs. 0.073 [0.024], P = 0.06). Changes in circulating lipid levels are another feature commonly observed in severe COVID-19 and a weak positive correlation was observed in the more severe group between reduced FABP2 and reduced relative lipid-CH3 and lipid-CH2 levels. FABP2 is a key regulator of enterocyte lipid import, a process which is inhibited by gut SARS-CoV-2 infection. We propose that the reduced circulating FABP2 in moderate to severe COVID-19 is a marker of infected enterocyte functional change rather than gut damage, which could also contribute to the development of hypolipidemia in patients with more severe disease.

Lombardy diagnostic and therapeutic network of thrombotic microangiopathy.

BACKGROUND: Thrombotic thrombocytopenic purpura (TTP) is a rare, life-threatening thrombotic microangiopathy (TMA) requiring urgent treatment. Standardization of its diagnosis and optimal management is challenging. This study aimed to evaluate the role of centralized, rapid testing of ADAMTS13 in patients experiencing acute TMAs requiring plasma-exchange (PEX) and to estimate the incidence of TTP in a large Italian Region. METHODS: We perfomed a cohort study in the frame of the project "Set-up of a Lombardy network for the study and treatment of patients undergoing apheresis", including 11 transfusion centers in the Region. Consecutive patients referred from 2014 to 2016 with acute TMAs requiring PEX were enrolled. Centralized ADAMTS13 activity testing was performed at the Milan Hemophilia and Thrombosis Center within 24 h. RESULTS: Forty-three TMA patients (44 events) were enrolled, of whom 35 (81%) had severe ADAMTS13 deficiency. Patients with severe ADAMTS13 deficiency were younger, mainly women, with a higher prevalence of autoimmune disorders and a lower prevalence of cancer. Clinical and laboratory characteristics of patients with and without severe ADAMTS13 deficiency largely overlapped, with a lower platelet count being the only baseline marker that significantly differed between the two patient groups (ADAMTS13 activity < 10% vs ≥ 10%: median difference of -27 × 109/l, 95% CI - 37 to - 3). PEX treatment was initiated in all patients, but soon discontinued in cases without severe ADAMTS13 deficiency. In this group, the mortality rate was higher and no episode exacerbations or relapses within 6 months occured. The estimated average annual incidence of acute acquired TTP events was 1.17 [0.78-1.55] per million people. CONCLUSIONS: Severe ADAMTS13 deficiency distinguished two groups of patients with largely overlapping clinical features but different treatment and disease course. This study provides a feasible model implemented in a large Italian region for the practical clinical approach to TMAs and underlines the importance of urgent ADAMTS13 activity testing for an accurate differential diagnosis and therapeutic approach.

Managing hematological cancer patients during the COVID-19 pandemic: an ESMO-EHA Interdisciplinary Expert Consensus.

BACKGROUND: The COVID-19 pandemic has created enormous challenges for the clinical management of patients with hematological malignancies (HMs), raising questions about the optimal care of this patient group. METHODS: This consensus manuscript aims at discussing clinical evidence and providing expert advice on statements related to the management of HMs in the COVID-19 pandemic. For this purpose, an international consortium was established including a steering committee, which prepared six working packages addressing significant clinical questions from the COVID-19 diagnosis, treatment, and mitigation strategies to specific HMs management in the pandemic. During a virtual consensus meeting, including global experts and lead by the European Society for Medical Oncology and the European Hematology Association, statements were discussed and voted upon. When a consensus could not be reached, the panel revised statements to develop consensual clinical guidance. RESULTS AND CONCLUSION: The expert panel agreed on 33 statements, reflecting a consensus, which will guide clinical decision making for patients with hematological neoplasms during the COVID-19 pandemic.

Evaluation of procoagulant imbalance in Cushing's syndrome after short- and long-term remission of disease.

OBJECTIVE: Patients with Cushing's syndrome (CS) are at high risk of venous thromboembolism related to a hypercoagulability due to procoagulant imbalance. However, whether these alterations are reversible after disease remission is still unclear. The endogenous thrombin potential (ETP) measured with and without the addition of thrombomodulin provides a global representation of coagulation and previous data confirmed hypercoagulable profile in patients with active hypercortisolism. Aim of this study was to assess the short- and long-term modification of ETP in patients with CS after disease remission. DESIGN AND METHODS: Nineteen patients with CS for whom surgical remission was achieved, were prospectively evaluated for clinical characteristics, cortisol secretion profile and ETP at different time points: (i) before surgical intervention; (ii) after 6 months and (iii) 5 years from the time of persistent remission. Nineteen healthy subjects matched for age and gender were also evaluated as control group. RESULTS: Before surgery, patients showed higher ETP-ratio (with/without thrombomodulin) than controls (0.62 ± 0.09-vs-0.56 ± 0.09, p = 0.034). No significant correlation between ETP-ratio and cortisol secretion was found. 6 months after remission, ETP-ratio was still significantly increased compared to controls (0.64 ± 0.09-vs-0.56 ± 0.09, p = 0.01), but was similar to baseline (0.64 ± 0.09-vs-0.62 ± 0.09, p = 0.87). At 5 years, ETP-ratio showed a significant decrease (0.55 ± 0.14-vs-0.62 ± 0.09, p = 0.02) and was comparable to controls (0.55 ± 0.14-vs-0.56 ± 0.09, p = 0.7). CONCLUSIONS: Plasma hypercoagulability detected in patients with active hypercortisolism persists at short-term evaluation and seems to be completely reversible after long-term remission of disease. These data, as part of a whole evaluation of thrombotic risk, can contribute to make appropriate therapeutic choice in these patients.

Evolution of Haemophilia Care in Europe: 10 years of the principles of care.

INTRODUCTION: The European principles of care in haemophilia marked their first decade in 2018. These guiding principles were the beginning of the European Haemophilia Consortium (EHC) review of countries' adherence to these principles in 2009, 2012, 2015 and 2018. The aim of this paper was to examine the implementation of the principles and how they have impacted the evolution of care in the last decade, as well as to identify remaining gaps and proposes future directions. METHODS: In 2018, the EHC distributed a survey to EHC national member organisations in English and Russian and encouraged them to discuss responses with local clinicians for accuracy. Data was also cross-referenced and validated for countries in earlier surveys using additional available resources. RESULTS: The 10-year-old European principles had a significant impact on the development of care for haemophilia and related bleeding disorders in Europe. They set objectives around which multi-stakeholder groups have established recommendations and specific steps for the progressive improvement of care for bleeding disorders. However, some have been promoted and implemented more than others. CONCLUSION: Monitoring adherence to, and impact of, the European Principles of Care significantly assists in tracking developments and highlighting gaps. Countries' inability to report consistent and coherent data remains a challenge and hinders both provision of treatment and care for patients as well as optimal national and European healthcare systems.

Phase 3 study of recombinant von Willebrand factor in patients with severe von Willebrand disease who are undergoing elective surgery.

Essentials Recombinant von Willebrand factor (rVWF) is effective in von Willebrand disease (VWD). A phase 3 study of rVWF, with/without recombinant factor VIII (rFVIII) before surgery in VWD. Overall rVWF's efficacy was rated excellent/good; rVWF was administered alone in most patients. rVWF was well-tolerated and hemostasis was achieved in patients with severe VWD undergoing surgery. SUMMARY: Background Recombinant von Willebrand factor (rVWF) has demonstrated efficacy for on-demand treatment of bleeding in severe von Willebrand disease (VWD), warranting evaluation in the surgical setting. Objectives This study (NCT02283268) evaluated the hemostatic efficacy/safety profile of rVWF, with/without recombinant factor VIII (rFVIII), in patients with severe VWD undergoing surgery. Patients/Methods Patients received rVWF 40-60 IU kg-1 , VWF ristocetin cofactor activity was measured 12-24 h before surgery. If endogenous FVIII activity (FVIII:C) target levels were achieved 3 h before surgery, rVWF was administered alone 1 h before surgery; rVWF was co-administered with rFVIII if target endogenous FVIII levels were not achieved. rVWF was infused postoperatively to maintain target trough levels. Overall and intraoperative hemostatic efficacy, the pharmacodynamics of rVWF administration and the incidence of adverse events (AEs) were assessed. Results All patients treated with rVWF for major (n = 10), minor (n = 4) and oral (n = 1) surgery had overall and intraoperative hemostatic efficacy ratings of excellent (73.3% and 86.7%) or good (26.7% and 13.3%). Most rVWF infusions (89.4%) were administered alone, resulting in hemostatically effective levels of endogenous FVIII within 6 h, which were sustained for 72-96 h; 70% (n = 7/10) of major surgeries were performed without rFVIII co-administration. Six patients reported 12 treatment-emergent AEs. Two patients each had one serious AE: diverticulitis (not treatment related) and deep vein thrombosis (sponsor-assessed as possibly treatment related). No severe allergic reactions or inhibitory antibodies were reported. Conclusions These data support the efficacy and safety profile of rVWF in patients with severe VWD undergoing elective surgery.

Hemostatic abnormalities in patients with Ehlers-Danlos syndrome.

Essentials Ehlers-Danlos Syndrome (EDS) is a rare heterogeneous group of inherited collagen disorders. A cohort of EDS patients was investigated for bleeding tendency and hemostatic abnormalities. EDS is associated with an increased risk of bleeding. EDS patients have platelet function abnormalities, whose severity correlates with bleeding risk. SUMMARY: Background Ehlers-Danlos syndrome (EDS) includes a heterogeneous group of connective tissue disorders affecting skin, bones, vessels, and other organs. Patients with EDS have an increased risk of bleeding, but a comprehensive study of hemostasis in EDS patients is lacking. Objective To investigate the bleeding tendency of a cohort of patients with EDS by using the Bleeding Assessment Tool of the ISTH, the bleeding severity score (BSS). Methods The BSS was defined as abnormal when it was ≥ 4 in men and ≥ 6 in women. Patients with a bleeding tendency were compared with those without in terms of type and number of hemostatic abnormalities. Results Fifty-nine of 141 patients with EDS (41.7%) had an abnormal BSS. Prothrombin time and activated partial thromboplastin time were slightly prolonged in 10 patients (7.1%) because of mild coagulation factor deficiencies, which were not responsible for the bleeding diathesis. von Willebrand factor antigen, ristocetin cofactor, endogenous thrombin potential and platelet count were normal in all patients. At least one platelet function abnormality was found in 53 patients (90%) with an abnormal BSS and in 64 (78%) with a normal BSS (adjusted odds ratio [OR] 2.55, 95% confidence interval [CI] 0.87-7.48). The risk of bleeding progressively increased with the number of platelet function abnormalities, reaching an OR of 5.19 (95% CI 1.32-20.45) when more than three abnormalities were detected. Conclusions Our results show that nearly half of patients with EDS have an abnormal BSS, which, in 90% of cases, appear, at least in part, to be attributable to platelet function abnormalities. Abnormalities of primary hemostasis may contribute to the risk of bleeding in patients with EDS.

Targeted sequencing to identify novel genetic risk factors for deep vein thrombosis: a study of 734 genes.

Essentials Deep vein thrombosis (DVT) has a large unknown genetic component. We sequenced coding areas of 734 hemostasis-related genes in 899 DVT patients and 599 controls. Variants in F5, FGA-FGG, CYP4V2-KLKB1-F11, and ABO were associated with DVT risk. Associations in KLKB1 and F5 suggest a more complex genetic architecture than previously thought. SUMMARY: Background Although several genetic risk factors for deep vein thrombosis (DVT) are known, almost all related to hemostasis, a large genetic component remains unexplained. Objectives To identify novel genetic determinants by using targeted DNA sequencing. Patients/Methods We included 899 DVT patients and 599 controls from three case-control studies (DVT-Milan, Multiple Environmental and Genetic Assessment of risk factors for venous thrombosis [MEGA], and the Thrombophilia, Hypercoagulability and Environmental Risks in Venous Thromboembolism [THE-VTE] study) for sequencing of the coding regions of 734 genes involved in hemostasis or related pathways. We performed single-variant association tests for common variants (minor allele frequency [MAF] ≥ 1%) and gene-based tests for rare variants (MAF ≤ 1%), accounting for multiple testing by use of the false discovery rate (FDR). Results Sixty-two of 3617 common variants were associated with DVT risk (FDR < 0.10). Most of these mapped to F5,ABO,FGA-FGG, and CYP4V2-KLKB1-F11. The lead variant at F5 was rs6672595 (odds ratio [OR] 1.58, 95% confidence interval [CI] 1.29-1.92), in moderate linkage with the known variant rs4524. Reciprocal conditional analyses suggested that intronic variation might drive this association. We also observed a secondary association at the F11 region: missense KLKB1 variant rs3733402 remained associated conditional on known variants rs2039614 and rs2289252 (OR 1.36, 95% CI 1.10-1.69). Two novel variant associations were observed, in CBS and MASP1, but these were not replicated in the meta-analysis data from the International Network against Thrombosis (INVENT) consortium. There was no support for a burden of rare variants contributing to DVT risk (FDR > 0.2). Conclusions We confirmed associations between DVT and common variants in F5,ABO,FGA-FGG, and CYP4V2-KLKB1-F11, and observed secondary signals in F5 and CYP4V2-KLKB1-F11 that warrant replication and fine-mapping in larger studies.

An international collaborative study to compare different von Willebrand factor glycoprotein Ib binding activity assays: the COMPASS-VWF study.

Essentials New VWF activity assays are increasingly used but information on their comparability is limited. This is an ISTH SSC-organized study (expert labs, 5 countries) to compare all available assays. VWF activity by six assays correlated well with each other. The new assays show improved characteristics - minor differences are noted. SUMMARY: Background Several new assays have become available to measure von Willebrand factor (VWF) activity. The new assays appear to have improved performance characteristics compared with the old reference standard, ristocetin cofactor activity (VWF:RCo), but information is limited about how they compare with VWF:RCo and each other. Methods The von Willebrand factor Subcommittee of the International Society for Thrombosis and Haemostasis (ISTH) Scientific and Standardization Committee (SSC) designed a collaborative study involving expert laboratories from several countries to compare available tests with each other and with VWF:RCo. Eight laboratories from five countries were provided with blinded samples from normal healthy individuals and well-characterized clinical cases. Laboratories measured VWF activity using all tests available to them; data from six laboratories, not affected by thawing during transportation, are included in this study. Results All tests correlated well with VWF:RCo activity (r-values ranged from 0.963 to 0.989). Slightly steeper regression lines for VWF:Ab and VWF:GPIbM were clinically insignificant. The new assays showed improved performance characteristics. Of the commercially available assays, the VWF:GPIbR using the AcuStar system was the most sensitive and could reliably detect VWF activity below 1 IU dL-1 . The lower limit of the measuring interval for the VWF:GPIbM and the VWF:GPIbR assays was in the 3-4 and 3-6 IU dL-1 range, respectively. Inter-laboratory variation was also improved for most new assays. Conclusion All VWF activity assays correlated well with each other and the VWF:RCo assay. The slight differences in characteristics found in the COMPASS-VWF study will assist the VWF community in interpreting and comparing activity results.

Recombinant factor XIII A-subunit in a patient with factor XIII deficiency and recurrent pregnancy loss.

Essentials Inherited factor XIII deficiency is a very rare bleeding disorder. We used recombinant factor XIII-A in a pregnant patient with factor XIII-A subunit deficiency. The patient had a successful pregnancy outcome with no pregnancy related complications. The dose of recombinant factor XIII-A was minimized by using frequent trough level monitoring. SUMMARY: Inherited factor XIII deficiency is a very rare bleeding disorder, and is one of the causes of recurrent pregnancy loss. The use of plasma-derived FXIII to improve pregnancy outcomes has been reported. We report a 26-year-old woman with FXIII A-subunit (FXIII-A) deficiency who was treated with recombinant FXIII-A and had a successful pregnancy outcome with no pregnancy-related complications. Our case illustrates that the dose of recombinant FXIII-A can be minimized and adjusted on the basis of frequent trough level monitoring.

"In vitro" correction of the severe factor V deficiency-related coagulopathy by a novel plasma-derived factor V concentrate.

INTRODUCTION: Severe congenital factor V (FV) deficiency is a rare bleeding disorder characterized by very low/undetectable levels of FV. Fresh frozen plasma is the standard treatment for bleeding manifestations. Recently, a novel plasma-derived FV concentrate has been developed. AIM: To evaluate the "in vitro" ability of the novel FV concentrate to normalize clotting times and generate normal amount of thrombin in plasma collected from patients with severe FV deficiency. METHODS: Prothrombin time (PT), activated partial thromboplastin time (aPTT), FV activity and antigen levels and thrombin generation were measured pre- and postspiking of plasma samples of 10 patients with increasing doses of FV concentrate (from 0 to 100 IU/dL). RESULTS: Prothrombin time and activated partial thromboplastin time ratios as well as all thrombin generation parameters were fully corrected by the addition of FV concentrate at a final concentration of 25 IU/dL. However, the addition of FV at a concentration of 1-3 IU/dL was already sufficient to correct peak height and endogenous thrombin potential (but not lag time and time to peak) after activation with 5 pmol/L tissue factor. FV activity and antigen levels showed a linear response to supplementation with the novel FV concentrate. CONCLUSION: The novel plasma-derived FV concentrate was effective to correct "in vitro" severe FV deficiency in patients. The optimal FV concentration to fully normalize both global clotting times and thrombin generation parameters using the novel plasma-derived FV concentrate was 25 IU/dL.

First-year results of an expanded humanitarian aid programme for haemophilia in resource-constrained countries.

INTRODUCTION: The gaps in haemophilia treatment around the world are enormous; approximately 60% of an estimated 475 000 individuals are not identified. Of the 187 000 diagnosed, 30% (57 000) access clotting factor replacement therapy. Since 1996, humanitarian aid distributed by the World Federation of Hemophilia (WFH) has played a minor, yet vital role providing life-saving clotting factor to countries in emergency situations. Donated amounts have been small and sporadic, often salvaging short-dated products, providing little opportunity to leverage donations with governments. In 2015, a prospective donation programme of 100 million I.U. per year of extended half-life factor VIII and IX over 10 years was established, necessitating the development of new logistics and training programmes by WFH. AIM: To measure the impact of a greatly expanded haemophilia humanitarian aid program. MATERIALS AND METHODS: In 2016, the first full year of the expanded programme, WFH, distributed products to 58 countries with factor VIII usage <1 I.U. per capita, a level incompatible with long-term survival and far below the 4 I.U. FVIII per capita minimum established in Europe. RESULTS: The scope of the programme and utilization data for 2016 indicate primarily use for acute bleeding, orthopaedic and emergency surgeries. Compared to 2014, 2016 data showed substantial increases in patients served (5.9-fold, from 2119 to 14 579), surgeries performed (37-fold) and bleeds treated (6.9-fold). Patients on prophylaxis rose from 0 to 852, including 458 children under 10 years old. DISCUSSION: The expanded humanitarian aid programme impacts an estimated 10% of individuals with haemophilia previously unable to access treatment. CONCLUSION: This programme represents an unprecedented public-private partnership to deliver medicines to individuals with no access. Further, the programme offers the prospective opportunity to engage governments to take more responsiblity for increasing training, medical management, and product supply in 58 resource constrained countries.

Prediction of factor VIII inhibitor development in the SIPPET cohort by mutational analysis and factor VIII antigen measurement.

Essentials A residual factor VIII synthesis is likely to be protective towards inhibitor (INH) development. Mutation type-inhibitor risk association was explored in 231 patients with severe hemophilia A. A 2-fold increase in INH development for in silico null vs. non-null mutations was found. A 3.5-fold increase in INH risk for antigen negative vs. antigen positive mutations was found. SUMMARY: Background The type of F8 mutation is the main predictor of inhibitor development in patients with severe hemophilia A. Mutations expected to allow residual synthesis of factor VIII are likely to play a protective role against alloantibody development by inducing immune tolerance. According to the expected full or partial impairment of FVIII synthesis, F8 variants are commonly classified as null and non-null. Objectives To explore the mutation type-inhibitor risk association in a cohort of 231 patients with severe hemophilia A enrolled in the Survey of Inhibitors in Plasma-Product Exposed Toddlers (SIPPET) randomized trial. Methods The genetic defects in these patients, consisting of inversions of intron 22 (n = 110) and intron 1 (n = 6), large deletions (n = 16), and nonsense (n = 38), frameshift (n = 28), missense (n = 19) and splicing (n = 14) variants, of which 34 have been previously unreported, were reclassified according to two additional criteria: the functional effects of missense and splicing alterations as predicted by multiple in silico analyses, and the levels of FVIII antigen in patient plasma. Results A two-fold increase in inhibitor development for in silico null mutations as compared with in silico non-null mutations (hazard ratio [HR] 2.08, 95% confidence interval [CI] 0.84-5.17) and a 3.5-fold increase in inhibitor development for antigen-negative mutations as compared with antigen-positive mutations (HR 3.61, 95% CI 0.89-14.74] were found. Conclusions Our findings confirm an association between the synthesis of minute amounts of FVIII and inhibitor protection, and underline the importance of investigating the residual FVIII antigen levels associated with causative variants in order to understand their clinical relevance.

The ISTH Bleeding Assessment Tool and the risk of future bleeding.

Essentials ISTH Bleeding Assessment Tool (ISTH-BAT) is used to assist the diagnosis of bleeding disorders. We examined whether the ISTH-BAT is capable of predicting the risk of future bleeding. 136 subjects were administered the ISTH-BAT and followed for up to four years. The ISTH-BAT score failed to predict the risk of future bleeding. SUMMARY: Background The ISTH Bleeding Assessment Tool (ISTH-BAT) is a diagnostic tool used in subjects with suspected inherited bleeding disorders. Aim To evaluate whether the ISTH-BAT, applied at first work-up in a tertiary-care center, predicts the risk of subsequent bleeding events. Methods This was an observational cohort study including all consecutive subjects, of either sex and any age, referred between 2011 and 2015 because of a suspected bleeding disorder. The analysis was restricted to those with an ISTH-BAT score of ≥ 3. Incidence rates (IRs) of major bleeding (MB) and clinically relevant non-major bleeding (CRNMB) events were calculated as the number of events over accrued person-years. The main analysis was performed with Cox regression analysis, assessing an ISTH-BAT score of ≤ 5 versus a score of > 5, as well as the score as a continuous variable, and various covariates (sex, age, and presence/absence of a final diagnosis). Results One hundred and thirty-six subjects had a median ISTH-BAT score of 4 (range 3-18). Eleven subjects (8.1%) had a bleeding event during follow-up (one MB event; 10 CRNMB events). The overall IR of bleeding events per 100 person-years was 3.7 (95% confidence interval [CI] 1.8-6.6). No difference was observed between subjects with an ISTH-BAT score of ≤ 5 and those with a score of > 5 (hazard ratio [HR] 1.2, 95% CI 0.3-4.6). The results were similar when the ISTH-BAT score was considered as a continuous variable (HR 1.1, 95% CI 0.9-1.4). The IR of bleeding was increased in individuals with a diagnosis of a hemostatic defect (IR of 7.5 per 100 person-years; HR 3.0, 95% CI 0.8-11.8). Conclusions The ISTH-BAT does not identify patients at increased risk of future bleeding events.