Inhibitors In Hemophilia Research Articles

Minor surgical procedures during immune tolerance induction in people with hemophilia A and inhibitors: results from the Brazilian Immune Tolerance (BrazIT) study cohort

IntroductionSurgeries are implicated in the development of anti-factor VIII (FVIII) neutralizing antibodies (inhibitors) in hemophilia A individuals with immune tolerance induction (ITI) treatment being the recommended therapy to eradicate these inhibitors. We evaluated the association of surgical procedures performed during ITI and treatment outcome. MethodsPatients were treated according to the Brazilian ITI Protocol with outcomes being defined as successful (i.e., recovered responsiveness to exogenous FVIII) and failed (i.e., unresponsiveness to exogenous FVIII thus requiring bypassing agents for bleeding control). Surgical procedures during induction therapy were managed following international recommendations. ResultsTreatment success rate was 68.7 % in 163 patients; 33 (20.2 %) were submitted to 43 (96 %) minor and two major surgeries. Personal, hemophilia, inhibitor, and treatment characteristics were similar between patients submitted to surgical procedures or not while on ITI; the success rates were 72.7 % and 67.7 % (p-value = 0.577), respectively. ConclusionNo association was found between having a minor surgical procedure and ITI treatment outcome.

HOMOEOPATHIC MANAGEMENT OF TARGETED HAEMARTHROSES IN INHIBITOR POSITIVE HAEMOPHILIAC: A CASE REPORT

Haemarthroses is common presentation in haemophilia patient. Due to recurrent use of clotting factor concentrates for treatment and prophylaxis, of which about 20 % of patients produce antibodies to factor VIII and factor IX. This results in development of inhibitors in such patients which complicates the case even more. Homoeopathy the alternative form of treatment can be used in such case. This case report presents the management of haemarthroses in a haemophilia inhibitor. Well selected homoeopathic similimum helped in reversing the haemophilic arthropathy in left knee joint of the patient and even controlled the recurrence of the haemarthroses in target joint. The result was assessed using Hemophilia joint health assessment scale (HJHS). The result of before and after treatment was extremely significant with a p value of 0.0009 The individual curative response of the case was assessed using Modified naranjo criteria for homoeopathic case reporting, casual attribution (MONARCH).

Recent Advances in Genetic and Non-Genetic Factors Related to the Development of Hemophilia Inhibitors

Recent Advances in Genetic and Non-Genetic Factors Related to the Development of Hemophilia Inhibitors

Hemophilia A Research Program (HARP): A Novel Study Design for a Multicenter and Decentralized Prospective Observational Longitudinal Cohort Study in Hemophilia A

Background: Hemophilia A is a rare X-linked bleeding disorder caused by factor VIII (FVIII) deficiency. There is a compelling need in hemophilia to better understand the causes of bleeding in females and the factors underlying FVIII inhibitor development. To meet this need we are establishing a new NHLBI funded Hemophilia A Analytical Cohort Research Program (HARP) as part of a public-private partnership to support an Intergenerational Precision Medicine Program for the study of factor VIII (FVIII) immunogenicity in severe hemophilia A (Harpf8.org). The HARP objectives are: 1) Longitudinal assessment of bleeding in females with a severe hemophilia A genotype beginning in pregnancy; 2) Longitudinal assessment of anti-FVIII antibody formation in severe hemophilia A from birth through at least the first 2 years of life; 3) Studies of causes of bleeding in females with a severe hemophilia A genotype; 4) Studies of the immune response to FVIII in severe hemophilia A; 5) Development of a well-curated annotated data set of genetic, hemophilia, immune, environmental, psychosocial, and quality of life metrics; 6) Development of a multi-omics and immunologic biospecimen resource; and 7) Development of the hemophilia analytical Portal (F8portal.org). Study design: The HARP is a multicenter and decentralized prospective observational longitudinal cohort study beginning in pregnancy. It will follow 50 mother-baby pairs where the babies have severe hemophilia A and the mothers have a severe hemophilia A genotype. The private Clinical Consortium (CC) will oversee the study throughout the U.S., including sites that care for the mothers (up to 200 sites) and sites that care for the child (up to 70 sites). The CC will also conduct decentralized aspects of the study including collection of patient reported outcomes and local sample collection. The primary maternal endpoint is primary postpartum hemorrhage; the primary pediatric endpoint is rate of development of humoral immune response to FVIII and proportion that progress to clinical inhibitors. Mother and child secondary endpoints reflect the diverse research the HARP resource will support. Inclusion criteria for the pregnant person are: Pregnant and prior to 37 weeks gestation, known or at-risk to have a severe hemophilia A genotype, pregnant with a fetus at-risk for severe hemophilia A, ability to give informed consent, and > 18 years old. Inclusion criteria for the child are: severe hemophilia A, born to a person in the study. Exclusion/discontinuation criteria are: Presence of another significant bleeding disorder in mother or child; mother age <18 years; no fetus is at-risk to inherit severe hemophilia A; the child does not have severe hemophilia A; the child has a severe immune disorder; the mother or child participates in another study with blood collection volumes that exceed safety limits; mother's or child's healthcare will be outside the U.S.; the mother is a prisoner; any other reason that in the opinion of the investigator a subject is unsuitable for the study Summary: The HARP innovates in multiple areas of clinical study design to conduct a national prospective longitudinal intergenerational study in a rare disorder beginning in pregnancy. The structure includes a new multidisciplinary CC, systematic inclusion of Lived Experience Experts, strategies for broad community engagement, recruitment, and retention, and decentralized operations. This cohort will enable deep characterization, including genomics and other -omics, on pregnancy-associated bleeding and drivers of FVIII inhibitors in hemophilia A. The HARP Shareable Resource will be comprised of well annotated data and biological samples accessible to researchers through a portal, F8portal.org, and a linked biorepository.

FVIII Inhibitors can Accurately be Measured in the Presence of Mim8 in Hemophilia A Plasma

Introduction: Mim8 (Denecimig; Novo Nordisk) is a next generation bispecific antibody being developed for prophylactic treatment of hemophilia A with or without inhibitors. Mim8 complexes with FIXa and FX to generate FXa in the absence of FVIII. There is a clinical need to monitor FVIII inhibitors in hemophilia A patients, however measuring FVIII inhibitors in the laboratory can be challenging. Using aPTT-based Modified Nijmegen-Bethesda Assays in the presence of FVIIIa Mimetics such as Mim8 is not recommended due to interference. While FVIII chromogenic substrate assays (CSA) containing human or human/bovine proteins vary in their sensitivity to FVIII Mimetics. This study investigated if bovine FIXa/FX FVIII CSA can accurately measure FVIII inhibitors in the presence of Mim8 in hemophilia A plasma. Methods: Two FVIII CSA kits using bovine FIXa/bovine FX reagents, Factor VIII Chromogenic Assay (Siemens Healthineers) and Chromogenix Coatest ® SP4 FVIII (Diapharma Group, Inc.) were evaluated to investigate whether Mim8 at 5, 10, 20 and 40 ug/mL (final concentration) interferes with the measurement of FVIII inhibitor (Affinity BioLogicals, Inc.) spiked into pooled congenital FVIII deficient plasma (George-King Bio-Medical, Inc.), at 0.2, 1.0 and 4.8 Bethesda Units/mL (BU/mL, final concentration). CSA were analyzed on BCS ®XP analyzers (Siemens Healthineers) using manufacturer or laboratory developed protocols. Inhibitor samples were prepared from a FVIII inhibitor working solution that was assigned a potency of 11.8 BU/mL (Siemens CSA). Mim8 was added to the FVIII inhibitor samples at 20x the final concentration so that the plasma matrix was 95% inhibitor plasma. CRYOcheck™ Factor VIII Inhibitor kit (Precision BioLogic, Inc.) was used to measure the FVIII inhibitor levels. Plasma containing FVIII inhibitor alone or in combination with Mim8 was heat inactivated at 56 ± 2°C for 30 minutes, followed by centrifugation at 2700xg for 5 minutes. The inactivated plasma was incubated at 37 ± 1°C for 2 hours with Imidazole buffered Pooled Normal Plasma (PNP). By standardizing the amount of FVIII in the PNP (⁓100%), the inhibitor levels were determined according to how much of the added FVIII was inactivated relative to a control tube mix with no inhibitor present. Mim8 interference was defined as a negative result (<0.6 BU/mL) becoming positive for samples containing 0.2 BU/mL of FVIII inhibitor, and a result outside of ±25% (%RE) of the corresponding un-spiked sample for the samples containing 1.0 or 4.8 BU/mL of FVIII inhibitor. Results: The data presented for each CSA is from 3 separate aliquots tested on 3 independent occasions. No Mim8 interference was observed at any FVIII inhibitor levels whichever CSA kit, using bovine FIXa or FX reagent, was used with all results for Mim8 spiked samples falling within ±25.0% of the target value. Using the Siemens CSA inhibitor samples at 0.2 BU/mL remained negative (<0.6 BU/mL) with no results reporting >0.4 BU/mL. Inhibitor samples with target values of 1.0 or 4.8 BU/mL, recovered 0.96, 0.99, 0.98 and 0.97 BU/mL and 5.66, 5.69, 5.66 and 5.62 BU/mL, as Mim8 concentrations increased, compared to the un-spiked samples at 1.01 and 5.74 BU/mL, respectively. The spiked samples at 1.0 and 4.8 BU/mL recovered within -2.0 to -5.0%, and -2.1 to -0.9%, of the un-spiked sample, whilst the un-spiked 1.0 and 4.8 BU/mL samples recovered within 1.0% and 19.6% of target, respectively. The Chromogenix CSA recovered 0.90, 0.95, 0.94 and 0.84, as Mim8 concentrations increased, compared to the un-spiked sample at 0.88 at 1.0 BU/mL. At 4.8 BU/mL the results were 5.22, 5.50, 5.57 and 5.72 BU/mL compared to the un-spiked sample of 5.07 BU/mL. The spiked sample at 1.0 BU/mL and 4.8 BU/mL recovered within -4.5 to 8.0%, and 3.0 to 12.8%, of the un-spiked sample. The un-spiked 1.0 and 4.8 BU/mL samples recovered within -12.0% and 5.6% of target, respectively. Conclusion: This study showed that by using FVIII chromogenic assays containing bovine reagents, FVIII inhibitors levels up to at least ⁓5.0 BU/mL can be accurately determined in severe hemophilia A plasma in the presence of up to 40 µg/mL Mim8.

Potential Predictive Biomarkers for the Development and Outcomes of Inhibitors in Hemophilia A Patients

Introduction Inhibitor development poses a significant challenge in the treatment of hemophilia A (HA). Approximately 30% of HA patients develop inhibitors in response to FVIII infusion therapy. While the risk is highest early in treatment days, patients are at risk throughout their lifetime. Regardless of certain known risk factors, the lack of biomarkers for inhibitor development hampers efforts to prevent their occurrence. Since inflammation is a well-known risk factor, one of our goals is to identify potential biomarkers among inflammatory cytokines. Another focus is glycosylation-related biomarkers, which has drawn great attention in biomarker discovery of various diseases including cancers. By analyzing the association of cytokines and plasma glycosylation with inhibitor formation, this research aims to provide valuable references for clinical care. Methods Plasma samples from 60 HA patients, and 23 healthy donors were analyzed to detect the levels of G-CSF, IFN-γ, IL-1β, IL-2, IL-6, IL-8, IL-10, IL12p70, IP-10 and TNF. The HA samples were divided into inhibitor negative (InhNeg, BU < 0.6, n = 26) and inhibitor positive (InhPos, BU ≥ 0.6, n = 34) groups. A predictive model of inhibitor development was established using logistic regression analysis. To investigate the correlation between cytokine levels and high inhibitor titers, the inhibitor-positive samples were further categorized into good risk (< 10 BU, n = 14) and poor risk (≥ 10 BU, n = 20) groups based on immune tolerance induction therapy prognosis. Total plasma N-glycans were released, methylated, and analyzed by using MALDI-TOF MS/MS. A synthesized N-glycan was spiked in to quantify each glycoform. The study evaluated the correlation between HA sample groups and various glycosylation ratios, including for sialylation, core-fucosylation, galactosylation and galactose. The cut-off value of each glycosylation ratio was based on the median of the inhibitor-negative group. Results The adjusted odds ratio of inhibitor development for elevated G-CSF, IL-6 and decreased IL-10 were 2.84 (95% CI: 1.45-8.48; p = 0.021), 1.50 (95% CI: 1.14-2.42; p = 0.069), and 0.28 (95% CI: 0.10-0.61; p = 0.006), respectively in HA patients. Among InhPos HA patients, the levels of G-CSF, IL-2, IL-6, IL-8, and IL-10 in poor risk group were significantly higher than in good risk group (p-value < 0.05, ANOVA). Since aging also causes glycosylation changes, we analyzed adult (≥ 18 years old, n = 31) and pediatric (< 18 years old, n = 17) HA patients separately. Among analyzed glycan ratios, 80% of adult HA patients with inhibitors had a higher core-fucosylation ratio. All pediatric patients with inhibitors had significant lower sialylation and higher galactosylation ratios. Strikingly, pediatric patients with inhibitors exhibited around 2-fold higher core-fucosylation than those without inhibitor. Conclusion Our finding of elevated levels of G-CSF and IL-6 along with decreased levels of IL-10 associated with the presence of inhibitors, suggests the potential for prediction of inhibitor development. Moreover, high levels of G-CSF, IL-2, IL-6, IL-8 and IL-10 in patients with high inhibitor titers (≥ 10 BU, poor risk group) but not in good risk group imply they could be explored in future studies as prognostic factors in immune tolerance therapy. Glycan ratios can also hold promise as biomarkers including higher core-fucosylation in adult patients. Additionally, significant lower sialyation, higher galactosylation and core-fucosylation ratios indicate higher possibility of inhibitor development in pediatric patients.

Development Of Inhibitors In Hemophilia A Patients Receiving Different Treatment Options

Introduction: Hemophilia A is the most common inherited bleeding disorder in Pakistan. Apart from frequent bleeds and joint deformities, the formation of inhibitors is the major complication of hemophilia. Inhibitors are antibodies formed against the coagulation factor. The purpose of this study is to determine the incidence of inhibitor development in hemophilia A patient receiving different treatment modalities namely fresh frozen plasma (FFP)/cryoprecipitate, plasma derived FVIII concentrates and long-acting recombinant FVIII concentrates. Aims & Objectives: To assess the development of inhibitors in hemophilia A patients receiving treatment modalities. Place and Duration of Study: The study was conducted at Chughtai Institute of Pathology from November 2021 to November 2023. Material &Methods: A total of 75 registered patients of the Hemophilia Patient Welfare Society, Lahore and Sundas Foundation were initially screened for FVIII inhibitors. Those who were found negative for these inhibitors were included in the study and then divided into three equal groups of 25 patients each. One group received only FFP/cryoprecipitate for treatment, the other group received plasma derived FVIII concentrates and the last group received recombinant factor VIII concentrates. These patients were observed for two years i.e after 15-20 exposures and following this period their inhibitors screening was done again to establish the effect of different treatment modalities on inhibitor development. The data collected was entered and analyzed by SPSS-21, a p-value of ?0.05 was considered significant. Results: Our study showed that only 3 patients out of 25, receiving FFP as treatment modality developed inhibitors. Whereas, none of the patients receiving plasma derived FVIII concentrates and recombinant factor VIII concentrates developed inhibitors. The P value calculated is 0.044 which is significant. Conclusion: Plasma derived FVIII concentrates and recombinant FVIII concentrates are safer as compared to FFP/Cryoprecipitate and have lesser risk for inhibitor development.

Inhibitors in hemophilia: association with surgery plans and outcomes in a retrospective cohort study

BackgroundThe development of antibodies (inhibitors) to clotting factors compromises the management of hemophilia A and B, resulting in resistance to clotting factor replacement and, in many cases, the need for bypassing agents to achieve hemostasis. ObjectivesTo evaluate the association between the presence of inhibitors and achievement of perioperative hemostasis, development of complications, and presurgical plan deviations. MethodsWe conducted a retrospective study using data from the Indiana Hemophilia and Thrombosis Center surgical database (1998-2019). Associations between perioperative outcomes and inhibitor status were assessed while controlling for patient and procedural characteristics. ResultsA total of 1492 surgeries were performed in 539 persons with hemophilia, with 72 procedures performed in 20 patients with inhibitors (15 with hemophilia A; 5 with hemophilia B). High-responding inhibitors (>5 BU/mL) were present in 27 procedures, low-responding inhibitors (≤5 BU/mL) were present in in 13 procedures, and 32 procedures were performed in patients with historically persistent inhibitors. Adjusting for age, diagnosis, surgery setting, hemostatic agent, data collection period, and surgery type (major/minor), inhibitors were associated with a higher risk of inadequate perioperative hemostasis (33.4% vs 8.6%; adjusted relative risk [adjRR], 3.78; 95% CI, 1.89-7.56; P < .001). Reported complications include hemorrhage, fever, pain, thrombosis, and infections. Complications were not statistically different based on inhibitor status (31.7% vs 14.6%; adjRR, 1.25; 95% CI, 0.63-2.49; P = .526). Presurgical plan deviations (eg, hemostatic medication dose adjustments, procedure rescheduling, and changes in the length of postoperative hospitalization) occurred more frequently in surgeries involving inhibitors (70.8 vs 39.5%; adjRR, 1.47; 95% CI, 1.12-1.93; P = .005). ConclusionInhibitors are associated with higher risks of adverse perioperative outcomes. Strategies to address inhibitor development should be prioritized to avoid undesirable perioperative outcomes.

Frequency of Inhibitors among known Hemophilia A Patients.

ABSTRACT&#x0D; Introduction: Inhibitors are IgG alloantibodies which are directed against exogenous clotting factor VIII that neutralize the function of infused clotting factor concentrate used for the treatment. The prevalence of inhibitors in Hemophilia A is around 30% worldwide. Inhibitors occurs as a result of natural immune process of the body as factor concentrates used for treatment are foreign to the body and patient’s own body either produces no factor or produces structurally abnormal protein.&#x0D; Objectives: To determine the frequency of inhibitors among known hemophilia A patients.&#x0D; Methods: Cross-sectional survey done including 100 male patients of hemophilia A and on treatment for more than 1 year, from 2 to 60 years of age, were included. All the patients were undergone aPTT based screening for inhibitor, both immediately and two hours after mixing the patients plasma with the normal plasma.&#x0D; Results: Age range in this study was from 2 to 60 years with mean age of 37.21 ± 14.73 years. Frequency of inhibitors among known hemophilia A patients was seen in 18 (18.0%) of cases.&#x0D; Conclusion: This study concluded that frequency of inhibitors among known hemophilia A patients is quite high.&#x0D; Keywords: Hemophilia A, inhibitors, severity.

A child diagnosed with severe hemophilia A presenting with nephrotic syndrome: a case report

BackgroundNephrotic syndrome occurring as a complication of immune tolerance therapy for inhibitors in hemophilia B is well recognized. It is also known to occur in association with factor borne infections, especially hepatitis C. This is the first case report of nephrotic syndrome occurring in a child receiving prophylactic factor VIII in the absence of inhibitors of hepatitis infection. However, the pathophysiology of this phenomenon is poorly understood.Case presentationA 7-year Sri Lankan boy diagnosed with severe hemophilia A on weekly factor VIII prophylaxis was diagnosed with three episodes of nephrotic syndrome, a condition in which there is leakage of plasma protein into urine. He had three episodes of nephrotic syndrome, all of which responded well to 60 mg/m2 daily dose of oral steroids, achieving remission within 2 weeks of starting daily prednisolone. He has not developed inhibitors for factor VIII. His hepatitis screening remained negative.ConclusionsThere is a possible link between factor therapy for hemophilia A and nephrotic syndrome, which can be a T-cell-mediated immune response. This case also highlights the importance of monitoring for renal involvement in patients treated with factor replacement.

The Use of Bypassing Treatment Strategies in Hemophilia and Their Effect on Laboratory Testing.

Factor VIII and IX inhibitors in congenital hemophilia A and B, respectively, neutralize the infused coagulation factor concentrate rendering them ineffective. Bypassing agents (BPAs) that circumvent the block imposed by the inhibitors are used for the prevention and management of bleeding. Activated prothrombin complex concentrate was the original BPA, recombinant activated factor VII was then introduced, and more recently nonfactor agents that target the procoagulant and anticoagulant systems have been developed and are in clinical use (e.g., emicizumab, a bispecific antibody for hemophilia A). Other BPAs are in clinical trials (e.g., fitusiran targets antithrombin, concizumab and marstacimab target tissue factor pathway inhibitor, and SerpinPC targets activated protein C). The BPAs have a varied effect on coagulation assays, and as more patients are exposed to these agents, it is important to be aware of the effects. Herein, we present an overview of the effect of BPAs on routine and specialized coagulation assays including thrombin generation and viscoelastic assays.

Factor VIII inhibitors in hemophilia A treated with emicizumab: longitudinal follow-up of outcomes

BackgroundUsing emicizumab in lieu of immune tolerance induction (ITI) for patients with hemophilia A (HA) and factor (F)VIII inhibitors has been well described. However, decisions regarding ITI initiation, regimen, and preservation of tolerance remain to be elucidated. ObjectivesTo study the course of FVIII inhibitors in patients with HA and a history of FVIII inhibitors receiving emicizumab prophylaxis. MethodsPatients with HA, with and without FVIII inhibitors, initiating emicizumab prophylaxis were prospectively followed up in our center. All patients with current or previous inhibitors were included in this analysis. Plasma samples for FVIII inhibitor assays were obtained every 3 to 6 months or following FVIII exposure. Patients documented annual bleeding rate and any FVIII exposure days (EDs). ResultsOf 162 emicizumab-treated participants, 51 met the inclusion criteria. A decrease in annual bleeding rate was observed in all 51 participants followed up for a median of 3.3 years, with 31 breakthrough bleeding episodes reported in 22 of 51 participants. FVIII inhibitor level transiently increased following FVIII exposures in 5 of 15 failed ITI participants. Eight of 21 participants who did not undergo ITI were exposed to FVIII (1-2 EDs)), and 1 of these 8 participants demonstrated increased FVIII inhibitor levels after head trauma (following 1 ED). Among participants who underwent successful ITI, 8 of 15 patients were exposed to FVIII over a total of 13 EDs (1-2 ED(s) each) for traumatic breakthrough bleeds. In all these participants, inhibitor levels remained zero, indicating successful tolerance maintenance. ConclusionOur longitudinal follow-up of emicizumab-treated patients with HA and FVIII inhibitors shows that occasional exposure to FVIII may induce a transient anamnestic response. Nonetheless, no FVIII inhibitor recurrence was noted following FVIII exposures in patients who underwent successful ITI.

Prospective Hemophilia Inhibitor PUP Study reveals distinct antibody signatures during FVIII inhibitor eradication.

Factor VIII (FVIII) inhibitor formation is a major clinical concern during replacement therapy in patients with hemophilia A. Immune tolerance induction (ITI) is the only therapeutic approach to attempt inhibitor eradication and establishment of long-term immune tolerance to FVIII. Hemophilia Inhibitor Previously Untreated Patient (PUP) Study (HIPS) was a prospective clinical trial to investigate changes in the immune system of PUPs with severe hemophilia A. Five patients who developed persistent FVIII inhibitors during HIPS entered an ITI extension arm (HIPS-ITI). During HIPS-ITI, inhibitor patients received ITI with the same FVIII product (a single source of recombinant, human full-length FVIII) used in HIPS until successful tolerance, declared failure, or a maximum of 2 years after HIPS-ITI enrollment, whichever came first. Blood samples and clinical data were collected monthly. Longitudinal FVIII-binding antibody signatures, associated binding specificities, and apparent affinities were determined for each patient at each sampling time point. ITI was successful or partially successful in 2 patients and failed in 3. Both groups presented with distinct FVIII-specific antibody signatures. ITI success required the disappearance of FVIII inhibitors, which was associated with the eradication or sustained titer minimization of high-affinity FVIII-specific antibodies, particularly of the immunoglobulin G1 (IgG1) and IgG4 subclasses. In contrast, ITI failure, as reflected by FVIII inhibitor persistence, was associated with persistent high-affinity FVIII-specific antibodies. Interestingly, 1 patient with partial ITI success and 1 patient with ITI failure developed apparent oligoreactive FVIII-binding antibodies during ITI. The explanation of the true nature of these antibodies requires more comprehensive follow-ups in future studies. This trial was registered at www.clinicaltrials.gov as #NCT01652027.

Economic impact of inhibitors in pediatric hemophilia A

Economic impact of inhibitors in pediatric hemophilia A

Association between interleukin-2 (rs2069762) gene polymorphism and FVIII inhibitor development in Indonesian patients with severe hemophilia A

BACKGROUND Factor VIII (FVIII) inhibitors in hemophilia A (HA) patients render FVIII replacement therapy ineffective. Although its development cause is unclear, it has been classified into therapeutic and genetic-related etiologies. Single nucleotide polymorphisms (SNPs) in several cytokine genes, including interleukin (IL)-2, could increase the risk of FVIII inhibitor development. This study aimed to evaluate the association between IL-2 (rs2069762) gene SNP and FVIII inhibitor development in Indonesian patients with severe HA.&#x0D; METHODS The IL-2 (rs2069762) gene SNP was examined in 119 HA patients. The presence of FVIII inhibitors was quantified using an enzyme-linked immunosorbent assay, with a titer of &lt;0.28 ng/ml considered negative. Patients were divided into two groups: 59 with FVIII inhibitors (positive group) and 60 without inhibitors (negative group). The genotype of the subjects was determined using peripheral blood mononuclear cells and tetra-primer amplification refractory mutation system-polymerase chain reaction.&#x0D; RESULTS There was no association between IL-2 (rs2069762) gene polymorphism and FVIII inhibitor development on genotypes (p = 0.138) and allele frequencies (p = 0.780).&#x0D; CONCLUSIONS IL-2 (rs2069762) gene polymorphism is not a risk factor in the development of FVIII inhibitors in Indonesian patients with severe HA. Thus, further polymorphism studies in other cytokine genes are required to gain a comprehensive understanding of the FVIII inhibitor development.

Occurrence of FVIII Inhibitors in Hemophilia A Patients Following an Institutional Switch to a Third Generation B-Domain-Deleted FVIII.

In 2018, Refacto AFR, a B-domain-deleted third-generation FVIII concentrate, became our preferential product. After the introduction, the development of inhibitors was prospectively monitored; retrospectively, we sought for risk factors in the patients who developed a de-novo inhibitor. Over a period of 15 months, 4/19 adult patients with non-severe haemophilia who were treated on demand for surgery, developed high titer antibodies to FVIII after administration of Refacto AFR; 5/52 mostly severe patients on prophylaxis, developed an inhibitor (3 ≥ 0.1 BU; 1 > 0.6 BU, 1 high titre) after they switched to Refacto AFR; all were children <14 years of age and with >100 exposure days, none related to surgery or intensive treatment; all received KovaltryR before. In conclusion: inhibitors were encountered in on demand patients and previously treated prophylaxis patients; this observation might be a coincidental finding, but also risk factors like genotype and surgery and/or that Refacto AFR is more immunogenic should be considered. For the patients on prophylaxis we hypothesize that loss of tolerance by preceding KovaltryR might have contributed to inhibitor development.

Factor IX inhibitors in haemophilia B: A report of National Haemophilia Registry in China.

The development of inhibitors against factor FIX (FIX) is the most serious complication of FIX replacement therapy in haemophilia B (HB) patients. Currently, only few cohorts of HB inhibitor patients have been reported worldwide. This Chinese nationwide study of HB inhibitor patients explored their risk factors for FIX inhibitor development and experience on their management. We retrospectively analysed patient characteristics, F9 genotypes, treatment strategies and outcomes of HB inhibitor patients registered to the Chinese National Registry and Patient Organization Registry. Forty-four unique HB inhibitor patients were identified in 4485 unique HB patients registered by year 2021 to the two Registries. Inhibitor diagnosis were usually delayed and the low prevalence (.98%) may suggest some inhibitor patients were not identified. Their median age at inhibitor diagnosis was 7.5 (IQR, 3.0-14.8) years. Most patients (95.5%) had high-titre inhibitors. Allergic/Anaphylactic reactions occurred in 59.1% patients. Large deletions and nonsense mutations were the most common F9 mutation types in our FIX inhibitor patients. Patients with large F9 gene deletions were more likely to develop inhibitors (p =.0002), while those with missense mutations had a low risk (p <.0001). Thirteen (29.5%) patients received immune tolerance induction (ITI) therapy using low-dose prothrombin complex concentrate regimens. Twelve completed ITI with three (25.0%) achieving success. Nephrotic syndrome developed in two (16.7%) patients during ITI. This study reports the largest Chinese cohort of HB inhibitor patients. Large deletions were most significantly associated with inhibitor development. Low-dose ITI might be feasible for FIX inhibitor eradication.

Can T-cell and B-cell excision circles predict development of inhibitors in pediatric hemophilia A?

Hemophilia A (HA) therapy requires intravenous replacement infusions of factor (F) VIII concentrate. Inhibitors are high-affinity immunoglobulin G that are directed against FVIII and thereby render replacement therapy ineffective. This complication has significant prognostic implications. We aimed to examine the immune system involvement in inhibitor formation specifically T-cell excision circles (TRECs) and B-cell excision circles (KRECs), markers of new T and B cells, respectively, and examine them as surrogate markers for inhibitor formation. Blood samples were collected from 35 children with severe HA. Children were divided into two groups: with FVIII inhibitors and without FVIII inhibitors. TRECs and KRECs were measured in peripheral blood. A total of 11 patients with inhibitors and 24 without were evaluated. Children with inhibitors had higher levels of TRECs however not statistically significant (p = 0.085). CjKREC levels were higher in the inhibitor patients (p = 0.003). Moreover, the sj/cjKREC ratio was lower in the inhibitor patients (p = 0.015). Our findings may add to the notion that inhibitor formation is attributed to humoral immunity due to peripheral B-cell expansion and loss of peripheral tolerance. Improved knowledge regarding the involvement of the immune system in the formation of FVIII inhibitors will enable better therapy tailoring in the era of non-replacement therapies. The etiology of FVIII inhibitor formation is multifactorial, in which the immune system plays a pivotal role. Our findings may add to the notion that inhibitor formation is attributed to humoral immunity due to peripheral B-cell expansion and production of antibodies against FVIII. Improved knowledge regarding the involvement of the immune system in the development of FVIII inhibitors will enable the identification of patients prone to inhibitor development and better therapy tailoring in the new era of non-replacement therapies.

Risk factors for inhibitors in hemophilia A based on RNA‐seq and DNA methylation

BackgroundThe development of factor VIII (FVIII) inhibitor is a severe complication during replacement therapy for hemophilia A patients. ObjectivesWe investigated the potential risk factors for FVIII inhibitor formation based on genome‐wide RNA‐sequencing and whole‐genome bisulfite sequencing analysis. MethodsRNA‐sequencing and whole‐genome bisulfite sequencing analysis were applied on 17 blood samples with F8 intron 22 inversion, including seven with inhibitors and 10 without. ResultsAltogether, 344 mRNA transcripts and 20 long noncoding RNAs (lncRNA) transcripts were differentially expressed. Among the differentially expressed transcripts, 200 mRNAs and 12 lncRNAs were upregulated, and 144 mRNAs and eight lncRNAs were downregulated. Gene ontology enrichment analysis of differentially expressed mRNAs showed that genes involved in immune stimulation, especially those for T‐cell activation, were upregulated, whereas genes involved in negative immune response regulation were downregulated. Coexpression analysis revealed that the targeted upregulated genes of differentially expressed lncRNA were similarly closely related to immune activation, especially T‐cell activation. Methylation analysis showed inhibitor patients exhibited a slightly lower methylation status in the CpG islands, 5′ untranslated region, and exon regions (p < 0.01). Genes with differentially methylated regions were also related to T‐cell activation. ConclusionsThere is an upregulation of genes involved in activation of the immune system in hemophilia A patients with inhibitors. The lncRNA and methylation modifications may play important roles in inhibitor production. These findings are potentially to reveal novel therapeutic targets for prevention and treatment of inhibitors.

Frequency of Specific and Non-specific Inhibitors in Haemophilia A Patients.

Objective: To determine the frequency of specific and non-specific inhibitors in haemophilia A patients.Study design: This is a cross-sectional study.Patients and methods: A total of 150 male haemophilia A patients were included in this cross-sectional study at the National Institute of Blood Diseases and Bone Marrow Transplant (NIBD), Karachi, Pakistan, from September 2019 to January 2022.Results: Among 150 patients included in this study, 23 (15.3%) had an inhibitor and 127 (84.6%) did not. All patients had specific inhibitors against Factor VIII (FVIII). Non-specific inhibitors were not identified in our population. Among the patients in the inhibitor group, there were 13 (56.5%) in the severe (<1%) category. There were 10 (43.5%) patients in the moderate (1-5%) category. There were no patients in the mild category. The median inhibitor level was 15.4 Bethesda unit (BU).Conclusion: The development of inhibitors has not been identified as a major problem in our population. However, it is noteworthy that only 15.3% of patients with haemophilia A developed inhibitors in this data set. They were essentially treated with plasma and its products.