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BACKGROUND: Persons with hemophilia (PWH) are at risk for chronic hemophilic arthropathy (HA). Joint replacement surgery may be used to relieve intractable pain and/or restore joint function. OBJECTIVES: This multicenter, prospective, observational cohort study evaluated the rate of bleeding during the postoperative period after total hip (THA) or knee arthroplasty (TKA). PATIENTS/METHODS: We included PWH of any severity ≥18 years of age who were undergoing THA or TKA. Clinical decisions were made at the discretion of the treating physician according to local standards of care. Clinical data were prospectively recorded. Major bleeding was defined as bleeding in a critical site, bleeding that resulted in either a 2 g/dL or greater decrease in hemoglobin during any 24-hour period, or transfusion of two or more units of packed red blood cells. RESULTS: One hundred thirty-one procedures (98 TKA and 33 THA) were performed, 39 (29.8%) of which were complicated by major bleeding, including 46% of THA and 25% of TKA. The risk of major bleeding was increased in THA compared to TKA (OR 2.50, p=0.05), and by the presence of an inhibitor (OR 4.29, p=0.04), increased BMI (OR 4.49 and 6.09 for overweight and obese, respectively, compared to normal BMI, each p

The direct oral anticoagulants (DOACs), apixaban and rivaroxaban, have been studied for extended-phase treatment of venous thromboembolism (VTE). Yet, scant evidence exists surrounding clinician practice and decision-making regarding dose reduction.Report clinician practice and characteristics surrounding dose reduction of DOACs for extended-phase VTE treatment.We conducted a 16-question REDCap survey between July 14, 2021, and September 13, 2021, among ISTH 2021 Congress attendees and on Twitter. We explored factors associated with dose reduction using logistic regression. We used k-means clustering to identify distinct groups of dose-reduction decision-making. Random forest analysis explored demographics with respect to identified groups.Among 171 respondents, most were attending academic physicians from North America. Clinicians who treated larger volumes of patients had higher odds of dose reduction. We identified five clusters that showed distinct patterns of behavior regarding dose reduction. Cluster 1 rarely dose reduces and likely prescribes rivaroxaban over apixaban; cluster 2 dose reduces frequently, does not consider age when dose-reducing, is least likely to temporarily reescalate dosing, and prescribes apixaban and rivaroxaban equally; cluster 3 dose reduces50% of the time, and temporarily reescalates dosing during increased VTE risk; cluster 4 dose reduces frequently, temporarily reescalates dosing, and is most likely to prescribe apixaban over rivaroxaban; and cluster 5 dose reduces most frequently, and takes the fewest risk factors into consideration when deciding to dose reduce.Most clinicians elect to dose-reduce DOACs for extended-phase anticoagulation. The likelihood of a clinician to dose reduce increases with volume of patients treated. Clinician prescribing patterns cluster around VTE risk factors as well as reescalation during high-risk periods.

Background: Patients with severe hemophilia A and inhibitors are at risk of bleeding during invasive procedures. The standard of care for preventing perioperative bleeding has been replacement therapy with FVIII concentrates or for patients with high-titer inhibitors, bypassing agents. However, there is no consensus on the appropriate management of surgery in patients receiving the novel agent emicizumab. The aim of this study was to demonstrate a case of a patient on emicizumab undergoing major surgery with bypassing agents with preoperative use of the thrombin generation assay (TGA) and thromboelastography (TEG). Methods: We report a patient with hemophilia A with inhibitors who had undergone a total knee replacement while on emicizumab combined with a bypassing agent. We utilized TEG and TGA to determine which bypassing agent to choose as well as to inform about the ideal dose. Results: We elected to use recombinant FVIIa as a bypassing agent for the surgery based upon the TGA results. Conclusion: The TGA can be utilized to support decision-making in patients on emicizumab undergoing major surgery to both predict efficacy and potentially minimize the risk of thrombotic events.

Background Factor VIII activity is determined by directly measuring the aPTT of a patient plasma sample and determining the percent activity from a standard curve generated from plotting the measured clotting time (in seconds) on a semi-log scale vs a known percent activity of the standard at several specific dilution points. Factor VIII activity for the patient samples is then performed on dilutions of patient plasma mixed with equal amounts of plasma deficient in the factor to be measured, and the percent of factor in the patient plasma is calculated from the standard curve by plotting the observed clotting time for a specific dilution of the patient sample. To minimize the potential for under-reporting an activity level or missing the presence of an inhibitor (defined as an antibody directed against Factor VIII), a subjective assessment of parallelism of the patient curve to the standard curve is performed. In absence of an inhibitor the standard and patient curves are parallel to each other with the patient curve’s slope (Ps) similar to the standard curve’s slope (Cs). In patients with an inhibitor, the clotting time is prolonged. Furthermore, with each subsequent dilution, the amount of inhibitor is diluted out, leading to shorter clotting times for subsequent dilutions. In practice this leads to a less steep patient curve slope (Ps) compared to the standard curve slope (Cs) and thus nonparallel lines. Aim Parallelism determination is currently a subjective assessment that leads to increased error in reporting, potential missed evaluation for inhibitors and potential unnecessary testing for inhibitors. We developed an objective and automated tool to assess parallelism as an added screening tool for the presence of a Factor VIII inhibitor. Methods We performed Factor VIII assays (Low Factor VIII assay modification on STA Compact using low curve calibration at 1:6, 1:15 and 1:30 dilution (STA Deficient VIII, Immuno-Depleted Plasma for Factor VIII:C assay by STA. Package insert 26217-revised September 1994)) at appropriate dilutions on Factor VIII deficient hemophilia patients. We examined curves for parallelism by comparing the ratio of the slope of the curve generated from patient dilutions without detectable inhibitor (disease-free state) and the slope of the curve generated from patient dilutions with a known inhibitor (disease state) vs the slope of the standard curve. We confirmed presence of an inhibitor for each patient sample by Bethesda assay utilizing the Nijmegen modification. Results Using a bell curve generated from a parameter simulation of obtained Ps/Cs ratios from screening 21 samples with and without an inhibitor, we determined a ratio of Ps/Cs of 0.45 to be a cut-off that was 100% sensitive and 80% specific for detection of an inhibitor. To confirm the validity of this cut-off, we screened 48 de-identified samples with and without low FVIII inhibitor (Bethesda titers Conclusion We developed and validated a new screening tool for detecting the presence of an inhibitor to Factor VIII during routine FVIII assays. This method has the potential to screen for the detection of inhibitors to other specific clotting factors such as FIX, lupus anticoagulants and the presence of the newer oral anticoagulants which directly inhibit Factor Xa or thrombin. Disclosures No relevant conflicts of interest to declare.

INTRODUCTION The need for factor concentrate (FC) prior to uncomplicated endoscopies in persons with hemophilia (PWH) is not well characterized. Infusion of FC is time consuming, costly, and increases risk of inhibitor development. There is no general consensus on management of PWH undergoing endoscopies and the incidence of bleeding is not well established. At our Hemophilia Treatment Center (HTC), our standard of care (SOC) for many years was to manage PWH with an infusion of FC prior to any endoscopy for any indication regardless of severity of hemophilia. In November 2017, we changed our SOC for persons with mild hemophilia (factor VIII or IX >5%) to use only antifibrinolytics pre-endoscopy for those in whom no procedural intervention was anticipated, followed by infusion of FC post-endoscopy if a biopsy or polypectomy was performed or if there was bleeding associated with the procedure. Persons with moderate or severe hemophilia (factor VIII or IX ≤5%) continued to receive infusion of FC prior to all endoscopies. We report our experience managing PWH of all severities undergoing EGD and colonoscopy between 2008 and 2019. We compare incidence of bleeding, cost of medical care, and outcomes before and after the change in SOC for persons with mild hemophilia. METHODS Subjects were identified from a database of patients treated at our HTC. Those with factor VIII inhibitors were excluded. Cost of FC and antifibrinolytics were determined from average wholesale pharmacy prices in 2019. Gastrointestinal (GI) bleeding was assessed by review of medical records within 30 days after endoscopy. Documentation of any patient or clinician reported hematemesis or rectal bleeding, requiring or not requiring medical attention or treatment, was recorded as GI bleeding. Adequacy of hemostasis, as assessed by review of medical records, was classified as poor if the subject had unexpected hypotension, required intensive or intermediate care, or transfusion; fair if unplanned doses of FC were required, but the event did not meet criteria for poor hemostasis; and excellent for events that did not meet any of the above criteria. RESULTS Sixty-seven subjects met eligibility criteria; 12 were excluded due to inadequate records of endoscopies. Data were collected on 110 endoscopies performed in 55 subjects, range 1 to 5 per subject. Subject demographics are noted in Table 1. Endoscopy characteristics and outcomes are shown in Table 2. GI bleeding occurred within 30 days after 10 endoscopies prior to the change in SOC. This included 7 endoscopies performed because of preexisting bleeding due to esophageal varices, or gastric, duodenal, or colonic ulcers. In addition, post endoscopy bleeding occurred in one patient after colonoscopy following the use of hot snare cautery resection of polyps, and in another after resection of a rectal polyp by cold snare following an EGD with colonoscopy. No GI bleeding occurred within 30 days of the endoscopies performed after we changed our SOC. FC was used prior to 97.1% of 68 endoscopies performed before we changed our SOC, compared to 73.8% of 42 endoscopies performed after. Hospital stay of 2 days was equivalent across all groups when subjects with preexisting bleeding were excluded. Hemostasis was excellent in 79.4% of endoscopies before we changed our SOC and 100% of endoscopies after. Endoscopy characteristics and outcomes in subjects with mild hemophilia before and after we changed our SOC are shown in Table 3. Reasons for using FC pre-endoscopy in these subjects included preexisting bleeding, planned intervention, or anticipated complex procedure. Treatment costs in subjects with mild hemophilia are noted in Table 4. CONCLUSION We describe our HTC's experience with endoscopies in PWH. In our cohort of persons with mild hemophilia, we demonstrate that the use of antifibrinolytics rather than FC pre-endoscopy did not result in any difference in adequacy of hemostasis or development of GI bleeding within 30 days post endoscopy. This practice was associated with significant cost-savings and no difference of hospitalization rate or length of stay. We conclude that persons with mild hemophilia, who are not actively bleeding or undergoing complex procedures, can be safely managed with antifibrinolytics alone pre-endoscopy. However, patient-specific and procedure-specific factors must be considered when determining the appropriate pre-endoscopy treatment for bleeding prophylaxis in PWH. Disclosures Eyster: Novo Nordisk: Research Funding; SPARK: Research Funding; Sanofi: Research Funding; Baxalta/Shire: Research Funding.

We studied the course of chronic HCV infections in a cohort of 222 persons with hemophilia (PWH) and von Willebrand disease followed at our center since 1973. Twenty two (10%) developed end stage liver disease (ESLD). Forty years after HCV infection, cumulative incidence of ESLD was 12.3% and overall survival was 45.5%. Those who were infected with HCV only (n = 100) had a survival of 75.2%, while those infected with HIV (n = 122) had a survival of 24% (P 95% of individuals. Am. J. Hematol. 91:E335-E340, 2016. © 2016 Wiley Periodicals, Inc.

INTRODUCTION: Dehydrated hereditary stomatocytosis (DHSt) is an autosomal dominant red blood cell membrane disorder characterized by hemolytic anemia and splenomegaly. DHSt has an estimated incidence of 1:50,000 births, and the degree of anemia varies within and between families. Although transfusion support during childhood is not uncommon, continued requirement into adulthood is rare. The most frequent cause of DHSt is a gain-of-function mutation of the PIEZO1 gene, leading to delayed channel inactivation that results in a monovalent cation leak and an increase in intracellular calcium (Ca2+). Many of these patients develop recurrent thromboses post splenectomy. Other DHSt patients have mutations in KCNN4, which encodes the Gardos channel, with mutations causing increased Ca2+ sensitivity and potassium efflux. To our knowledge, 42 patients from ten families have been described with four distinct KCNN4 mutations: Arg352His, Val282Met or Val282Glu, and a 28bp deletion encompassing the exon-intron 7 junction. We report herein the eighth family with the Arg352His locus mutation. CASE REPORT: Five subjects from a single family were enrolled in this study (affected proband, unaffected husband, two affected children, and an unaffected grandchild). The proband has had hemolytic anemia since childhood. She had undergone splenectomy at age three and cholecystectomy at eight. Her anemia persisted, with a mean hemoglobin (Hgb) of 10.5g/dL and a reticulocyte count (retic ct) of 12.6%, with no need for transfusion support or iron chelation therapy (Table 1). Extensive testing revealed slightly decreased osmotic fragility and mildly elevated intracellular sodium concentration, of 19.4 mEq/L. Her daughter has mild splenomegaly and anemia, with a mean Hgb of 11.1g/dL and retic ct of 6.7%, whereas her son has more severe disease, with a mean Hgb of 9.3g/dL and a retic ct of >22%. He underwent splenectomy as a teenager for immune thrombocytopenia, and has required chelation therapy with deferasirox since age 31, when his ferritin rose to >1000ng/mL. Similar to results reported by others, splenectomy did not alter the severity of hemolysis in either the proband or her son, and neither developed thrombotic complications 57 and 7 years post splenectomy, respectively. The proband's daughter has a child with a normal Hgb, and is presumed unaffected. Stomatocytes were rarely seen on the peripheral blood smears of the proband and her children. METHODS: A clinically available 39 gene hemolytic anemia panel on the proband failed to identify the mutation underlying this disorder. We therefore performed whole exome sequencing on all five family members. We prioritized the analysis of 23 additional genes that are included in hemolytic anemia panels from two other reference laboratories and are involved in disorders of red blood cell membrane or cytoskeletal proteins of potential clinical relevance to the study population. RESULTS: A single missense mutation, Arg352His within KCNN4, was identified in all three affected individuals. This heterozygous mutation was present in the proband and her affected two children, and absent in her unaffected grandchild and husband. CONCLUSIONS: To our knowledge, this Pennsylvania family is only the eleventh described to have DHSt secondary to a KCNN4 mutation. This disorder is likely much more prevalent than reported, due to the rarity of stomatocytes on peripheral blood smears, the omission of the KCNN4 gene from hemolytic panels offered by some reference laboratories, and variable clinical presentation. KCNN4 mutations should be investigated if other causes are not identified in patients with lifelong hemolytic anemia suspected of having a red cell membrane protein or cytoskeletal disorder. Appropriate diagnosis may allow severely affected patients to be considered for treatment with the experimental Gardos channel inhibitor senicapoc. Additionally, mutational diagnosis is especially important when considering the adverse outcomes post splenectomy in PIEZO1 as compared to KCNN4 mutations. Disclosures Eyster: SPARK:Research Funding;Sanofi:Research Funding;Novo Nordisk:Research Funding;Baxalta/Shire:Research Funding.

Men with hemophilia were initially thought to be protected from cardiovascular disease (CVD), but it is now clear that atherothrombotic events occur. The primary objective of the CVD in Hemophilia study was to determine the prevalence of CVD and CVD risk factors in US older men with moderate and severe hemophilia and to compare findings with those reported in age-comparable men in the Atherosclerosis Risk in Communities (ARIC) cohort. We hypothesized if lower factor levels are protective from CVD, we would see a difference in CVD rates between more severely affected and unaffected men. Beginning in October 2012, 200 patients with moderate or severe hemophilia A or B (factor VIII or IX level ≤ 5%), aged 54 to 73 years, were enrolled at 19 US hemophilia treatment centers. Data were collected from patient interview and medical records. A fasting blood sample and electrocardiogram (ECG) were obtained and assayed and read centrally. CVD was defined as any angina, any myocardial infarction by ECG or physician diagnosis, any self-reported nonhemorrhagic stroke or transient ischemic attack verified by physicians, or any history of coronary bypass graft surgery or coronary artery angioplasty. CVD risk factors were common in the population. Compared with men of similar age in the ARIC cohort, patients with hemophilia had significantly less CVD (15% vs 25.8%; P < .001). However, on an individual patient level, CVD events occur and efforts to prevent cardiovascular events are warranted. Few men were receiving secondary prophylaxis with low-dose aspirin, despite published opinion that it can be used safely in this patient population.

The frequency of anemia, iron deficiency, and the long-term need for IV iron following Roux-en-y gastric bypass (RYGB) surgery has not been well characterized. Three-hundred and nineteen out of 904 consecutive subjects who underwent RYGB at Penn State Hershey Medical Center from 1999 to 2006 met the inclusion criteria for a preoperative complete blood count (CBC) and at least one CBC >6 months following surgery. Cumulative incidence of anemia 7 years post procedure was 58%. Menstruation status and presence of preoperative anemia were predictive of anemia by univariate analysis and multivariable Cox regression ( P= 0.0014 and 0.044, respectively). Twenty-seven subjects, primarily premenopausal women, representing 8.5% of the cohort and 22% of the 122 anemic subjects, needed intravenous (IV) iron a mean of 51 months postoperatively for anemia unresponsive or refractory to oral iron. The risk for development of anemia necessitating IV iron therapy following RYGB is highest in menstruating women and continues to increase for many years, even in post-menopausal women. Well-designed prospective studies are needed to identify the incidence of iron deficiency anemia and the patient populations at increased risk for requiring IV iron replacement after RYGB surgery.

The development of neutralizing antibodies, referred to as inhibitors, against factor VIII is a major complication associated with FVIII infusion therapy for the treatment of hemophilia A (HA). Previous studies have shown that a subset of HA patients and a low percentage of healthy individuals harbor non-neutralizing anti-FVIII antibodies that do not elicit the clinical manifestations associated with inhibitor development.To assess HA patients' anti-FVIII antibody profiles as potential predictors of clinical outcomes.A fluorescence immunoassay (FLI) was used to detect anti-FVIII antibodies in 491 samples from 371 HA patients.Assessments of antibody profiles showed that the presence of anti-FVIII IgG1 , IgG2 or IgG4 correlated qualitatively and quantitatively with the presence of an FVIII inhibitor as determined with the Nijmegen-Bethesda assay (NBA). Forty-eight patients with a negative inhibitor history contributed serial samples to the study, including seven patients who had negative NBA titers initially and later converted to being NBA-positive. The FLI detected anti-FVIII IgG1 in five of those seven patients prior to their conversion to NBA-positive. Five of 15 serial-sample patients who had a negative inhibitor history and had anti-FVIII IgG1 later developed an inhibitor, as compared with two of 33 patients with a negative inhibitor history without anti-FVIII IgG1 .These data provide a rationale for future studies designed both to monitor the dynamics of anti-FVIII antibody profiles in HA patients as a potential predictor of future inhibitor development and to assess the value of the anti-FVIII FLI as a supplement to traditional inhibitor testing.

Men and boys who present with bleeding associated with low factor VIII levels and normal von Willebrand studies are assumed to have hemophilia A until proven otherwise. However, routinely available coagulation assays cannot easily distinguish mild hemophilia A from the 2N variant of von Willebrand disease. We present a case that highlights the difficulties of recognizing this diagnosis, the role of genetic testing, and the identification of a 2N variant that has not been previously described.

Introduction: While Factor VIII (FVIII) activity correlates with phenotype in most Hemophilia A (HA) males, the study of rare individuals with discrepant bleeding phenotypes can provide insight into F8 genotype-bleeding phenotype relationships, refine HA classification and improve clinical management. We present an adult male subject who carries a F8 missense mutation, c.5999G>C (p.Gly2000Ala), but remarkably has had no bleeding despite numerous challenges, including multiple surgeries. To resolve genotype-phenotype discrepancy, we evaluated F8 transcripts and screened for additional gene variants by exome sequencing. Methods: We performed clinical laboratory assays including one-stage FVIII:C and chromogenic FVIII assays, FVIII antigen ELISA, VWF antigen and activity assays, and von Willebrand Disease (VWD) Type 2 Normandy binding activity assay. The F8 c.5999G>C mutation is proposed to affect F8 splicing, and result in a transcript that deletes exon 19. To evaluate F8 splice variants, quantitative reverse transcription polymerase chain reaction (qRT-PCR) was performed using primers that distinguish FVIII isoforms. Samples were normalized relative to a housekeeping gene, GAPDH. Normalized relative fold expression for each primer pair was calculated by the 2−ΔΔCT Livak method. Whole exome sequencing was performed at 80X coverage. Sequencing reads were mapped to the GRCh37/hg19 reference genome with BWA-MEM, and high-quality variant calls were made using GATK4.0 with annotation based on refSeq 1.9, employing best practice recommendations. Results: Clinical assays confirmed reduced FVIII activity levels consistent with mild HA. By qRT-PCR, the F8 mutation results in a partial splicing defect. Exon 19 deleted transcripts predominate, but are likely not secreted. Indeed, the reduced levels of full-length F8 isoform closely mirror measured FVIII protein and activity levels. These findings explain FVIII levels, but to do not explain the absent phenotype. Exome sequencing analysis first confirmed the F8 mutation and excluded other genetic causes of FVIII deficiency. Additionally, we excluded thrombophilic mutations proposed to attenuate bleeding severity in HA, including Factor V Leiden (c.1601G>A, p. R534Q) and the c.*97G>A (p.G20210A) mutation in the F2 gene encoding prothrombin. To identify additional variants that may modify hemostasis, we then screened for non-pathogenic variants in 96 genes that are known to be mutated in bleeding disorders or that alter FVIII clearance or half-life. Of 352 single nucleotide variants (SNVs) in these genes, we prioritized 10 non-synonymous variants with annotated effects on hemostasis. Of these, four are associated with functional effects that would potentiate bleeding phenotype in coagulation factor deficiency. In contrast, five variants are associated with prothrombotic effects and are strong candidates for ameliorating bleeding. Of these, two non-pathogenic VWF variants, rs1063856 and rs7962217, have been shown to increase FVIII. Three other variants in F5 reside within the R2 haplotype. R2 has effects in the absence of Factor V Leiden and can increase thrombin generation and FVIII levels. Conclusion: The identification of multiple variants that are expected to attenuate bleeding suggests that a single gene variant is unlikely to ameliorate bleeding phenotype in this subject, particularly in the presence of additional variants that promote bleeding. These results suggest evaluation of non-pathogenic variants in non-F8 genes may further explain other cases of discrepant HA not resolved by clinical assays. While these new modulators of HA phenotype require functional confirmation, they provide new avenues for therapeutic development. Disclosures Eyster: Bayer: Other: research support; Baxalta: Other: research support; Shire: Other: research support; NovoNordisk: Other: research support; SPARK: Other: research support.

Gene transfer for hemophilia A offers the potential for a one-time disease altering treatment, eliminating the risk of bleeds while freeing patients from the burden of lifelong chronic therapy. SPK-8011 consists of a bioengineered AAV capsid expressing B domain-deleted factor VIII (FVIII) under the control of a liver-specific promoter. In pre-clinical studies, we showed a dose-dependent increase in circulating FVIII levels in non-human primates infused with SPK-8011. We conducted a Phase I/II study of SPK-8011 in 12 men (ages 18-52 years) with severe (n=11) or moderately severe (n=1) hemophilia A. Prior to gene therapy, 8/12 subjects were on prophylaxis, and 4/12 received on-demand treatment. Subjects were enrolled in 1 of 3 dose cohorts, 5E11 vg/kg (n=2), 1E12(n=3), or 2E12(N=7). Safety analysis showed no inhibitor formation. A single serious adverse event (SAE) was reported, associated with an immune response to AAV capsid characterized by simultaneous decline in FVIII, transaminase elevation peaking at Grade 2, and development of positive IFN-g ELISPOTs to capsid was observed beginning at week 6.5 after vector infusion. The asymptomatic transaminase elevation did not respond promptly to initiation of oral steroids and the subject received two infusions of IV methylprednisolone in hospital, thereby fulfilling SAE criteria. The SAE has resolved. All vector doses led to expression of FVIII levels adequate to prevent bleeding and allow cessation of prophylaxis. Across the 12 subjects at 3 doses, there was a 97% reduction in annualized bleeding rate (ABR), and a 97% reduction in annualized infusion rate (AIR). In the 5E11 dose cohort, mean FVIII levels beginning 12 weeks post vector infusion are 13%, with no bleeding events, no elevated transaminase levels, no use of steroids, and stable FVIII expression out to 66 weeks (ongoing). In the 1E12 dose cohort, mean FVIII levels are 15% beginning at 12 weeks post-infusion and stable out to 46 weeks (ongoing). The first subject in the 1E12 dose infused a single dose of factor concentrate for a spontaneous joint bleed at day 159, and the second received multiple infusions for a traumatic bleed beginning at day 195. Declining FVIII levels triggered initiation of a course of tapering steroids in both subjects, at 12 and 7 weeks post vector infusion respectively, which led to stabilization of FVIII levels. The third subject has had no bleeding and did not receive factor infusions or steroids. In the 2E12 (highest) dose cohort, 5/7 subjects currently have FVIII levels 16-49%; their mean FVIII level beginning 12 weeks post-infusion is 30%. No bleeds have been reported among these subjects beginning 4 weeks post vector infusion. Additionally, 5/7 subjects in the 2E12 dose cohort received a course of steroids, initiated at 6-11 weeks post vector infusion, for one or more of the following: declining FVIII levels, rise in ALT above subject baseline, or elevated IFN-g ELISPOTs to AAV capsid. Steroid initiation normalized ALT levels and extinguished the ELISPOT signal in all cases; 2 subjects showed limited stabilization of FVIII levels, which fell to Our data indicate that the kinetics of SPK-8011 expression are similar to those observed with investigational SPK-9001 for hemophilia B. All subjects demonstrated durable transgene expression for up to 66 weeks post vector administration (data cutoff 7/13/18). On cumulative follow up of 345 weeks, SPK-8011 demonstrated a favorable safety profile with no evidence of FVIII inhibitor formation, a single SAE, and 2/12 subjects who experienced ALT elevation above the upper limit of normal that resolved with steroid initiation. Data from the 5E11 (lowest) dose cohort are consistent with published natural history data indicating FVIII:C 12% is adequate to prevent spontaneous bleeding events. Given that 2 subjects in the 2E12 dose cohort lost some FVIII expression, which then stabilized on steroids, and 5/7 subjects in this cohort required steroids, prophylactic steroids may be warranted. We conclude that infusion of SPK-8011 in 12 subjects with severe or moderately severe hemophilia A resulted in safe, durable, dose-dependent FVIII expression resulting in an excellent preliminary efficacy profile with an overall 97% reduction in ABR and AIR. Disclosures High: Spark Therapeutics: Employment, Equity Ownership, Patents & Royalties. George:University of Pennsylvania: Equity Ownership; Pfizer: Consultancy. Ragni:CSL Behring: Research Funding; Alnylam: Membership on an entity's Board of Directors or advisory committees, Research Funding; Sangamo: Research Funding; Shire: Research Funding; Biomarin: Membership on an entity's Board of Directors or advisory committees, Research Funding; Novo Nordisk: Research Funding; Bioverativ: Consultancy, Research Funding; MOGAM: Membership on an entity's Board of Directors or advisory committees; SPARK: Consultancy, Research Funding. Croteau:Novo Nordisk: Consultancy; Octapharma: Consultancy, Honoraria, Research Funding; Pfizer: Research Funding; Spark Therapeutics: Research Funding; Tremeau Pharmaceuticals: Consultancy; Genetech: Consultancy, Research Funding; CSL-Behring: Consultancy; Catalyst Biosciences: Consultancy; Bioveritiv: Consultancy; Biomarin: Consultancy; Bayer: Consultancy; Baxalta/Shire: Consultancy, Research Funding. Joseney-Antoine:Spark Therapeutics: Employment. Macdougall:Spark Therapeutics: Employment. Tompkins:Spark Therapeutics: Employment. Hait:Spark Therapeutics: Employment. Couto:Spark Therapeutics: Employment. Bassiri:Spark Therapeutics: Employment. Valentino:Spark Therapeutics: Employment. Carr:Spark Therapeutics: Employment. Hui:Spark Therapeutics: Employment. Wachtel:Spark Therapeutics: Employment. Takefman:Spark Therapeutics: Employment. Mingozzi:Spark Therapeutics, Inc.: Employment. Anguela:Spark Therapeutics, Inc.: Employment. Reape:Spark Therapeutics: Employment.

Introduction: Over a three-year period, U.S. men with hemophilia were found to be 50 times more likely to die from renal disease than the general population (SMR 50; 95% CI 26.8-92.8) (Soucie et al., Blood 2000). Despite this finding, data regarding chronic kidney disease (CKD) and its risk factors in patients with hemophilia remain limited. The objective of this study is to determine the prevalence of CKD and CKD risk factors among older men with moderate and severe hemophilia. Methods: This CKD cohort study is an extension of a U.S. national study sponsored by the American Thrombosis and Hemostasis Network (ATHN). The study, entitled ATHN 1: A Cross-Sectional Analysis of Cardiovascular Disease (CVD) in the Hemophilia Population, began enrollment in 10/2012. Inclusion criteria are men with moderate or severe congenital hemophilia A or B (FVIII or IX level ≤ 5%), age 54-73. Men with an additional bleeding disorder (besides liver dysfunction) were excluded. In this extension study, CKD risk factors, historical creatinine levels, and history of renal events were obtained from patient interview and chart review after obtaining informed consent. Glomerular filtration rate (GFR) values were calculated using the CKD-EPI equation and compared to values in the general population using the NHANES dataset (Coresh et al., JAMA 2007). CKD is defined as the presence of either kidney damage or decreased kidney function with GFR < 60 ml/min/1.73 m2 for ≥ 3 months, irrespective of cause. Results: As of 6/30/2018, 134/200 planned subjects have been enrolled and interim analysis on 134 subjects from 18 U.S. hemophilia treatment centers (HTCs) is presented here. The majority were white (119; 88.8%) or African-American (13; 9.7%). Mean age was 64 years (SD: 5; range: 56-77). Most used factor on demand, with only 38.8% (52/134) on prophylaxis, defined as ≥2 doses of FVIII or ≥ 1 dose FIX/week. Four (3.0%) had a current inhibitor. Viral infection was common; 28.4% currently had hepatitis C, and 19.4% HIV. Hypertension (HTN) was reported in 51.5% of subjects, 14.9% diabetes mellitus (DM); and average BMI was 28.2 kg/m2 (36.6% obese). 11.6% (16/134) were found to have CVD (defined as angina, MI, TIA, or ischemic or embolic stroke). Acute kidney injury was common. Fasting blood work showed an abnormally elevated creatinine in 26.9% subjects (mean 1.1 mg/dl, SD 0.4). Mean historical maximum creatinine reported in the cohort was 1.0 (range 0.5-4.8), with mean GFR 67 (range 11-126). 11.4% (13/114) met the definition of CKD. Stages of CKD by GFR in the hemophilia cohort were similar to the NHANES general population (p=0.561). See Table 1 for subject reported history of renal events, and Table 2 for specific diagnoses of renal disease. In our cohort, hemophilia subjects with CKD tended to have a diagnosis of intrinsic kidney disease (60.0% vs 11.6%, p=0.02), and non-significantly tended to have DM (23.1% vs 12.8%), age ≥65 years (21.1% vs 9.4%), and HTN (18.0% vs 9.8%) compared to subjects without CKD. No other significant trends were identified, including no association with CVD, HCV, HIV, BMI, or hematuria. Conclusions: In this interim analysis of an ongoing national prospective cohort study, older men with moderate to severe hemophilia commonly report risk factors for CKD, including HTN (51.5%), DM, viral infection, and potential renal damaging medication use. Only 11.6% had CVD. Urological symptoms were also common, including hematuria and obstructive symptoms with urination. In our cohort, 11.4% met the definition of CKD, defined as the presence of either kidney damage or GFR < 60 ml/min/1.73 m2 for ≥ 3 months. The distribution of GFR values appeared similar to the general population. As with risk factors associated with CKD in the general population, diagnosis of intrinsic kidney disease was significantly associated with CKD in hemophilia subjects, with non-significant trend for increased DM, older age, and HTN compared to subjects without CKD. It is reassuring that the prevalence of CKD does not appear to be increased in men with hemophilia compared to the general population, despite a known and unexplained high incidence of HTN in the hemophilia population. We plan to formally compare the prevalence of CKD and CKD risk factors with similarly aged men in the ARIC database once enrollment is complete, as understanding the risk factors that contribute to CKD is essential to halt its progression. Disclosures Sood: Bayer: Research Funding. Shapiro:BioMarin: Research Funding; Shire: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Novo Nordisk: Membership on an entity's Board of Directors or advisory committees, Research Funding; Prometic Life Sciences: Consultancy, Research Funding; Kedrion Biopharma: Consultancy, Research Funding; Sangamo Biosciences: Consultancy; Octapharma: Research Funding; OPKO: Research Funding; Daiichi Sankyo: Research Funding; Bayer Healthcare: Other: International Network of Pediatric Hemophilia; Bio Products Laboratory: Consultancy; Genetech: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Bioverativ, a Sanofi Company: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding. Kessler:Biomarin: Research Funding; Dimension Advisory boards: Membership on an entity's Board of Directors or advisory committees; DSMB: Membership on an entity's Board of Directors or advisory committees; Sangamo: Research Funding; Novo Nordisk: Honoraria, Research Funding; Octapharma: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Genentech: Research Funding; Pfizer: Honoraria, Membership on an entity's Board of Directors or advisory committees; Bayer: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding. Key:UniQure BV: Research Funding. Quon:Bioverativ, a Sanofi Company: Speakers Bureau; NovoNordisk: Consultancy, Speakers Bureau; Bayer: Consultancy; Shire: Speakers Bureau; Genetech: Consultancy, Speakers Bureau; Octapharma: Consultancy. Manco-Johnson:Novo Nordisk: Honoraria; CSL Behring: Honoraria; Bayer AG: Honoraria, Research Funding; Biogentek: Honoraria; Baxalta, now part of Shire: Honoraria. Cuker:Kedrion: Membership on an entity's Board of Directors or advisory committees; Genzyme: Consultancy; Spark Therapeutics: Research Funding; Synergy: Consultancy. Ragni:Novo Nordisk: Research Funding; Shire: Research Funding; MOGAM: Membership on an entity's Board of Directors or advisory committees; Sangamo: Research Funding; Alnylam: Membership on an entity's Board of Directors or advisory committees, Research Funding; Sangamo: Research Funding; SPARK: Consultancy, Research Funding; CSL Behring: Research Funding; Biomarin: Membership on an entity's Board of Directors or advisory committees, Research Funding; Bioverativ: Consultancy, Research Funding; SPARK: Consultancy, Research Funding. von Drygalski:UniQure BV, Bayer, Bioverativ/Sanofi, Pfizer, Novo Nordisk, Biomarin, Shire, CSL Behring: Consultancy. Kouides:UniQure: Other: DSMB; Octapharma: Research Funding. Escobar:Bayer, CSL Behring, Genentech, Hemabiologics, Kedrion, Novo Nordisk, Octapharma, Pfizer and Shire: Consultancy; Pfizer: Research Funding. Wang:Daiichi Sankyo: Consultancy, Other: Travel. Konkle:Shire: Research Funding; Genentech: Consultancy; Bioverativ: Research Funding; BioMarin: Consultancy; Pfizer: Research Funding; Gilead: Consultancy; Spark: Consultancy, Research Funding; CSL Behring: Consultancy; Sangamo: Research Funding.

Introduction: Although FVIII activity (FVIII:C) correlates with phenotype in most Hemophilia A males, some have bleeding phenotypes milder than predicted by FVIII:C. Analysis of these individuals may provide additional insight into the relationship between F8 genotype and bleeding phenotype, identify additional factors that modulate FVIII, refine the classification of Hemophilia A and improve clinical management. We present a male subject who carries a c.5999G>C mutation in F8. This mutation has been reported in an individual with a severe bleeding phenotype in the EAHAD F8 Variant Database (www.factorviii-db.org). Our subject was serendipitously diagnosed at age 80, and has never had abnormal bleeding despite numerous challenges, including multiple dental extractions, adult circumcision, internal hemorrhoid ligation and cardiac catheterization. In an effort to resolve this genotype-phenotype discrepancy, we performed FVIII and von Willebrand Factor (VWF) assays, quantitated the extent of alternative splicing in F8, and screened for mutations in additional genes known to impact hemostasis. We validated a partial splicing defect that is consistent with mild Hemophilia A, but surprisingly cannot explain the lack of a bleeding phenotype. Methods: Peripheral blood was tested by one-stage FVIII:C and chromogenic FVIII assays, FVIII antigen ELISA, VWF antigen and activity assays, and von Willebrand Disease (VWD) Type 2 Normandy binding assay. To evaluate F8 splice variants, quantitative RT-PCR (qRT-PCR) was performed on RNA isolated from peripheral blood using primers that distinguish FVIII isoforms. Samples were tested in quadruplicate, with sample input normalized relative to GAPDH. Screening for exonic mutations in F8, F5, F2 and VWF genes was undertaken by Next Generation Sequencing (NGS). Results and Conclusions: In contrast to the previously reported severe c.5999G>C patient, this subject has reduced FVIII activity consistent with mild Hemophilia A measured by both FVIII activity assays. FVIII antigen assessment by ELISA gave similar results (0.35 IU/mL). To better understand FVIII:C levels in this individual, we sought to characterize F8 transcript isoforms. The c.5999G>C resides within the splice acceptor site of exon 19 and may result in two FVIII isoforms. One isoform produces a full-length FVIII containing a missense mutation (p.Gly2000Ala). The second isoform skips the 117 bp exon 19, but is predicted to retain an open reading frame; however, its location in the A3 domain likely disrupts secretion or function (Donadon et al., Haematologica 2018). The relative abundance of F8 variants evaluated by qRT-PCR demonstrated that despite a modest overall increase in F8 transcript relative to a normal control (1.6 fold higher expression of F8 at exon 18), the predominant isoform in this subject lacks exon 19, resulting in substantial depletion of the full-length isoform (0.3 relative to a normal control). These data indicate the full-length protein likely retains function, with transcript levels that explain residual FVIII activity. In an effort to explain absent bleeding phenotype, NGS was performed. The c.5999G>C F8 mutation was confirmed, while the thrombophilic Factor V Leiden and Prothrombin G20210A mutations were excluded. Sequencing also excluded mutations implicated in Type 2 Normandy VWD. Additional clinical testing may provide an explanation. VWF was elevated as indicated by VWF antigen and Ristocetin cofactor activity (>400%, >100%, respectively). Furthermore, VWF:FVIII binding activity by ELISA showed a higher than normal binding ratio (>1.42). VWF multimer analysis indicated normal distribution. Therefore, we propose elevated VWF abrogates this subject's bleeding phenotype. Altogether, our study expands our understanding of the functional consequences of this mutation by demonstrating that this mutation can result in FVIII levels predicting mild Hemophilia A. This case also underscores the complexities of predicting phenotype for mutations that result in partial splicing defects. Perhaps more intriguing is the lack of bleeding phenotype in this elderly individual with numerous hemostatic challenges, that might be related to increased VWF. These data support additional studies investigating the role of VWF in ameliorating bleeding risk in Hemophilia A. Disclosures No relevant conflicts of interest to declare.

SummaryBolus infusion (BI) recombinant factor VIIa (rFVIIa) administration is safe and effective in the surgical management of haemophilia patients with inhibitors but has not been compared directly with continuous infusion (CI). We conducted an open-label, randomized, multicenter trial comparing the efficacy and safety of rFVIIa administered by BI or CI for the surgical management of haemophilia A or B patients with inhibitors to FVIII or FIX. Safety was compared with that of a control group of noninhibitor patients receiving FVIII or FIX concentrates for major surgery. All inhibitor subjects received an initial bolus dose of 90 μg/kg rFVIIa and were then randomly assigned to BI (n=12) or CI (n=12). The BI group received 90 μg/kg rFVIIa every two hours (h) during surgery through day 5, then every four hours for days 6–10. The CI group received 50 μg/kg/h rFVIIa through day 5, then 25 mg/kg/h for days 6–10. The control group (n=12) received FVIII or FIX per institutional protocols. Twenty-two major surgeries included orthopedic procedures on the knee (n=13), hip (n=3), and abdominal/pelvis procedures (n=4). One patient with an autoimmune FVIII inhibitor randomized to the BI arm was excluded from efficacy analysis. Haemostatic efficacy of rFVIIa in each group was comparable: effective in 8/11 and 9/12 subjects in the BI and CI arms, respectively, and ineffective in three subjects in each arm. Serious adverse events were related to continued or increased bleeding. In conclusion, haemostatic efficacy and safety of BI and CI of rFVIIa are comparable for the surgical management of haemophilia subjects with inhibitors.

SummaryPerioperative clotting factor replacement is administered to reverse the inherent haemostatic defect in persons with haemophilia (PWH), potentially increasing their risk for developing venous thromboembolism (VTE) postoperatively. It was our objective to determine the prevalence of VTE in PWH undergoing total hip or knee arthroplasty (THA, TKA). Patients with haemophilia A or B who underwent THA or TKA were enrolled in this prospective, multicentre observational cohort study. Lower extremity venous duplex ultrasound was performed prior to surgery and 4–6 weeks after surgery. Eleven centres enrolled 51 subjects, 46 of whom completed the study. Six subjects (13.0 %) were treated with bypass agents perioperatively; the remaining 40 subjects received factor VIII or IX replacement. Intermittent pneumatic compression devices were utilised postoperatively in 23 subjects (50 %), and four subjects (8.7 %) also received low-molecular-weight heparin prophylaxis. One subject (2.2 %) with moderate haemophilia A was diagnosed with symptomatic distal deep-vein thrombosis (DVT) on day 6 following TKA. One subject (2.2 %) with severe haemophilia A was diagnosed with pulmonary embolism on day 9 following bilateral TKA. No subjects had asymptomatic DVT. Eighteen subjects (39.1 %) had major bleeding, and three subjects (6.5 %) experienced minor bleeding. The observed prevalence of ultrasound-detectable, asymptomatic DVT in PWH following TKA or THA in this study was low, but the incidence of symptomatic VTE (4.3 %, 95 % CI, 0.5–14.8 %) appeared similar to the estimated incidence in the general population without thromboprophylaxis.

Before the mid-1980s, haemophilia often was unknowingly treated with contaminated plasma products, resulting in high rates of human immunodeficiency virus (HIV-1), hepatitis C virus (HCV) and hepatitis B virus (HBV) infections. To estimate the impact of these infections, a new cohort was established. All HCV-seropositive patients, age 13-88 years, at 52 comprehensive haemophilia treatment centres were eligible. Cross-sectional data collected during April 2001 to January 2004 (median June 2002) were analysed. Plasma HIV-1 and HCV RNA were quantified by polymerase chain reaction. Highly active antiretroviral therapy (HAART) was defined as use of at least three recommended medications. Among 2069 participants, 620 (30%) had HIV-1. Of 1955 with known HBV status, 814 (42%) had resolved HBV and 90 (4.6%) were HBV carriers. Although 80% of the HIV-1-positive participants had > or = 200 CD4+ cells microL(-1), only 59% were on HAART. HIV-1 RNA was undetectable in 23% of those not taking antiretroviral medications. Most (72%) participants had received no anti-HCV therapy. HCV RNA was detected less frequently (59%) among participants treated with standard interferon plus ribavirin (P = 0.0001) and more frequently among HIV-1-positive than HIV-1-negative participants (85% vs. 70%, P < 0.0001). HIV-1-positive participants were more likely to have pancytopenia and subclinical hepatic abnormalities, as well as persistent jaundice, hepatomegaly, splenomegaly and ascites. HAART recipients did not differ from HIV-negative participants in the prevalence of ascites. The clinical abnormalities were more prevalent with older age but were not confounded by HBV status or self-reported alcohol consumption. Eleven participants presented with or previously had hepatocellular carcinoma or non-Hodgkin lymphoma. Although prospective analysis is needed, our data reveal the scale of hepatic and haematological disease that is likely to manifest in the adult haemophilic population during the coming years unless most of them are successfully treated for HIV-1, HCV or both.

Inhibitors are a rare but serious complication of treatment of patients with haemophilia. Phase III clinical trials enrol too few patients to adequately assess new product inhibitor risk. This project explores the feasibility of using a public health surveillance system to conduct national surveillance for inhibitors. Staff at 17 U.S. haemophilia treatment centres (HTC) enrolled patients with haemophilia A and B into this prospective study. HTC staff provided detailed historic data on product use and inhibitors at baseline, and postenrolment patients provided monthly detailed infusion logs. A central laboratory performed inhibitor tests on blood specimens that were collected at baseline, annually, prior to any planned product switch or when clinically indicated. The central laboratory also performed genotyping of all enrolled patients. From January 2006 through June 2012, 1163 patients were enrolled and followed up for 3329 person-years. A total of 3048 inhibitor tests were performed and 23 new factor VIII inhibitors were identified, 61% of which were not clinically apparent. Infusion logs were submitted for 113,205 exposure days. Genotyping revealed 431 distinct mutations causing haemophilia, 151 of which had not previously been reported elsewhere in the world. This study provided critical information about the practical issues that must be addressed to successfully implement national inhibitor surveillance. Centralized testing with routine monitoring and confirmation of locally identified inhibitors will provide valid and representative data with which to evaluate inhibitor incidence and prevalence, monitor trends in occurrence rates and identify potential inhibitor outbreaks associated with products.

Persons with hemophilia or other HIV-1 risk factors may be more likely to have idiopathic CD4+ T-lymphocytopenia (ICL). We determined the frequency of ICL in prospectively followed cohorts of HIV-1 seronegative hemophilic men and seronegative female sex partners of HIV-1 infected hemophilic men, and examined factors potentially associated with ICL. Seven of 304 (2.3%) seronegative hemophilic men and one of 160 (0.6%) female partners met the ICL definition, but the condition resolved for two of the men and for the sole female partner. All five men with persistent ICL had lymphocytopenia (< 1,200 total lymphocytes/microliters) and < 300 total CD4+ lymphocytes/microliters; only one had a low CD4+ percentage. On the most recent measurement, 14.5% of the 304 seronegative hemophilic men had lymphocytopenia. Compared with matched hemophilic controls, men with persistent ICL more often had a history of liver disease (3/5 cases, 0/21 controls, P = 0.007) or splenomegaly (3/5 cases, 4/21 controls; P = 0.04), but not severe hemophilia, greater clotting factor concentrate exposure, high alanine aminotransferase levels, hepatitis B virus antigenemia, or detectable hepatitis C virus RNA in plasma. All five cases and 20/21 controls had antibodies to hepatitis C virus present in their serum. In this cohort of hemophilic men, ICL was related to lymphocytopenia associated with liver disease rather than selective loss of CD4+ lymphocytes.

To evaluate the safety and efficacy of didanosine (ddl) monotherapy and three different combinations of zidovudine (ZDV) and ddl in asymptomatic human immunodeficiency virus-1 (HIV-1) infection, we conducted an open-label, phase I/II study in 126 asymptomatic HIV-1-infected hemophilic and nonhemophilic subjects with a CD4 count of 200 to 500/mm3 stratified for prior zidovudine treatment and baseline CD4 count. Study arms included arm A, low-dose combination (ZDV 150 mg and ddl 134 mg, daily); arm B, moderate-dose combination (ZDV 300 mg and ddI 334 mg, daily); arm C, high-dose combination (ZDV 600 mg and ddl 500 mg, daily), and arm D, ddl monotherapy (ddl 500 mg, daily). Earlier, more frequent hepatotoxicity was experienced by hemophilic subjects (P = .008), but there were no differences in toxicity between treatment arms (P = .51), nor were there any differences in the rate of development of clinical endpoints by treatment (P = .41). Smaller median CD4 increases occurred over the first 12 weeks for arms A and D, 44/mm3 and 42/mm3, than arms B and C, 105/mm3 and 114/mm3, respectively, (P = .015). Hemophilia status (P = .0004) and prior ZDV experience (P = .044) independently predicted weaker CD4 responses during the first 12 weeks of treatment. Using a regression model and adjusting for hemophilia status, prior ZDV treatment, and baseline CD4, there was a significant reduction in quantitative viral load from baseline by week 12 for all treatment arms combined (P = .0001), with significantly lower median percent reduction for arm A (56.3%) than arms B, C, and D (94.6%, 98.5%, and 91.9%, respectively, P = .015). Although greater hepatoxicity and weaker CD4 responses occur in hemophilic subjects, didanosine monotherapy and combination therapy with zidovudine are safe and effective in asymptomatic HIV-1-infected patients.

Background: Hemophilia A (HA) is an X-linked recessive disorder that affects males, whereas female carriers are presumed asymptomatic if Factor VIII activity levels (FVIII:C) fall within normal range. However, FVIII:C does not always correlate with bleeding phenotype, leading to an underappreciation of bleeding sequelae in females. Therefore, it is clinically important to identify HA carriers at higher bleeding risk. FVIII expression in HA carriers is influenced by X chromosome inactivation (XCI), a process that silences one X in XX females such that each cell has a random probability of inactivating either parental X. However, rare female carriers of X-linked disorders can be severely affected if XCI is skewed and the mutant X is preferentially active. How XCI skewing modulates bleeding in mild/moderate HA is less well understood. HA bleeding may be also affected by variants in factors influencing FVIII binding and clearance, including Von Willebrand Factor (VWF) and ABO blood type. To better understand HA carrier bleeding tendency, we analyzed a family that segregates a mild/moderate HA mutation (F8: c.2167G>A). Four carriers in this pedigree have FVIII:C that approach affected males, necessitating prophylaxis prior to surgical procedures. We hypothesized that bleeding in these carriers can be largely explained by XCI skewing, but additional variants may fine tune FVIII:C. Methodology: FVIII levels were assessed by one stage (FVIII:C) and chromogenic (FVIII:CR) assays. At least two plasma samples spanning >3 years from each female were tested in duplicate with each FVIII assay. To address XCI skewing, we performed methylation-based assays at the Androgen Receptor (AR) and Fragile X Mental Retardation 1 (FMR1) loci. At least three independent PBMC DNA samples from each female were evaluated. Additionally, we screened VWF regions known to influence FVIII:C (exons 18-20, 24-27). Results: For each female, results between XCI assays were indistinguishable (r2 = 0.99). Two of four females had pronounced skewing (≥80:20); a third had measurable skewing (67:33). Importantly, all three predominantly expressed the mutant paternal allele. Familial XCI skewing argues for a genetic cause. However, XIST, the major regulator of XCI, lacked promoter alterations. Importantly, there was linear correlation between XCI skewing and FVIII:C measured by FVIII:C or FVIII:CR assays (r2 = 0.77 and 0.83, respectively). One female with random XCI, had FVIII:C considered hemostatic (median 51%, range 43-67), whereas the other females with skewed XCI had levels Conclusions: Our results indicate that HA carrier bleeding phenotypes are multifaceted, and the major determinant of FVIII:C is XCI. These results also suggest that even moderate XCI skewing could influence clinical bleeding in HA carriers. However, random XCI in one female explains FVIII:C but does not fully negate bleeding tendency, emphasizing the complexity of carrier phenotype. These findings provide justification for an expanded study of carriers in unrelated pedigrees using a comprehensive approach to include XCI assays. Disclosures No relevant conflicts of interest to declare.

Background: Hemophilia A (HA) is an X-linked disorder that primarily affects males, although female carriers can exhibit bleeding phenotypes. Factor VIII activity levels (FVIII:C) in XX females are influenced by X chromosome inactivation (XCI), a process that silences each parental X in a proportion of cells. XCI skewing can decrease FVIII expression by preferentially inactivating the normal X chromosome. FVIII:C is further modulated by factors such as ABO blood groups and von Willebrand factor (VWF). The D'/D3 domain of VWF binds circulating FVIII protein, preventing proteolytic degradation. In type 2 Normandy von Willebrand disease (2N VWD), D'/D3 mutations decrease affinity to FVIII and result in bleeding events similar to mild/moderate HA. Variants in VWF also affect the pharmacokinetics of recombinant FVIII. Current clinical screening tests detect the 3 VWF mutations responsible for >90% of 2N, but report variants not directly responsible for 2N as clinically benign. However, common polymorphisms are known to affect FVIII:C in normal individuals. Therefore, a better understanding of how specific alterations in VWF modulate HA phenotype is necessary to interpret clinical presentation and refine management with factor concentrates. This is particularly important in HA individuals carrying mild/moderate mutations. To evaluate VWF variants in HA carriers, we focused on an extended pedigree that includes four obligate carriers from a family with mild/moderate HA (FVIII: p.Ala723Thr). FVIII:C varied and largely correlated with XCI skewing. Nevertheless, the FVIII:VWF interaction prompted us to identify VWF gene variants that could further modulate FVIII:C and contribute to bleeding in this family. Methods: To identify D'/D3 VWF variants that impact FVIII binding, primers were designed to amplify exons 17-20 and 24-27 on chromosome 12. A chromosome 22 pseudogene, with 97% identity to VWF exons 24-34, complicated primer design. Primers specific to VWF were selected by targeting regions that differed from the pseudogene and were verified by digestion with a restriction enzyme unique to each chromosome 12 exon. The PCR products were amplified and sequenced from the four female carriers and a control male relative. Results: After excluding the three most common mutations responsible for 2N, seven other variants were identified. Four of these were intronic polymorphisms and a synonymous variant at p.Asn1138 not associated with VWD and presumed to be clinically benign. All but one of these have been described in normal individuals. Two females were heterozygous for missense variant rs1063856 (p.Thr789Ala) and synonymous polymorphism rs1063857 (p.Tyr795=) that are in linkage disequilibrium and are likely to impact FVIII:C and VWF antigen (VWF:Ag) levels. These common variants, found in up to 36% of Caucasians and 58% of African Americans, are reported to increase VWF:Ag and FVIII:C jn heterozygotes (9 IU/dL and 7 IU/dL respectively). Neither ABO blood groups nor XCI skewing could fully explain the differences in FVIII:C activity observed with this variant. Conclusions: We propose that VWF variants rs1063856/rs1063857 may contribute to FVIII:C differences between two females in the pedigree with similar XCI skewing. We conclude that consideration of VWF variants is important for fully understanding bleeding phenotype and treatment responses in female carriers and males in families with mild/moderate HA. These findings support the need for expanded studies into the role of FVIII and VWF variant interactions in additional unrelated HA individuals. Disclosures No relevant conflicts of interest to declare.

Patients with inherited bleeding disorders who received clotting factor concentrates before 1987 have high rates of hepatitis C virus (HCV) or HCV/HIV infection. We evaluated HCV quasispecies evolution in longitudinally collected specimens comparing those from patients with progression to end-stage liver disease (ESLD; cases) to those with compensated chronic hepatitis (controls). Plasma samples were obtained from the National Cancer Institute Multicenter Hemophilia Cohort Study. Controls were matched for age, sex, infection duration, and presence/absence of HIV. Samples from early infection were compared to those obtained after onset of ESLD in the cases. The first hypervariable (HVR1) and core proteincoding regions were amplified, subcloned, and sequenced. Complexity and diversity were determined. More than 700 sub-clones from 10 pairs of patients (8 with HIV) followed over approximately 9.3 years were evaluated. HVR1 complexity narrowed over time in the cases, whereas it increased in controls (P = .01). Similar trends were observed for diversity within HVR1 and the core region (P = .04). HCV-infected patients with inherited bleeding disorders undergo quasispecies evolution over time. Evolution patterns differ for progressors and nonprogressors. Further understanding of these mechanisms may help identify factors related to progression rate and treatment response.

[Graphic][1] Introduction: Factor VIII (FVIII) products with a longer half-life may allow for longer intervals between treatments for patients with hemophilia A and may facilitate prophylaxis tailored to an individual’s bleeding phenotype. BAY 94-9027, a PEGylated FVIII product, demonstrated an extended half-life in a phase 1 trial and was well tolerated and efficacious in a phase 2/3 study with dosing intervals up to every 7 days. In this subanalysis of the phase 2/3 trial, bleeding frequency calculated based on the BAY 94-9027 prophylactic dosing regimen during the study was compared with reported bleeding frequency in the 12 months before enrollment. Also, on-study annualized bleeding rates (ABRs) for joint, spontaneous, and trauma bleeds are presented by treatment group. Methods: PROTECT VIII was a multinational, partially randomized, open-label, 36-week study in previously treated patients aged 12–65 years with severe hemophilia A and no history of FVIII inhibitors. Patients received BAY 94-9027 for 36 weeks either on demand or prophylactically. Patients were assigned to 1 of 3 prophylaxis dosing regimens based on the number of bleeds observed during a 10-week run-in period, during which all patients in the prophylaxis arm were treated with 25 IU/kg BAY 94-9027 2x/week. Patients with ≤1 breakthrough bleed during the 10-week period were randomized 1:1 to BAY 94-9027 45–60 IU/kg every 5 days or 60 IU/kg every 7 days. Patients with ≥2 breakthrough bleeds received 30–40 IU/kg BAY 94-9027 2x/week. ABR and annualized joint bleeding rate (AJBR) for the 12 months before the study (collected retrospectively at screening) were compared with values calculated in patients previously treated with prophylaxis who used BAY 94-9027 prophylaxis during the study (weeks 0–36 for the combined prophylaxis groups [including the 10-week period]; weeks 10–36 for the 3 assigned prophylaxis dosing regimens). ABRs for joint, spontaneous, and trauma bleeds during the study were analyzed for the on-demand and combined prophylaxis groups (weeks 0–36) and in relation to patients’ BAY 94-9027 dosing regimen (weeks 10–36). Results : The intent-to-treat population comprised132 patients (prophylaxis, n=112; on demand, n=20). In patients previously treated with prophylaxis, median ABR and AJBR during BAY 94-9027 prophylaxis (weeks 0–36) were lower than corresponding prestudy values; ABR and AJBR during weeks 10–36 for every-5-day, every-7-day, and 2x/week BAY 94-9027 dosing were also lower than or comparable to prestudy values ( Table ). Median ABRs for joint, spontaneous, and trauma bleeds were lower for the combined prophylaxis groups (weeks 0–36) compared with the on-demand group (combined prophylaxis groups: 1.5, 1.4, and 0.0, respectively; on-demand group: 16.3, 14.3, and 9.1). In the prophylaxis arms (weeks 10–36), median ABRs for joint, spontaneous, and trauma bleeds were 2.1, 0.0, and 0.0 for 2x/week dosing; 1.9, 0.0, and 0.0 for every-5-day dosing; and 1.9, 1.9, and 0.0 for every-7-day dosing. | | Combined Prophylaxis, | 2x/week, week 10–36 | Every 5 Days, | Every 7 Days, | | ------------ | ---------------------------- | ------------------------ | ------------------------------ | ---------------------------- | ---------------------------- | | | week 0 – 36 (n=87)* | Required† (n=9) | Not Randomized‡ (n=6) | week 10 – 36 (n=34) | week 10 – 36 (n=37) | | | | | | ABR, median | | | | | | | Prestudy | 5 | 12 | 5.5 | 3 | 2 | | Study | 2.82 | 8.7 | 0.75 | 1.48 | 2.88 | | P value | 0.0015 | 0.2445 | 0.0766 | 0.0039 | 0.4981 | | AJBR, median | | | | | | | Prestudy | 2 | 9 | 3.5 | 2 | 2 | | Study | 1.46 | 7.24 | | 1.40 | 1.39 | | P value | 0.0045 | 0.3484 | 0.0673 | 0.0131 | 0.4111 | * P values (paired Student’s t test) are nominal, as no multiplicity control was applied. * *n=86 for AJBR. * †Patients with ≥2 breakthrough bleeds in weeks 0–10. * ‡Patients with ≤1 bleeds in weeks 0–10 who were not randomized (randomized arms were filled). Abstract 1526. Table. Bleeding Frequency During BAY 94-9027 Prophylaxis vs Prestudy Values Conclusions: BAY 94-9027 prophylaxis resulted in lower ABRs and AJBRs during the 36-week study period compared with prestudy values in patients previously treated with prophylaxis. Subgroup analyses based on prophylactic dosing regimens (including dosing intervals of up to every 7 days) showed that patients who were randomized based on bleeding phenotype during the 10-week run-in period achieved bleeding control that was better than or comparable to their prestudy levels, highlighting the value of individualized phenotype-based dosing with BAY 94-9027. In addition, prophylaxis with BAY 94-9027 resulted in reduced joint, spontaneous, and trauma bleeds compared with on-demand treatment. Disclosures Boggio: Bayer, Baxter, Novo Nordisk, CSL Behring, Grifols, and Pfizer: Consultancy. Hong: Bayer HealthCare: Employment. Wang: Bayer HealthCare Pharmaceuticals: Employment. Michaels: Bayer HealthCare Pharmaceuticals: Employment. [1]: /embed/inline-graphic-2.gif

Introduction: Cardiovascular disease (CVD) is a disease of aging. While men with hemophilia were initially thought to be protected from CVD, it is now clear that atherothrombotic events do occur. The objective of this study is to determine the prevalence of CVD and CV risk factors among older men with moderate and severe hemophilia. Methods: A U.S. national cross-sectional study began enrollment in 10/2012. Inclusion criteria are men with moderate or severe congenital hemophilia A or B (FVIII or IX level ≤ 5%), age 54-73. Men with an additional bleeding disorder (besides liver dysfunction) were excluded. After obtaining informed consent, CV risk factors, medications, and history of thrombotic events were obtained from patient interview and chart review. A fasting blood sample was assayed centrally. Results: As of 8/1/2014, 160/200 planned subjects were recruited and interim analysis on 126 subjects from 18 U.S. Hemophilia Treatment Centers is presented here. The majority were white (109; 86.9%) or African American (15; 11.5%). Mean age was 62 years (SD: 5; range: 54-74). Most used factor on demand, with only 34.1% (43/126) on prophylaxis, defined as ≥2 doses of FVIII or ≥ 1 dose FIX/week. Five (4.0%) had a current inhibitor. Viral infection was common; 68.3% currently had hepatitis C, and 25% HIV. Hypertension (HTN) was reported in 65.6% of subjects, 37.3% dyslipidemia and 24.6% diabetes (DM); 44.5% had ever smoked, 56.3% denied engaging in at least moderate physical activity and 43.7% had a family history of CVD. Average BMI was 28 kg/m2 (29.4% obese) and waist circumference 97 cm. Fasting blood work showed an abnormally elevated: creatinine in 27.6% subjects (mean 1.09 mg/dl, SD 0.5), CRP in 6.3% (4.15 mg/L, 10.6), total cholesterol in 18.1% (169.49 mg/dl, 35.8), triglycerides in 25.2% (122.10 mg/dl, 58.2), LDL in 19.7% (102.27 mg/dl, 32.3); and low HDL in 45.7% (42.80 mg/dl, 12.2). Ten subjects (7.9%) reported prior angina; 7 (5.6%) atrial fibrillation/flutter; 3 (2.4%) leg deep venous thrombosis; 2 (1.6%) myocardial infarction (MI), transient ischemic event (TIA), or pulmonary emboli; and 1 (0.8%) coronary artery angioplasty, stent placement, CABG, or peripheral arterial angioplasty. In total, 11 subjects had CVD (defined as angina, MI, TIA, or ischemic or embolic stroke), a prevalence rate of 8.7%. This is significantly lower than the reported prevalence of 23% CVD in similar aged men without hemophilia in the longitudinal ARIC cohort (p-value Due to the small number of events, individual CV risk factors thus far did not achieve statistical significance in predicting CVD. Compared to never smokers, ever smokers had an odds ratio (OR) of 3.7 (95% CI: 0.9-14.8) of CVD. For HTN, dyslipidemia, and DM, the OR (95% CI) of CVD were 2.5 (0.5-12.1), 2.2 (0.6-7.5), and 1.9 (0.5-6.8), respectively. Positive family history (OR 2.4 (0.7-8.8)) and low-level of physical activity (1.4 (0.4-5.0)) also suggested some association with increased CVD risk. Obese BMI and large waist circumference were not significant. Men using prophylaxis appeared less likely to have CVD (1/43, 2.3%) than men not on prophylaxis (10/83, 12.1%), OR 0.2 (0.02-1.4), although the difference was not statistically significant. HIV+ men (1/32, 3.1%) were also less likely to have CVD compared to non-HIV+ men (10/92, 10.9%), OR 0.3 (0.2-10.9), but not significantly so. Lastly we investigated the role of anti-HTN (used in 36.5% of all subjects), cholesterol lowering (16.7%), and DM medications (10.3%) in reducing CVD. Not taking anti-HTN, cholesterol or DM medications non-significantly increased CVD risk with an OR in all subjects of 1.5, 3.3, or 3.9 respectively. Conclusions: In this interim analysis of an ongoing national cross-sectional study, older men with moderate to severe hemophilia commonly report risk factors for CVD, including HTN (65.6%), dyslipidemia (37.3%) and renal insufficiency (27.6%). Despite this, the prevalence of reported CVD is low at 8.7%, suggesting that men with hemophilia may be protected from forming pathogenic thrombi. More data is needed to determine if the approach to prophylaxis or other therapies should be altered in this population. We plan to formally compare the prevalence of CVD and CV risk factors with similarly aged men in the ARIC database once enrollment is complete. Disclosures Sood: Bayer: Research Funding. Shapiro:Baxter: Consultancy, Global Steering Committee Other, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Novo Nordisk: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Bayer: Consultancy, Global Steering Committee, Global Steering Committee Other, Research Funding; CSL Behring: Research Funding; Biogen Idec: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Kedrion: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding. Kempton:Baxter Healthcare, Inc: Membership on an entity's Board of Directors or advisory committees; CSL Behring: Membership on an entity's Board of Directors or advisory committees; Kedrion Biopharma: Membership on an entity's Board of Directors or advisory committees; NovoNordisk, Inc: Research Funding. Fogarty:Amgen Inc: Consultancy; Bayer HealthCare Pharmaceuticals: Consultancy; Baxter: Consultancy; Biogen Idec Inc.: Consultancy; Chugai Pharma USA: Consultancy; Pfizer Inc: Consultancy; Baxter: Research Funding; Biogen Idec Inc.: Research Funding; CSL Behring: Research Funding; Pfizer Inc: Research Funding; Medscape LLC: Honoraria; VindicoMed: Honoraria. Ragni:Biogen Idec: Consultancy, Research Funding; Bristol-Myers Squibb: Consultancy, Research Funding; GlaxoSmithKline: Consultancy, Research Funding; Tacere Benitec: Consultancy, Drug Safety Monitoring Board, Drug Safety Monitoring Board Other; Baxter: Research Funding; Bayer: Research Funding; CSL Behring: Research Funding; Merck: Research Funding; Novartis: Research Funding; Novo Nordisk: Research Funding; Pfizer: Research Funding; Spark Therapeutics: Research Funding; Vascular Medicine Institute, PIttsburgh, PA: Research Funding. Neff:Baxter: Membership on an entity's Board of Directors or advisory committees. Konkle:CDC: Research Funding, This work was supported by CDC grant 1U01DD000761-01 Other.

Background The ability of a person with hemophilia to adhere to treatment recommendations aimed at preventing and controlling bleeding likely impacts clinical outcomes and resource utilization. Annual cost of replacement factor for persons with severe hemophilia on prophylaxis is estimated to be as high as $300,000 per year (Johnson KA and Zhou ZY. ASH Annual Meeting Educational Program. 2011; 413-418). Hemophilia care is optimized when delivered by a highly specialized hemophilia treatment center (HTC) (Soucie et al. Blood. 2000; 96: 437-442). Consolidation of care for persons with hemophilia from a large geographic region within an HTC results in demographic heterogeneity that may impact an individual's ability to adhere to treatment recommendations. Little is known regarding the impact of socio-demographic factors on treatment outcomes for persons with hemophilia. The purpose of this study was to identify disease-related, treatment-related and demographic variables that have a significant impact on morbidity and resource utilization. Methods We identified 69 persons with severe hemophilia treated at our HTC between June 2009 and June 2012. We collected data for the variables listed below in table 1. To assess morbidity and resource utilization, we collected data for total factor use, total outpatient cost, total inpatient cost, frequency of inpatient and outpatient encounters and length of stay when hospitalized. Risk factors were identified using linear regression. A subset analysis was performed to evaluate the impact of prophylaxis on patients without inhibitors. All statistical analyses were performed using STATA (version 12.0 College Station, TX). Statistical significance was defined as P < 0.05. Results Age and inhibitor status were the only variables to significantly impact cost in both inpatient and outpatient settings (Table 1). The statistically significant beneficial impact of prophylaxis was confirmed by subgroup analysis of patients without inhibitors with respect to number of hospitalizations and length of stay, but not total cost. Conclusions Identifying and addressing potential barriers to care will likely reduce morbidity and cost for persons with hemophilia. Caregiver status and age are demographic variables that may contribute to poor medical adherence resulting in increased morbidity and cost of care for persons with severe hemophilia. Disclosures: No relevant conflicts of interest to declare.

Background Patients undergoing major orthopedic surgery such as hip and knee arthroplasty are at particularly high risk for venous thromboembolism (VTE) due to the nature of the surgery and postoperative immobilization, and thus routine thromboprophylaxis has been the standard of care for more than 15 years. Peri-operative clotting factor concentrate replacement is administered to reverse the inherent hemostatic defect in persons with hemophilia (PWH), potentially rendering these patients at risk for developing deep vein thrombosis (DVT) postoperatively. Currently, no published guidelines address the risk of DVT or the indications for thromboprophylaxis in PWH undergoing hip or knee arthroplasty. Consequently, given their underlying bleeding risk, thromboprophylaxis practice in individuals with hemophilia undergoing orthopedic procedures varies considerably among Hemophilia Treatment Centers (HTCs). Objective To determine the rate of ultrasound-detectable lower extremity DVT (asymptomatic or symptomatic) or clinically apparent pulmonary embolism (PE) in PWH undergoing total hip or knee arthroplasty. Methods This was a prospective, multicenter observational study. All patients with hemophilia A or B scheduled for knee or hip replacement at participating centers were eligible to participate. Patients with a prior history of VTE or those receiving anticoagulation prior to surgery were excluded. Baseline venous duplex ultrasounds of the lower extremities (proximal and distal) were obtained within 7 days prior to surgery, and follow-up ultrasounds were performed 4-6 weeks after surgery. Patients were otherwise evaluated and treated per the local institutional standard of care. Coagulation factor dosing and plasma factor activity levels were recorded, as was use of mechanical (intermittent pneumatic compression) or pharmacologic thromboprophylaxis. Major bleeding was defined as bleeding that resulted in a decrease of hemoglobin of 2 g/dL or more over a 24 hour period, bleeding that led to transfusion of 2 or more units of packed red blood cells, bleeding into a critical site, or bleeding that led to death. Minor bleeding was defined as clinically overt bleeding that did not satisfy any of the major bleeding criteria. Results 11 HTCs enrolled a total of 46 patients between February 2010 and June 2013. 44 patients completed the study (2 withdrew due to surgery cancellation); 13 had hip replacement (median age 50.9 years) and 31 had knee replacement (median age 49.1 years). 15 patients (34.1%) had a history of inhibitor prior to surgery. 6 patients (13.6%) were treated with bypassing agents perioperatively; 3 of these patients had active inhibitors present at the time of surgery. The remaining 38 patients received factor VIII or IX replacement. 22 patients (50%) received some form of mechanical prophylaxis during hospitalization, and 3 patients (6.8%) also received low molecular weight heparin (LMWH) prophylaxis. 22 patients (50%) did not receive any form of prophylaxis. 1 patient (2.3%, 95% CI 0-12%) with moderate hemophilia A and no history of inhibitor who received mechanical prophylaxis was diagnosed with symptomatic distal DVT on postoperative day 6 (confirmed by ultrasound). No patients (0%, 95% CI 0-8.0%) had asymptomatic DVT or clinically apparent PE. 16 patients (36.4%) had major bleeding during hospitalization. 4 of the 6 patients treated with a bypassing agent had major bleeding, and 1 of these patients required a re-operation due to bleeding in the replaced joint. 2 of the 3 patients who received LMWH prophylaxis had major bleeding events. 4 patients (9.1%) experienced minor bleeding. Conclusions In this prospective study, the observed prevalence of ultrasound detectable DVT in PWH following total knee or hip arthroplasty was very low. In the general population, when thromboprophylaxis is not administered, DVT prevalence (measured by venography) after knee and hip replacement is 40-84% and 40-57%, respectively. With mechanical prophylaxis, these rates decrease to 20.3% for hip replacement and 28.3% for knee replacement. Although these prevalence rates are expected to be somewhat lower when ultrasound is used for diagnosis, the low prevalence of DVT (2.3%) and the high rate of bleeding complications (36.4% major; 9.1% minor) in this study suggest that pharmacologic prophylaxis should not be used following knee or hip replacement in PWH with no prior history of VTE. Disclosures: Kempton: Baxter Healthcare: Membership on an entity’s Board of Directors or advisory committees; Novo Nordisk: Research Funding.

Objective: To assess the efficacy of Alphanate® as replacement therapy in subjects with congenital VWD undergoing surgical or invasive procedures. Methodology: Retrospective data from 5 hospitals and accounting for 8 years of chart review has been collected. The study protocol and the subject’s informed consent were approved by the local Institutional Review Boards. To date, 35 treated events (27 subjects) have been evaluated. Events were classified by local investigators and an independent referee committee as major or minor surgery, and invasive procedures. Treatment efficacy was rated using a 4-point verbal rating scale. Replacement therapy was considered effective if the treatment outcome was rated as excellent or good, and non-effective when the outcome was rated as poor or none. Results: Efficacy results (number and percentage) obtained by local investigators and the referee committee are presented in Tables 1 and 2. Conclusions: Since there may be bias in the interpretation of responses made by the participating physicians, it was felt that true responses might better be assessed retrospectively by an adjudication committee. Therefore, both Tables were presented here. Much of the discrepancies between these 2 tables was generated by oral surgery. There was a high level of efficacy agreement between investigators and the referree committee. Therefore, it is concluded that Alphanate® is effective in preventing excessive bleedings during surgeries and invasive procedures in subjects with congenital VWD. | | Major surgery (n=9) | Minor surgery (n=17) | Invasive procedure (n=9) | Total (n=35) | |:--------- | ------------------- | -------------------- | ------------------------ | ------------ | | Excellent | 6 (66.7%) | 14 (82.4%) | 8 (88.9%) | 28 (80.0%) | | Good | 3 (33.3%) | 1 (5.9%) | 1 (11.1%) | 5 (14.3%) | | Poor | 0 (0.0%) | 1 (5.9%) | 0 (0.0%) | 1 (2.9%) | | None | 0 (0.0%) | 1 (5.9%) | 0 (0.0%) | 1 (2.9%) | Table 1. Efficacy Results Obtained by Investigators | | Major Surgery (n=12) | Minor Surgery (n=11) | Invasive Procedure (n=12) | Total (n=35) | |:--------- | -------------------- | -------------------- | ------------------------- | ------------ | | Excellent | 8 (66.7%) | 11 (100%) | 11 (91.7%) | 30 (85.7%) | | Good | 1 (8.3%) | 0 (0.0%) | 0 (0.0%) | 1 (2.9%) | | Poor | 3 (25.0%) | 0 (0.0%) | 1 (8.3%) | 4 (11.4%) | | None | 0 (0.0%) | 0 (0.0%) | 0 (0.0%) | 0 (0.0%) | Table 2. Efficacy Results Obtained by Referee Committee

The fibrinolytic system was assessed in 28 hemophiliacs using the chromogenic substrate H-D-Val-Leu-Lys-pNA. To determine whether a state of hyperfibrinolysis might be associated with Factor VIII replacement therapy, 14 patients with severe disease who were intensively treated with Factor VIII concentrates were compared with 14 patients with mild disease who were receiving infrequent transfusions with cryoprecipitate or fresh frozen plasma. Seventeen normal males served as controls. With the exception of an elevated level of plasminogen activator and a decreased level of immediate antiplasmin in the mild group only, no evidence of enhanced fibrinolysis was found. Other components of the fibrinolytic system were either normal (plasmin) or increased (progressive antiplasmin containing both alpha 2PI and alpha 2M, and plasminogen). The elevated plasminogen levels were found only in the severe intensively transfused group. The elevated progressive antiplasmin levels were found in both groups of patients and did not appear to be related to transfusions. These findings do not support the concept of enhanced fibrinolysis associated with intensive Factor VIII replacement therapy.

Summary. We recently observed an increase in factor-VIII clot promoting activity as measured by a one-stage assay (VIII AHF1) in a haemophiliac with hepatitis. However, VIII AHF as measured by a two-stage assay (VIII AHF2) was 0.013 u/ml at a time when VIII AHF1 measured 0.38 u/ml. We then studied seven non-haemophiliacs with liver disease, and attempted to correlate the levels of VIII AHF1 and VIII AHF2 with factor VIII-like antigen (VIII AGN) as measured by quantitative immunoelectrophoresis. In four of the seven patients, disproportionate elevations of VIII AHF2 compared to VIII AHF1 were found. Furthermore, VIII AHF2 values correlated well with VIII AGN values. No such discrepancy was apparent in four normal control subjects. These findings emphasize the necessity for performing two-stage assays in haemophiliacs as well as non-haemophiliacs with liver disease to assess factor-VIII levels. In addition, they suggest that confirmation of the diagnosis of haemophilia may not be possible in the haemophiliac with hepatitis unless VIII AHF2 determinations are performed. The reason for the disparity between VIII AHF1 and VIII AHF2 levels is not apparent. However, the correlation of VIII AGN and VIII AHF2 levels in the non-haemophiliacs with liver disease provides further support for the concept that VIII AGN and VIII AHF are closely related or identical molecular entities.

In Pennsylvania, the prevalence of hemophilia is one per 10,000 males. Factor VIII deficiency is five times more frequent than Factor IX deficiency, and 34% of the patients have no relatives affected with the disease. The mean age is 23 years old, and 50% of the patients are less than 20 years old. Approximately one-third of the patients with Factor VIII deficiency and one fourth of the patients with Factor IX deficiency have levels of less than 0.01 mu/ml. By clinical criteria, 55% of those with Factor VIII deficiency are severe compared to 45% of those with Factor IX deficiency. Factor VIII-deficient patients are treated an average of 18 times per year compared to ten times per year for patients with Factor IX deficiency. Hemarthroses account for 70% of the hemorrhages treated and for 40% of the concentrate usage. Home therapy patients use an average of 45,950 Factor VIII units per year at a cost of ø170 per patient and their use accounts for 60% of the total Factor VIII usage of 1.7 million units. Less than five days per patient per year are lost from school or work because of bleeding, and patients are hospitalized for bleeding an average of only two to three days per patient year. Adverse immediate reactions to therapy are infrequent. Five percent of patients have persistence of the hepatitis B virus, and 7.5% have inhibitors. The mortality rate is 0.04% per year, with half of the deaths being hemophilia-related.

Proportionate twofold or greater increases in both Factor VIII procoagulant activity (VIII AHF) and Factor VIII-like antigen (VIII AGN) have been observed to follow the administration of adrenalin in normal individuals. Levels of factor VIII procoagulant or clot-promoting activity (VIII AHF) and VIII AGN were measured immediately before and one hour after the subcutaneous injection of adrenalin (0.35 ml) in ten adults with hemophilia A documented by low VIII AHF levels (ranging from 0.01-0.25 units/ml, normal VIII AGN levels, and normal VIII VWF levels as measured by the washed-platelet ristocetin assay). Eighteen normal adults served as controls. Results were analyzed by the paired t test. It was shown that VIII AGN increased significantly, while VIII AHF did not change, in hemophiliacs after adrenalin stimulation, regardless of the base line level. This fixed rate of VIII AHF production, release, or activation may help to explain the constancy of VIII AHF levels in individual hemophiliacs and their affected family members.

Up to the late 1980s, hepatitis C virus (HCV) infected the majority of people with hemophilia (PWH) in the United States (US), via contaminated plasma clotting factor therapy. Approximately two-thirds of the HCV-infected PWH in the US also were infected with human immunodeficiency virus (HIV). The Second Multicenter Hemophilia Cohort Study (MHCS-II) investigated complications of HCV and HIV infections in people with hemophilia. From 2001 - 2005, data and specimens were collected from 2561 anti-HCV positive participants, including 763 who were co-infected with HIV. These data were analyzed for correlates of spontaneous HCV clearance. Results. Of the 763 HIV-positive, anti-HCV-positive participants, 285 were eliminated from the analysis because they had received interferon treatment to clear HCV, lacked HCV treatment data, or lacked sufficient specimens to determine HCV clearance status. Of the remaining 478 interferon-untreated participants, 61 (12.8%) had cleared HCV RNA from plasma. HCV had cleared in 8.6% of the 116 who were never infected with hepatitis B virus (HBV), 11.6% of 207 with resolved HBV infection, and 8.9% of 124 with missing HBV status (P≥ 0.41). In contrast, HCV had cleared in 16 (51.6%) of the 31 participants with chronic HBV surface antigen (HBsAg) in serum. Adjusted for gender and race, HCV clearance was increased 7.4-fold (95% CI 3.7 - 15.0) for those with chronic HBsAg. In the HBsAg negative group, HCV clearance was significantly lower among participants of African ancestry (1.6%, p=0.01) and was significantly lower in PWH with severe compared to mild or moderate coagulopathy (8.2% vs 15.9%, p=0.02). Clearance was unrelated to type of initial clotting factor replacement therapy, year of birth, year of primary HCV infection, age at primary HCV infection, age at HIV infection, and year of enrollment in the cohort. HCV clearance occurred in 2 (12.5%) of the 14 participants in whom primary HCV infection occurred after or within one year of HIV infection, compared to 9.1% when HCV infection occurred more than one year before HIV (p=0.65). HCV clearance was unrelated to CD4 count at cohort enrollment, history of immunologically or clinically defined AIDS, and detection of HIV RNA in plasma at cohort enrollment. HCV clearance tended to be higher among the 231 participants on HAART (12.6%) than in those on less intensive (9.1%), anomalous (6.7%) or no anti-HIV therapy (6.5%, Ptrend=0.07), although when restricted to non-African ancestry males, HIV therapy was not associated with clearance (Ptrend=0.26). Conclusion. Unlike in HIV-negative people with hemophilia, HCV clearance in the current study was unrelated to age at or year of primary HCV infection. We were unable to evaluate repeated exposure to and likely re-infection with HCV, although the lower clearance in patients with severe hemophilia, who would have been treated, and thus exposed more frequently, supports the hypothesis of lower clearance with re-infection. While reduced overall, spontaneous HCV clearance was markedly increased by the presence of chronic HBsAg carriage. These findings will increase our understanding of interactions between these viruses and may lead to improved approaches to treatment of co-infected patients.