Objective:
Repeated joint bleeding in patients with hemophilia leads to hemophilic arthropathy (HA), which cannot be entirely prevented by clotting factor replacement. Vascular remodeling and permeability are associated with hemarthrosis and may contribute to HA progression; however, the underlying mechanisms and effects of FVIII replacement are poorly understood. Here, we explored vascular changes and synovial gene expression profiles in FVIII-deficient mice after induced hemarthrosis +/- FVIII replacement.
Methods:
Hemarthrosis was induced in FVIII-deficient mice by sub-patellar needle puncture +/- 100-200 IU/kg recombinant human FVIII (rhFVIII) intravenously 2 hours before and 6 hours after injury. Vascularity and gene expression were analyzed at baseline and 2 weeks post-injury. Vascularity was assessed by histology with Safranin-O-Fast Green and α-smooth muscle actin (αSMA) staining, and by musculoskeletal ultrasound with Power Doppler to detect microvascular flow. The permeability of synovial vessels was determined by quantification of extravasated albumin-bound Evans blue dye in knee joints at near-infrared fluorescence. For gene expression studies, RNA was extracted from synovial tissue and cDNA libraries were prepared using the NEBNext Ultra II DNA Library Prep Kit and sequenced on an Illumina NextSeq500 platform (single-end; 75bp reads). The R BioConductor packages tximport, edgeR and limma were used to read Salmon transcript files and implement the limma-voom method for differential expression analyses. Functional enrichment was performed using Signaling Pathway Impact Analysis.
Results:
Knee injury caused profound hemarthrosis in vehicle-treated mice that was largely prevented with rhFVIII prophylaxis (day 2 hematocrit: 26.4% and 46.3%, respectively). Soft tissue proliferation increased to a similar extent in both groups, as did various vascular parameters: microvascular flow (vehicle: 1.8-fold; rhFVIII: 1.5-fold), vessel number (vehicle: 2.2-fold, p=0.0005; rhFVIII: 2.0-fold, p=0.004), vessels with diameter ≥ 20 µm (vehicle: 2.9-fold, p=0.02; rhFVIII: 2.7-fold, p=0.02), and αSMA area per vessel (vehicle: 2.3-fold, p>0.05; rhFVIII: 3.6-fold, p=0.0006). Vascular permeability also increased significantly after joint bleeding (1.7-fold, p=0.0007) and was only partially rescued by rhFVIII prophylaxis (1.3-fold, p>0.05). RNA sequencing revealed a strong transcriptional response (1527 differentially expressed genes (DEG), 13 perturbed pathways), that was not substantially dampened in rhFVIII-treated mice (891 DEG, 20 perturbed pathways). Notably, perturbation of extracellular matrix (ECM)-receptor interactions was highly significant with vehicle (pGFWER=1.7x10-11) and rhFVIII prophylaxis (pGFWER=1.4x10-10). STRING analysis of the top 20 DEG revealed enrichment of ECM components and organization, and numerous genes encoding ECM constituents, including collagens and MMPs, were significantly upregulated. Changes in ECM gene expression may facilitate the observed synovial tissue and vascular remodeling after hemarthrosis.
Conclusions:
Hemarthrosis in FVIII-deficient mice triggers profound changes in synovial gene expression that may drive associated tissue and vascular remodeling processes. These changes are incompletely mitigated by rhFVIII prophylaxis. Further exploration will enable identification of disease markers and targeted drug discovery to intercept the progression of HA.
Background:
Inherited factor X deficiency is an autosomal recessive bleeding disorder with an estimated occurrence rate of 1:1,000,000¹. Historically, bleeding symptoms have been treated with topical therapies, antifibrinolytic agents, fresh frozen plasma (FFP) or plasma-derived FIX concentrates (PCCs). In 2015, the first factor X (FX) concentrate was approved in the U.S.
Objective:
This organization was interested in reviewing clinical outcomes such as perceived pain and unplanned hospitalizations of adults and children with FX disease currently being treated in the home with Coagadex®. METHODS: This organization conducted a retrospective review of a population of seven adult and pediatric patients. Patients were surveyed for pain, bleeding episodes, hospitalizations/ ER visits, dosing parameters and administration methods pre/post initiation of FX therapy. There were 3 children, 12 years old and under and four adults. Ages ranged from 5-60 years old with the average age of 27.9. There were five males and two females. The average length of treatment was 6.4 months. One patient was naïve, six converted from other therapies. Dose ranges administered by caregivers or self-infusion were 750 -2800 IU (26-61 IU/Kg). One patient was on-demand and six were administering prophylaxis therapy.
Results:
There were two converted prophylaxis patients reported pain with PCC’s and none with FX; one on-demand naïve patient stated his pain was markedly improved with prn administration of FX; four converted prophylaxis patients with no prior pain history reported no changes in pain on FX therapy. For on-demand patients treating bleeding episodes, three reported a decrease in the number of bleeding episodes, three were unchanged and one reported one additional bleeding episode. A total of ten hospitalizations or emergency room visits were reported during the six months prior to initiation of FX treatment and only one in the six months following initiation of treatment.
Conclusion:
Early recognition and home treatment with FX concentrate allows for prompt resolution of bleeding symptoms, decreased pain and decreased hospitalization or emergency room visits. Further investigation is needed to determine cost-savings for decreased hospitalization/ ER visits.References:Brown, D.L. & Kouides, P.A. (2008). Diagnosis and treatment of Inherited Factor X deficiency. Haemophilia. (14). 1176-1182. Retrieved from: https://www.hemophilia.org/sites/default/files/document/files/DiagnosisAndTreatmentOfInheritedFact…
Klaus Bonazza received his Ph.D. in chemistry from Vienna University of Technology. He is currently a postdoctoral researcher at Boston Children's Hospital and appointed at Harvard Medical School, mentored by Dr. Timothy Springer. His field of interest is the ultra-large concatemeric protein von Willebrand factor (VWF), which accounts for the adaptability of hemostasis to different flow conditions in the blood vessels.
At moderate, physiological flow VWF has a packed, "bird nest's" shape whereas strong elongational flow conditions, occurring downstream of vascular restrictions or injuries, induce a transition to a threat-like, elongated state. On top of this overall unpacking, tensile forces, which are exerted on the chain and transmitted by its A1 domain, cause local conformational changes which activate binding of thrombocyte receptor Glycoprotein Ib (GPIbα) to initiate coagulation. With his JGP fellowship award, Dr. Bonazza will pioneer a new method to obtain structural insights into force dependent VWF unpacking, A1 deformation and GPIbα binding based on hydrogendeuterium exchange under elongational flow conditions.
As a social worker at Gulf States Hemophilia & Thrombophilia Treatment Center, I have the privilege of serving patients across their lifespan. I would like to initiate grass roots education about hemophilia in Houston, Texas, by offering educational programming to specialized health care professionals who work directly with the aging population in nursing homes and assisted living communities. This would include executive directors and administrators of these facilities as well as direct clinical staff.
Objective:
BAY 94-9027 is an extended–half-life recombinant factor VIII (FVIII) product. In the PROTECT VIII Kids trial, BAY 94-9027 was efficacious for the prevention and treatment of bleeding episodes in previously treated children with severe hemophilia A. We report interim long-term efficacy and safety data from the PROTECT VIII Kids extension study.
Methods:
In PROTECT VIII Kids, previously treated patients (PTPs) aged <12 years with severe hemophilia A received BAY 94-9027 prophylaxis twice weekly (25‒60 IU/kg), every 5 days (45‒60 IU/kg), or every 7 days (60 IU/kg). Patients completing ≥50 exposure days (EDs) and ≥6 months in the main study or a 12-week safety substudy (part 2) that enrolled PTPs aged <6 years could continue in the optional extension for an additional ≥50 EDs.
Summary:
Fifty-nine of 73 patients treated with BAY 94-9027 in PROTECT VIII Kids (main study or part 2) continued in the extension (median [range] age at enrollment in the main study, 5.0 [2–11] years). At data cutoff (January 2018), patients had a median (range) of 1456 (351–1665) days in the trial (main study or part 2 plus extension). Patients in the extension received prophylaxis twice weekly (n=20), every 5 days (n=20), every 7 days (n=8), or switched prophylaxis frequency during the extension (variable frequency; n=11). Median (range) dose/infusion was 52.1 (19–62) IU/kg. Median annualized bleeding rate (ABR) for total bleeds was 1.8 for all patients and 0.8, 1.1, 2.1, and 3.2 for those treated twice weekly, every 5 days, every 7 days, or with varying frequency, respectively. Median ABR for joint bleeds in all patients was 0.7. During the extension, 3 patients (5.1%) experienced treatment-related adverse events (AEs) classified as mild (n=1), moderate (n=1), or severe (n=1). One patient discontinued because of a serious AE that was not related to treatment. No confirmed FVIII inhibitors or anti-PEG antibodies were observed; no patients had sustained levels of detectable PEG in plasma.
Conclusions:
Long-term treatment (up to ~4.5 years) with BAY 94-9027 prophylaxis was efficacious and well tolerated in previously treated pediatric patients with severe hemophilia A.
