Chantal Rothschild

Impact of 5 years of maximally successful highly active antiretroviral therapy on CD4 cell count and HIV-1 DNA level.

Objectives: To evaluate the impact on CD4 cell count and HIV-1 DNA level in peripheral blood mononuclear cells (PBMC) of long-term highly active antiretroviral therapy (HAART) in the setting of maximal success, i.e., constant plasma HIV-1 RNA load suppression. Design: Retrospective analysis of patients selected for a constantly undetectable plasma HIV-1 RNA load since HAART initiation. Methods: HIV-1 DNA was measured in PBMC using a real-time polymerase chain reaction assay. Loess estimates and regression analysis were used for modelling the variations of the CD4 cell count and HIV DNA level over time. Results: The study included 41 patients chronically infected with HIV-1 who had been taking HAART for a median duration of 60.4 months and had an undetectable plasma HIV RNA load ever since the first 6 months of HAART; 25 were tested for HIV-1 DNA. The mean CD4 cell count increase was high during the first 18 months on therapy (168 × 106 cells/l per year), much lower afterwards (38 × 106 cells/l per year), independently of the baseline CD4 cell count. Most of the patients (73.2%) reached a CD4 cell count constantly ⩾ 400 × 106/l during follow-up. HIV-1 DNA showed a mean decrease of 0.48 log10 copies/106 PBMC during the first year, of 0.18 log10 copies/106 PBMC per year during the 2nd and 3rd years, but no significant decrease afterwards. Conclusions: These results question the benefit of very long-term maintenance of HAART in terms of CD4 gain and HIV-1 DNA reduction.

Plasma and albumin-free recombinant factor VIII: pharmacokinetics, efficacy and safety in previously treated pediatric patients.

Summary.  Background: The pharmacokinetics of factor VIII replacement therapy in preschool previously treated patients (PTPs) with hemophilia A have not been well characterized. Objectives: To assess the pharmacokinetics, efficacy and safety of a plasma‐free recombinant FVIII concentrate, ADVATE [Antihemophilic Factor (Recombinant), Plasma/Albumin‐Free Method, rAHF‐PFM], in children < 6 years of age with severe hemophilia. Patients/methods: Fifty‐two boys, one girl, mean (± SD) age 3.1 ± 1.5 years and ≥ 50 days of prior FVIII exposure, were enrolled in a prospective study of ADVATE rAHF‐PFM at 23 centers. Results: The mean terminal phase half‐life (t1/2) was 9.88 ± 1.89 h, and the mean adjusted in vivo recovery (IVR) was 1.90 ± 0.43 IU dL−1 (IU kg−1)−1. Over the 1–6‐year age range, t1/2 of rAHF‐PFM increased by 0.40 h year−1. IVR increased by 0.095IU dL−1(IU kg−1)−1 (kg m−2)−1 in relation to body mass index (BMI). Patients primarily received prophylaxis. Median (range) annual joint bleeds were 0.0 (0.0–5.8), 0.0 (0.0–6.1) and 14.2 (0.0–34.5) for standard prophylaxis, modified prophylaxis and on‐demand treatment, respectively. Bleeds were managed in 90% (319/354) of episodes with one or two rAHF‐PFM infusions; response was rated excellent/good in 93.8% of episodes. Over a median 156 exposure days, no FVIII inhibitors were detected and no related severe adverse events or unusual non‐serious adverse events were seen. Conclusions: Children < 6 years of age appear to have shorter FVIII t1/2 and lower IVR values than older subjects. However, these parameters increased with age (t1/2) and BMI (adjusted IVR), respectively. rAHF‐PFM was clinically effective and well tolerated, with no signs of increased immunogenicity in previously treated young children with hemophilia A.

Recombinant factor VIII products and inhibitor development in previously untreated boys with severe hemophilia A.

Six recombinant factor VIII (rFVIII) products have been marketed worldwide. In 2013, the Research of Determinants of Inhibitor Development (RODIN) study group reported an unexpectedly high risk of inhibitor development with a second-generation full-length rFVIII (Product D) in previously untreated patients (PUPs) with severe hemophilia A (HA). In 1994, French public health authorities established a prospective cohort to monitor hemophilia treatment safety. A PUP subgroup was designed to investigate inhibitor risk factors. We analyzed this subcohort in view of the RODIN findings. After excluding 50 patients who participated in the RODIN study, the primary analysis focused on 303 boys with severe HA first treated with a rFVIII product. A clinically significant inhibitor was detected in 114 boys (37.6%). The inhibitor incidence was higher with Product D vs the most widely used rFVIII product (adjusted hazard ratio [aHR], 1.55; 95% confidence interval [CI], 0.97-2.49). Similar results were found for high-titer inhibitors and in 10 sensitivity analyses. No heterogeneity was observed between RODIN and our results. Combined aHRs were 1.58 (95% CI, 1.17-2.14) for all inhibitors and 1.70 (95% CI, 1.15-2.52) for high-titer inhibitors. Our results confirm the higher immunogenicity of Product D vs other rFVIII products in PUPs with severe HA.

Absence of collagen-induced platelet activation caused by compound heterozygous GPVI mutations

The glycoprotein VI (GPVI)/FcRgamma complex is a key receptor for platelet activation by collagen. We describe, for the first time, 2 genetic abnormalities in one patient. This 10-year-old girl presented ecchymoses since infancy, a prolonged bleeding time despite a normal platelet count and no antiplatelet antibodies. Collagen-induced platelet activation was null, whereas GPVI quantification by flow cytometry evidenced an incomplete deficiency. Immunoblotting showed an abnormal migration of residual GPVI, and no FcRgamma defect. GPVI DNA sequencing revealed (1) an R38C mutation in exon 3 of one allele and (2) an insertion of 5 nucleotides in exon 4 of the other allele, leading to a premature nonsense codon and absence of the corresponding mRNA. Introduction of the R38C mutation into recombinant GPVI-Fc resulted in abnormal protein migration and a loss of collagen binding. Thus, this composite genetic GPVI deficiency and dysfunction cause absence of platelet responses to collagen and a mild bleeding phenotype.

ReFacto® and Advate®: a single‐dose, randomized, two‐period crossover pharmacokinetics study in subjects with haemophilia A

Summary.  ReFacto is a recombinant B‐domain‐deleted, monoclonal antibody‐purified, solvent‐detergent‐treated factor VIII (BDDrFVIII) with no albumin added to the final formulation. Although ReFacto has been shown to be bioequivalent to a plasma‐derived FVIII product (Hemophil‐M) in a randomized, crossover pharmacokinetic (PK) study, the comparability of ReFacto with the full‐length (complete sequence) recombinant FVIII (FLrFVIII, Advate) product has not been previously examined in this manner. The primary objective of this study was to compare the PKs of ReFacto with those of Advate in patients with severe haemophilia A. This was a third‐party unblinded, randomized, multicentre, two‐period crossover PKs study of ReFacto and Advate in subjects with severe haemophilia A (FVIII:C ≤1%). Blood samples were collected over a 48‐h period after i.v. administration of each of the FVIII products. FVIII:C was determined using the chromogenic substrate assay (CSA) in a central laboratory. The plasma FVIII:C PK parameters of ReFacto and Advate were determined using non‐compartmental analysis. Bioequivalence was assessed on maximum plasma concentration (Cmax) and the area under the plasma concentration vs. time curves (AUCs) using an anova. The two products were judged to be equivalent if the 90% confidence limits of the ratio of the geometric mean values of Cmax and AUCs fell within the interval of 80–125%. Results from this PKs comparison of two different rFVIII products, using chromogenic substrate assay to measure FVIII:C, showed that ReFacto and Advate are bioequivalent to each other.

European data of a clinical trial with a sucrose formulated recombinant factor VIII in previously treated haemophilia A patients.

Summary.  To increase the safety of antihaemophilic treatment, the production process of full‐length recombinant factor VIII (FVIII) KOGENATE® Bayer (Kogenate®FS)has been modified. Human albumin is no longer added as stabilizer during purification and in final formulation. Instead, the new KOGENATE® Bayer production process uses sucrose as a stabilizer in the formulation and adds solvent/detergent virus inactivation step. An European clinical trial was carried out in Germany and France in previously treated patients with severe haemophilia A who had more than 100 exposure days to exogenous FVIII. Pharmacokinetic data was analysed according to one‐stage and chromogenic assays. Efficacy and safety during home therapy and in surgical procedures were evaluated; inhibitor formation was carefully monitored. Safety and efficacy were evaluated in 33 European patients for 24 months. Patients received more than 13 million IU KOGENATE® Bayer. Over 75% of patients accrued more than 100 exposure days with the new product. Of 875 bleeding episodes, 90.7% were treated with 1 or 2 infusions and 75.8% of responses to treatment were rated as ‘excellent’ or ‘good’. Prophylactic treatment was the most common mode of therapy (60.7% of infusions). The product was well‐tolerated and FVIII recovery studies were consistent throughout the study period. Only 0.26% of adverse events were reported to be drug related. No evidence of de novo inhibitor formation was observed. Overall, KOGENATE® Bayer was efficacious, safe and well‐tolerated for the treatment of haemophilia A in multitransfused patients.

Development of inhibitory antibodies to therapeutic factor VIII in severe hemophilia A is associated with microsatellite polymorphisms in the HMOX1 promoter

Induction of heme oxygenase-1, a stress-inducible enzyme with anti-inflammatory activity, reduces the immunogenicity of therapeutic factor VIII in experimental hemophilia A. In humans, heme oxygenase-1 expression is modulated by polymorphisms in the promoter of the heme oxygenase-1-encoding gene (HMOX1). We investigated the relationship between polymorphisms in the HMOX1 promoter and factor VIII inhibitor development in severe hemophilia A. We performed a case-control study on 99 inhibitor-positive patients and 263 patients who did not develop inhibitors within the first 150 cumulative days of exposure to therapeutic factor VIII. Direct sequencing and DNA fragment analysis were used to study (GT)n polymorphism and single nucleotide polymorphisms located at −1135 and −413 in the promoter of HMOX1. We assessed associations between the individual allele frequencies or genotypes, and inhibitor development. Our results demonstrate that inhibitor-positive patients had a higher frequency of alleles with large (GT)n repeats (L: n≥30), which are associated with lesser heme oxygenase-1 expression (odds ratio 2.31; 95% confidence interval 1.46–3.66; P<0.001]. Six genotypes (L/L, L/M, L/S, M/M, M/S and S/S) of (GT)n repeats were identified (S: n<21; M: 21≤n<30). The genotype group including L alleles (L/L, L/M and L/S) was statistically more frequent among inhibitor-positive than inhibitor-negative patients, as compared to the other genotypes (33.3% versus 17.1%) (odds ratio 2.21, 95% confidence interval 1.30–3.76; P<0.01). To our knowledge, this is the first association identified between HMOX1 promoter polymorphism and development of anti-drug antibodies. Our study paves the way towards modulation of the endogenous anti-inflammatory machinery of hemophilia patients to reduce the risk of inhibitor development

Deaths associated with acquired haemophilia in France from 2000 to 2009: multiple cause analysis for best care strategies.

Summary.  Deaths occurring in the context of acquired haemophilia (AH) may be related to inter‐connected causes and mechanisms including bleeding, specific or older patient co‐morbidities or iatrogenic complications. However, their magnitude remains unknown. This study aimed to determine the respective weight and frequency of the various causes of death in AH. Multiple‐cause analysis based on death certificates data is used in this purpose. Over a 10‐year period (2000–2009), 121 deaths with AH as a cause were registered in France. All the deaths were of adults (extremes: 47 and 99 years; mean age: 80.7 years). The average number of causes per death certificate was 4.7. AH was the underlying cause of death (UCD) in 69.4% of the cases, and was more frequent in the older subjects. In contrast, before age of 75 years, AH was more often a contributing cause of death. No postpartum or obvious thromboembolism‐related deaths were registered. Haemorrhagic shock was the most frequent direct cause of death (DCD), followed by infectious events, cardiac dysfunction, metabolic and nutritional disorders with muscle wasting and decubitus complications, and cancers (52.9%, 26.4%, 7.5%, 5.8% and 4.1%, respectively). However, when AH was not reported as an UCD, infections become the first DCD (32.4%) followed by bleeding events (16.2%). Best prophylactic and curative strategies for infections are particularly required to improve the prognosis in AH. Moreover, as several of its DCD correspond also to steroids side effects, best tolerated immunosuppressant regimen with steroid‐sparing agents adjoining are particularly awaited in AH population.

Characteristics, mechanisms of action, and epitope mapping of anti-factor VIII antibodies.

The development of anti-factor VIII (FVIII) antibodies (Abs), also called inhibitors, is currently one of the most serious complications arising during the treatment of hemophilia A patients. Improved prevention and eradication of these Abs remain a challenge both for clinicians and scientists. Numerous studies in the literature have reported on their epitope specificity, on their mechanism of FVIII inactivation, as well as on the methods used for their detection. In this review, we summarize the current knowledge on the nature (isotypes, kinetic properties), epitope properties, and mechanisms of action of anti-FVIII Abs. Furthermore, we present methods for detection and epitope characterization of anti-FVIII Abs with emphasis on the Luminex technique susceptible to facilitate the monitoring of changes in the epitope specificity of these Abs.

Analyses of the FranceCoag cohort support immunogenicity differences among one plasma-derived and two recombinant factor VIII brands in boys with severe hemophilia A

Around one third of boys with severe hemophilia A develop inhibitors (neutralizing antibodies) against their therapeutic factor VIII product. This adverse effect may result in more life-threatening bleeding, disability, impaired quality of life, and costly care. We compared the incidence of inhibitors in boys treated with the three factor VIII products most used in France: one plasma-derived (Factane) and two recombinant products (Advate and Kogenate Bayer). A previously untreated cohort of patients was created in 1994 to investigate risk factors for inhibitor development. We selected boys with severe hemophilia A (factor VIII <1 IU/dL) first treated with one of the three factor VIII products studied. Details of product infusions, inhibitor assays and main fixed and time-varying inhibitor risk factors were recorded for the first 75 exposure days. Three outcomes (all inhibitors, high-titer inhibitors and subsequently treated inhibitors) were analyzed by univariate and multivariate Cox models. We studied 395 boys first treated between 2001 and 2016 (131, 137, and 127 with Factane, Advate, and Kogenate Bayer, respectively). Clinically significant inhibitors were diagnosed in 121 patients (70 high-titer). The incidence of high-titer inhibitors was significantly associated with the factor VIII product received (P=0.005): the cumulative incidence at 75 exposure days was 12.7% (95% CI: 7.7–20.6) with Factane, 20.4% (95% CI: 14.0–29.1) with Advate, and 31.6% (95% CI: 23.5–41.7) with Kogenate Bayer. The low inhibitor incidence observed with Factane is concordant with recent findings from the SIPPET randomized trial. These consistent results from observational and experimental studies should lead to improved care for previously untreated patients and cost savings for healthcare systems worldwide.