The thrombin generation assay distinguishes inhibitor from non-inhibitor patients with severe haemophilia A.

INTRODUCTION: Patients with haemophilia A (HA) have impaired thrombin generation (TG) capacity and TG assay (TGA) values are linearly related to plasma factor VIII (FVIII) levels. AIM: This study carried out in patients with unmeasurable FVIII (<1 IU dL(-1) ) was aimed at unravelling any difference in TG capacity in patients with or without inhibitors. METHODS: Blood samples were collected from patients in a non-bleeding state, after a 5-day wash-out period from last treatment. RESULTS: TGA was performed in 102 patients with severe HA (15% with high-responding inhibitors; 51% with null F8 mutations, that as expected were more prevalent in inhibitor than in non-inhibitor patients). TG capacity was significantly lower in inhibitor than non-inhibitor patients and in those with null mutations than in those with non-null mutations. When the TG capacity was evaluated only in patients with null mutations with and without inhibitors it was lower in the presence of inhibitors. CONCLUSIONS: This study shows a greater TG impairment in inhibitor patients irrespective of FVIII levels, inhibitor titre and F8 mutation type, suggesting a role for the TGA in unravelling functional interferences of anti-FVIII inhibitors on coagulation system activation.

New predictive approaches for ITI treatment.

Immune tolerance induction (ITI) therapy in patients with haemophilia A and inhibitors constitutes a huge burden for affected patients and families and poses a large economic burden for a chronic disease. Concerted research efforts are attempting to optimize the therapeutic approach to the prevention and eradication of inhibitors. The Italian ITI Registry has provided data on 110 patients who completed ITI therapy as at July 2013. Analysis of independent predictors of success showed that, together with previously recognized factors - namely inhibitor titre prior to ITI, historical peak titre and peak titre on ITI - the type of causative FVIII gene mutation also contributes to the identification of patients with good prognosis and may be useful to optimize candidate selection and treatment regimens. Numerous studies have demonstrated that inhibitor reactivity against different FVIII products varies and is lower against concentrates containing von Willebrand factor (VWF). An Italian study compared inhibitor titres against a panel of FVIII concentrates in vitro and correlated titres with the capacity to inhibit maximum thrombin generation as measured by the thrombin generation assay (TGA). Observations led to the design of the PredictTGA study which aims to correlate TGA results with epitope specificity, inhibitor reactivity against different FVIII concentrates and clinical data in inhibitor patients receiving FVIII in the context of ITI or as prophylactic/on demand treatment. At the immunological level, it is known that T cells drive inhibitor development and that B cells secrete FVIII-specific antibodies. As understanding increases about the immunological response in ITI, it is becoming apparent that modulation of T-cell- and B-cell-mediated responses offers a range of potential new and specific approaches to prevent and eliminate inhibitors as well as individualize ITI therapy.

A Scan of Pleiotropic Immune Mediated Disease Genes Identifies Novel Determinants of Baseline FVIII Inhibitor Status in Hemophilia-A.

Hemophilia-A (HA) is caused by heterogeneous loss-of-function factor (F)VIII gene (F8)-mutations and deficiencies in plasma-FVIII-activity that impair intrinsic-pathway-mediated coagulation-amplification. The standard-of-care for severe-HA-patients is regular infusions of therapeutic-FVIII-proteins (tFVIIIs) but ~30% develop neutralizing-tFVIII-antibodies called "FVIII-inhibitors (FEIs)" and become refractory. We used the PATH study and ImmunoChip to scan immune-mediated-disease (IMD)-genes for novel and/or replicated genomic-sequence-variations associated with baseline-FEI-status while accounting for non-independence of data due to genetic-relatedness and F8-mutational-heterogeneity. The baseline-FEI-status of 450 North American PATH subjects-206 with black-African-ancestry and 244 with white-European-ancestry-was the dependent variable. The F8-mutation-data and a genetic-relatedness matrix were incorporated into a binary linear-mixed model of genetic association with baseline-FEI-status. We adopted a gene-centric-association-strategy to scan, as candidates, pleiotropic-IMD-genes implicated in the development of either ³2 autoimmune-/autoinflammatory-disorders (AADs) or ³1 AAD and FEIs. Baseline-FEI-status was significantly associated with SNPs assigned to NOS2A (rs117382854; p=3.2E-6) and B3GNT2 (rs10176009; p=5.1E-6), which have functions in anti-microbial-/-tumoral-immunity. Among IMD-genes implicated in FEI-risk previously, we identified strong associations with CTLA4 assigned SNPs (p=2.2E-5). The F8-mutation-effect underlies ~15% of the total heritability for baseline-FEI-status. Additive genetic heritability and SNPs in IMD-genes account for >50% of the patient-specific variability in baseline-FEI-status. Race is a significant determinant independent of F8-mutation-effects and non-F8-genetics.

F8 gene mutation profile in Indian hemophilia A patients: Identification of 23 novel mutations and factor VIII inhibitor risk association.

'FVIII inhibitors', especially in severe hemophilia A (HA) patients, is a serious adverse effect that complicates their clinical management. Many genetic and non-genetic risk factors have been proposed for FVIII inhibitor development, diverse in different population groups. This is the first study in Indian hemophiliacs that analyzes inhibitor risk in relation to the complete F8 mutation profile, in a case-control study that included 145 Indian severe HA patients, i.e. 69 inhibitor positive (with 18 inhibitor concordant/discordant family members), and 58 inhibitor negative patients, after informed consent. While 53.54% (68/127) index cases were positive for intron 22 or intron 1 inversions, 55 causative F8 mutations were detected in the 59 inversion negative patients, of which 23 were novel mutations (in 24 patients) and 32 were reported earlier (in 35 patients). A higher incidence of mutations, in the C1 and C2 domains in inhibitor positive patients, and in the A1 domain in inhibitor negative patients was observed, though not significantly different. The study suggests that large F8 rearrangements (significantly higher in the inhibitor positive patients) pose the highest risk, while missense mutations (significantly higher in the inhibitor negative patients) pose the lowest risk of inhibitor development in Indian hemophilia A patients.