Clinical characteristics.

Hemophilia B is characterized by deficiency in factor IX clotting activity that results in prolonged oozing after injuries, tooth extractions, or surgery, and delayed or recurrent bleeding prior to complete wound healing. The age of diagnosis and frequency of bleeding episodes are related to the level of factor IX clotting activity. In any individual with hemophilia B, bleeding episodes may be more frequent in childhood and adolescence than in adulthood.

• Individuals with severe hemophilia B are usually diagnosed during the first two years of life. Without prophylactic treatment, they may average up to two to five spontaneous bleeding episodes each month, including spontaneous joint or muscle bleeds, and prolonged bleeding or excessive pain and swelling from minor injuries, surgery, and tooth extractions.
• Individuals with moderate hemophilia B seldom have spontaneous bleeding, although it varies between individuals; however, they do have prolonged or delayed oozing after relatively minor trauma and are usually diagnosed before age five to six years. The frequency of bleeding episodes varies from once a month to once a year.
• Individuals with mild hemophilia B do not have spontaneous bleeding episodes; however, without pre- and postoperative treatment, abnormal bleeding occurs with surgery or tooth extractions. The frequency of bleeding may vary from once a year to once every ten years. Individuals with mild hemophilia B are often not diagnosed until later in life.

Approximately 30% of heterozygous females have factor IX clotting activity lower than 40% and are at risk for bleeding (even if the affected family member has mild hemophilia B), although symptoms are usually mild. After major trauma or invasive procedures, prolonged or excessive bleeding usually occurs, regardless of severity.

Diagnosis/testing.

The diagnosis of hemophilia B is established in individuals with low factor IX clotting activity. Identification of a hemizygous F9 pathogenic variant on molecular genetic testing in a male proband confirms the diagnosis. Identification of a heterozygous F9 pathogenic variant on molecular genetic testing in a symptomatic female confirms the diagnosis.

Management.

Treatment of manifestations: Referral to a hemophilia treatment center (HTC) for assessment, education, genetic counseling, and treatment.

Targeted therapy: For those with severe disease, prophylactic infusions of factor IX concentrate to maintain factor IX clotting activity higher than 1% or as needed to prevent bleeding and allow normal activity improves outcomes and prevents chronic joint disease. Some individuals with moderate hemophilia bleed frequently enough to benefit from prophylaxis. Longer-acting products that allow weekly or biweekly dosing are now available. Intravenous infusion of plasma-derived or recombinant factor IX for acute bleeding episodes should be given as soon as possible after symptoms occur. Training in home infusion should be provided for individuals/families affected by moderate and severe hemophilia.

Supportive care: Physical therapy for evaluation and treatment of musculoskeletal disease; standard treatments for pain with help from a pain specialist as needed; standard treatments for transfusion-related infections contracted prior to virucidal treatment of plasma-derived concentrates.

Surveillance: For young children with severe or moderate hemophilia B, assessments every six to 12 months at an HTC; older children and adults with severe or moderate hemophilia B benefit from at least annual assessment at an HTC; for individuals with mild hemophilia B, assessment at an HTC every one to two years. Individuals with comorbidities may require more frequent visits. The assessment should include a review of bleeding episodes, adjustment of treatment plans as needed, a joint and muscle evaluation, an inhibitor screen, viral testing if indicated, and a discussion of any other issues related to the individual's hemophilia B as well as family and community support. Screening for alloimmune inhibitors is performed after treatment with factor IX concentrates has been initiated and in any individual with a suboptimal clinical response to treatment.

Agents/circumstances to avoid: Circumcision of at-risk males until hemophilia B is either excluded or treated with factor IX concentrate regardless of severity; activities with a high risk of trauma, particularly head injury; aspirin and all aspirin-containing products. Cautious use of other medications and herbal remedies that affect platelet function. Use precaution with intramuscular injections (apply pressure and ice; intramuscular injection may be scheduled after factor IX treatment or while on prophylaxis).

Evaluation of relatives at risk: It is appropriate to evaluate asymptomatic male and female at-risk relatives of an affected individual in order to identify as early as possible those who would benefit from prompt initiation of targeted therapy, supportive care, and surveillance. It is recommended that the genetic status of at-risk females be established prior to pregnancy or as early in a pregnancy as possible.

Pregnancy management: Maternal factor IX levels do not increase during pregnancy and heterozygous females may need factor IX infusion support for delivery and/or to treat or prevent postpartum hemorrhage. Heterozygous mothers should be monitored for delayed bleeding postpartum. Tranexamic acid can be used to prevent postpartum hemorrhage.

Other: Vitamin K does not prevent or control bleeding in hemophilia B.

Genetic counseling.

Hemophilia B is inherited in an X-linked manner. The risk to sibs of a male proband depends on the genetic status of the mother. The risk to sibs of a female proband depends on the genetic status of the mother and father. If the mother of the proband has an F9 pathogenic variant, the chance of the mother transmitting it in each pregnancy is 50%. If the father of the proband has an F9 pathogenic variant, he will transmit it to all his daughters and none of his sons. Males who inherit the pathogenic variant will be affected; females who inherit the pathogenic variant are heterozygotes and may be at risk for bleeding. Once the F9 pathogenic variant has been identified in an affected family member, genetic testing for at-risk family members, prenatal testing for a pregnancy at increased risk, and preimplantation genetic testing are possible.

Suggestive Findings

Hemophilia B should be suspected in a male or female proband with any of the following clinical features, laboratory features, and/or family history.

Clinical features

• Hemarthrosis, especially with mild or no antecedent trauma
• Deep-muscle hematomas
• Intracranial bleeding in the absence of major trauma
• Neonatal cephalohematoma or intracranial bleeding
• Prolonged oozing or renewed bleeding after initial bleeding stops following tooth extractions, mouth injury, or circumcision *
• Prolonged or delayed bleeding or poor wound healing following surgery or trauma *
• Unexplained gastrointestinal bleeding or hematuria *
• Heavy menstrual bleeding, especially with onset at menarche
• Prolonged nosebleeds, especially recurrent and bilateral *
• Excessive bruising, especially with firm, subcutaneous hematomas

* Of any severity, or especially in more severely affected persons

Laboratory features

• Normal platelet count
• Prolonged activated partial thromboplastin time (aPTT) in severe and moderate hemophilia B; normal or mildly prolonged aPTT in mild hemophilia B
• Normal prothrombin time

Family history is consistent with X-linked inheritance (e.g., no male-to-male transmission). Absence of a known family history does not preclude the diagnosis.

Establishing the Diagnosis

Male proband. The diagnosis of hemophilia B is established in a male proband by identification of decreased factor IX clotting activity.

• Severe hemophilia B. <1% factor IX clotting activity
• Moderate hemophilia B. 1%-5% factor IX clotting activity
• Mild hemophilia B. 6%-40% factor IX clotting activity

Note: (1) The normal range for factor IX clotting activity is approximately 50%-150% [Khachidze et al 2006]. Individuals with factor IX clotting activity higher than 40% usually have normal coagulation in vivo. (2) Somatic mosaicism in males with hemophilia B has been described [Ketterling et al 1999, Miller 2021].

Identification of a hemizygous pathogenic (or likely pathogenic) variant in F9 by molecular genetic testing can help predict the clinical phenotype, assess the risk of developing a factor IX inhibitor, and allow family studies (see Table 1).

Female proband. The diagnosis of hemophilia B may be established in a female proband with bleeding symptoms and decreased factor IX clotting activity, and/or by identification of a heterozygous pathogenic (or likely pathogenic) variant in F9 by molecular genetic testing (see Table 1). Note: Factor IX clotting activity does not reliably identify heterozygous females, as only approximately 30% of females heterozygous for an F9 pathogenic variant have factor IX clotting activity lower than 40% [Plug et al 2006].

Note: (1) Per ACMG/AMG variant interpretation guidelines, the terms "pathogenic variant" and "likely pathogenic variant" are synonymous in a clinical setting, meaning that both are considered diagnostic and can be used for clinical decision making [Richards et al 2015]. Reference to "pathogenic variant" in this GeneReview is understood to include any likely pathogenic variant. (2) Identification of a hemizygous or heterozygous F9 variant of uncertain significance does not establish or rule out the diagnosis.

Clinical Description

Hemophilia B in the untreated individual is characterized by spontaneous bleeding including intracranial bleeding, muscle and joint bleeding (usually in severe disease), immediate or delayed bleeding or prolonged oozing after injuries, tooth extractions, or surgery, or renewed bleeding after initial bleeding has stopped [Berntorp et al 2021, Mancuso et al 2021]. Intermittent oozing may last for days or weeks after tooth extraction. Prolonged or delayed bleeding or wound hematoma formation after surgery is common. After circumcision, males with hemophilia B of any severity may have prolonged oozing, or they may heal normally. In severe hemophilia B, spontaneous joint bleeding is the most frequent sign.

The age of diagnosis and frequency of bleeding episodes are generally related to the factor IX clotting activity (see Table 2). In any affected individual, bleeding episodes may be more frequent in childhood and adolescence than in adulthood. To some extent, this greater frequency is a function of both physical activity levels and vulnerability during more rapid growth.

Individuals with severe hemophilia B are usually diagnosed as newborns due to birth- or neonatal-related procedures or during the first year of life [Kulkarni et al 2009]. In untreated toddlers, bleeding from minor mouth injuries and large "goose eggs" from minor head bumps are common; these are the most frequent presenting symptoms of severe hemophilia B. Intracranial bleeding may also result from head injuries. The untreated child almost always has subcutaneous hematomas; some have been referred for evaluation of possible nonaccidental trauma.

As the child grows and becomes more active, spontaneous joint bleeds occur with increasing frequency unless the child is on a prophylactic treatment program. Spontaneous joint bleeds or deep-muscle hematomas initially cause pain or limping before swelling appears. Children and young adults with severe hemophilia B who are not treated have an average of two to five spontaneous bleeding episodes each month. Joints are the most common sites of spontaneous bleeding; other sites include the muscles, kidneys, gastrointestinal tract, brain, and nose. Without prophylactic treatment, individuals with hemophilia B have prolonged bleeding or excessive pain and swelling from minor injuries, surgery, and tooth extractions.

Individuals with moderate hemophilia B seldom have spontaneous bleeding, although there is significant variability between individuals, and bleeding episodes may be precipitated by relatively minor trauma. Without pretreatment (as for elective invasive procedures) they do have prolonged or delayed oozing after relatively minor trauma and are usually diagnosed before age five to six years. The frequency of bleeding episodes requiring treatment with factor IX concentrates varies from once a month to once a year. Signs and symptoms of bleeding are otherwise similar to those found in severe hemophilia B.

Individuals with mild hemophilia B do not have spontaneous bleeding. However, without treatment, abnormal bleeding occurs with surgery, tooth extractions, and moderate to major injuries. The frequency of bleeding may vary from once a year to once every ten years. Individuals with mild hemophilia B are often not diagnosed until later in life when they undergo surgery or tooth extraction or experience major trauma.

Heterozygous females with a factor IX clotting activity level lower than 40% are at risk for bleeding that is usually comparable to that seen in males with mild hemophilia B. However, more subtle abnormal bleeding may occur with baseline factor IX clotting activity levels between 30% and 60% [Plug et al 2006, van Galen et al 2021].

Complications of untreated bleeding. The leading cause of death related to bleeding is intracranial hemorrhage [Zwagemaker et al 2021]. The major cause of disability from bleeding is chronic joint disease [Berntorp et al 2021]. Currently available treatment with clotting factor IX concentrates is normalizing life expectancy and reducing chronic joint disease for children and adults with hemophilia B [Mancuso et al 2021]. Prior to the availability of such treatment, the median life expectancy for the most severely affected individuals was in childhood. Excluding death from HIV, life expectancy for those severely affected individuals receiving adequate treatment was 63 years in 2000 [Darby et al 2007]. A more recent analysis from the Netherlands found life expectancy in men with hemophilia to be 77 years, six years less than the Dutch male population [Hassan et al 2021].

Other. Since the late 1960s, the mainstay of treatment of bleeding episodes has been factor IX concentrates that initially were derived solely from donor plasma. By the late 1970s, more purified preparations became available, reducing the risk for thrombogenicity but allowing viral transmission. HIV transmission from concentrates occurred between 1979 and 1985. Approximately half of these individuals died of AIDS prior to the advent of effective HIV therapy.

Viral inactivation methods and donor screening of plasmas were introduced by 1990 and a recombinant factor IX concentrate became available shortly thereafter [Monahan & Di Paola 2010]. A second recombinant factor IX concentrate was licensed by the FDA in 2013. Three long-acting modified recombinant factor IX concentrates are now FDA approved, extending the factor IX half-life three- to fivefold compared to unmodified products [Hart et al 2022]. In November 2022 the FDA approved the first gene therapy product (etranacogene dezaparvovec) for adults with hemophilia B.

Hepatitis B transmission from earlier plasma-derived concentrates was eliminated with donor screening and then vaccination introduced in the 1970s. Most individuals exposed to plasma-derived concentrates prior to the late 1980s became chronic carriers of hepatitis C virus. Viral inactivation methods implemented in concentrate preparation and donor screening assays developed by 1990 have essentially eliminated hepatitis C transmission from plasma-derived concentrates.

Alloimmune inhibitors occur much less frequently in hemophilia B than in hemophilia A although are more common than previously appreciated. Earlier data suggested a rate of 3%-5%, but more recent data suggests that it may be closer to 10%, almost exclusively in individuals with severe disease. The incidence appears to vary by population and underlying genetic variant [Puetz et al 2014, Male et al 2021, Johnsen et al 2022, Kihlberg et al 2022]. These individuals usually have partial- or whole-gene deletions or certain nonsense variants (see Genotype-Phenotype Correlations and Table A, Locus-Specific Databases). At times, the onset of an alloimmune response has been associated with anaphylaxis to transfused factor IX or development of nephrotic syndrome [DiMichele 2007, Chitlur et al 2009].

Genotype-Phenotype Correlations

Disease severity

• Large deletions, nonsense variants, and most frameshift variants cause severe disease.
• Missense variants can cause severe, moderate, or mild disease depending on their location and the specific substitutions involved.

Alloimmune inhibitors

• Alloimmune inhibitors occur with the greatest frequency (40%-60%) in individuals with large partial (>50 bp) deletions, whole-gene deletions, or early-termination variants (<100 predicted amino acids) [Goodeve 2015, Saini et al 2015].
• Missense variants are rarely associated with inhibitors.

Unlike hemophilia A, severe hemophilia B is often caused by a missense variant, and several of these are associated with normal cross-reacting material (factor IX antigen) levels (see Table A, Locus-Specific Databases).

Uncommon variants within the carboxylase-binding domain of the propeptide cause increased sensitivity to warfarin anticoagulation in individuals without any baseline bleeding tendency [Oldenburg et al 2001] (see Management).

In hemophilia B Leyden, more than 20 different causative variants in the proximal F9 promoter region have been described [Funnell & Crossley 2014, Miller 2021]. The severity of disease decreases after puberty; mild disease disappears, and severe disease becomes mild, depending on the specific pathogenic variant.

Penetrance

All males with an F9 pathogenic variant are affected and will have hemophilia B of approximately the same severity as all other affected males in the family; however, other genetic and environmental effects may modify the clinical severity to some extent.

Approximately 30% of heterozygous females have factor IX clotting activity below 40% and are at risk for a bleeding disorder; mild bleeding can occur in carriers with low-normal factor IX activity [Plug et al 2006].

Nomenclature

Newly recommended terminology for heterozygous females designates five clinical- and laboratory-based categories [van Galen et al 2021]. For females with decreased (≤40%) factor IX clotting activity, the terminology is the same as that used for hemizygous males:

• Severe hemophilia B (<1% factor IX clotting activity)
• Moderate hemophilia B (1%-5% factor IX clotting activity)
• Mild hemophilia B (6%-40% factor IX clotting activity)

For heterozygous females with normal factor IX clotting activity:

• Individuals with a bleeding phenotype are termed "symptomatic hemophilia carriers";
• Individuals who do not have a bleeding phenotype are termed "asymptomatic hemophilia carriers."

Prevalence

The birth prevalence of hemophilia B has been calculated to be five in 100,000 live male births, and 1.5 in 100,000 for severe hemophilia B [Iorio et al 2019].

The birth prevalence is thought to be approximately the same in all countries and all ethnicities, presumably because of the high spontaneous mutation rate of F9 and its presence on the X chromosome.

Hemophilia B is about one fifth as prevalent as hemophilia A.

Evaluations Following Initial Diagnosis

To establish the extent of disease and needs in an individual diagnosed with hemophilia B, the evaluations summarized in Table 4 (if not performed as part of the evaluation that led to the diagnosis) are recommended.

Treatment of Manifestations

The World Federation of Hemophilia has published treatment guidelines for the management of individuals with hemophilia. Treatment should be coordinated through a hemophilia treatment center (HTC).

Surveillance

Persons with hemophilia followed at an HTC (see Resources) have lower mortality than those who are not [Soucie et al 2000, Pai et al 2016].

Young children with severe or moderate hemophilia B should be evaluated at an HTC (accompanied by their parents/guardians) every six to 12 months and as needed to review their history of bleeding episodes and adjust treatment plans. Early signs and symptoms of possible bleeding episodes are reviewed. The assessment should also include a joint and muscle evaluation, an inhibitor screen, viral testing if indicated, and a discussion of any other issues related to the individual's hemophilia B as well as family and community support.

Because of the risk of severe allergic reactions with development of alloantibodies, it is recommended that the first 20 factor replacement treatments be given in a medical setting where resuscitation medications and equipment are available. Risk can be stratified if the genetic variant is known. Those with large partial deletions, complete gene deletions, and early termination variants (<100 predicted amino acids) are at highest risk [Hart et al 2022]. Screening for alloimmune inhibitors is performed after treatment with factor IX concentrates has been initiated for either bleeding or prophylaxis (see Genotype-Phenotype Correlations). Testing for inhibitors should also be performed in any individual with hemophilia B whenever a suboptimal clinical response to treatment is suspected.

Older children and adults with severe or moderate hemophilia B benefit from at least annual assessment at an HTC (see Resources) and periodic assessments to review bleeding episodes and treatment plans, evaluate joints and muscles, screen for inhibitors, perform viral testing if indicated, provide education, and discuss other issues relevant to the individual's hemophilia B.

Individuals with mild hemophilia B can benefit from an assessment at an HTC every one to two years. Affected individuals with comorbidities and other complications or treatment challenges may require more frequent visits.

Agents/Circumstances to Avoid

The following agents/circumstances should be avoided:

• Infant males with a family history of hemophilia B should not be circumcised unless hemophilia B is excluded; or, if present, the infant should be treated with factor IX concentrate directly before and after the procedure.
• Use precaution with intramuscular injections without factor IX treatment. Pressure on the site after intramuscular injection in children has been reported without factor IX coverage. Individuals on prophylactic factor IX infusions may be given intramuscular injections after factor IX treatment or factor IX may be given specifically for this indication.
• Activities that involve a high risk of trauma, particularly of head injury, should be avoided.
• Medications and herbal remedies that affect platelet function, including aspirin, should be avoided unless there is strong medical indication, such as individuals with a cardiovascular indication. Individuals with severe hemophilia usually require clotting factor prophylaxis to allow aspirin and other platelet-inhibitory drugs to be used safely [Angelini et al 2016].

Older, intermediate-purity plasma-derived "prothrombin complex" concentrates should be used cautiously (if at all) in hemophilia B because of their thrombogenic potential.

Evaluation of Relatives at Risk

It is appropriate to evaluate asymptomatic male and female at-risk relatives of an affected individual in order to identify as early as possible those who would benefit from prompt initiation of targeted therapy, supportive care, and surveillance. A thorough family history may identify relatives who are at risk but have not been tested (particularly in families with mild hemophilia B).

Evaluation of at-risk males

• Assay of factor IX clotting activity from a cord blood sample obtained by venipuncture of the umbilical vein (to avoid contamination by amniotic fluid or placenta tissue), assessment of factor IX clotting activity in the neonatal period, or molecular genetic testing for the family-specific F9 pathogenic variant can establish or exclude the diagnosis of hemophilia B in newborn males at risk.
  Note: (1) The cord blood for factor IX clotting activity assay should be drawn into a syringe containing one tenth volume of sodium citrate to avoid clotting and to provide an optimal mixing of the sample with the anticoagulant. (2) Factor IX clotting activity in cord blood in a normal-term newborn is lower than in adults (mean: ~30%; range: 15%-50%); thus, the diagnosis of hemophilia B can be established in an infant with activity lower than 1% but is equivocal in an infant with moderately low (15%-20%) activity.
• Infants with a family history of hemophilia B should not be circumcised unless hemophilia B is either excluded or, if present, factor IX concentrate is administered immediately before and after the procedure to prevent delayed oozing and poor wound healing. The benefit versus the risk of exposure to factor IX concentrate in early childhood should be considered.

At-risk females. Approximately 30% of heterozygous females have factor IX clotting activity lower than 40% and may have abnormal bleeding. In a Dutch survey of heterozygous females, bleeding symptoms correlated with baseline factor clotting activity; there was suggestion of a very mild increase in bleeding even in those with 40% to 60% factor IX clotting activity [Plug et al 2006]. Joint range of motion in female carriers with factor VIII or factor IX activity lower than 40% was significantly different from that measured in normal controls and inversely related to factor level [Sidonio et al 2014].

• All daughters and mothers of an affected male and other at-risk females should have molecular genetic testing for the family-specific F9 pathogenic variant. Heterozygous females should have a baseline factor IX clotting activity assay to determine if they are at increased risk for bleeding.
• It is recommended that the genetic status of at-risk females be established prior to pregnancy or as early in a pregnancy as possible.

See Genetic Counseling for issues related to testing of at-risk relatives for genetic counseling purposes.

Pregnancy Management

Obstetric issues. It is recommended that the genetic status of a female at risk for hemophilia B be established prior to pregnancy or as early in a pregnancy as possible.

Unlike for factor VIII, maternal factor IX levels do not increase during pregnancy, and heterozygous females are more likely to need factor IX infusion support for delivery and/or to treat or prevent postpartum hemorrhage. In females with hemophilia B, postpartum hemorrhage has been a prominent feature, even in women without heavy menstrual bleeding [Yang & Ragni 2004]. To prevent postpartum hemorrhage, tranexamic acid 1 gm intravenously immediately following cord clamping and then orally for seven to 14 days postpartum or as needed with or without factor IX concentrate as indicated by factor IX clotting activity can be used.

If the female has a baseline factor IX clotting activity below approximately 40%, she by definition has hemophilia B and is at risk for excessive bleeding, particularly postpartum, and may require therapy with factor IX concentrate [Yang & Ragni 2004].

Newborn males. Controversy remains as to indications for cesarean section versus vaginal delivery [Leebeek et al 2020]. In a retrospective study of 580 males age birth to two years with hemophilia A and hemophilia B, 17 suffered intracranial hemorrhages with delivery, and all but one were delivered vaginally [Kulkarni et al 2009]. This finding supports the recommendation of cesarean section for infants with hemophilia; however, 12 of the 17 were born to women not known to be heterozygous, suggesting that a planned delivery may mitigate risks. A more recent large study showed a similar risk of intracranial hemorrhage after planned vaginal delivery as reported in the general population [Andersson et al 2019]. The relative risks of cesarean section versus vaginal delivery should be considered and discussed with the family and obstetrician so that a coordinated plan can be developed. Regardless of delivery mode, instrumentation with vacuum assistance or forceps must be avoided.

Therapies Under Investigation

Gene therapy for hemophilia B. Clinical trials for gene therapy for hemophilia B are under way, and one product was approved by the FDA in 2022. Those that are currently in or have recently completed Phase III clinical trials use liver-targeted adeno-associated vectors (AAV) and include the F9 Padua variant (p.Arg384Leu), which results in a transgene with increased specific activity. These vectors use liver-restricted promoters to target synthesis to the natural site of factor IX synthesis. Therapeutic levels have been achieved in many, although not all, individuals in studies to date, and short-term steroids are often needed for elevated transaminases. Long-term durability and safety need further study [Leebeek & Miesbach 2021].

Hemostasis rebalancing agents. Several products are under study that alter the balance of hemostasis so that individuals in which hemostasis is defective, such as those with hemophilia B, can have a more normal hemostatic response [Mancuso et al 2021].

Hemostasis rebalancing agents in late-phase clinical trials include antithrombin inhibitors and tissue factor pathway inhibitors. These have shown efficacy in hemophilia B and are particularly promising for individuals with hemophilia B and prevalent inhibitors.

Search ClinicalTrials.gov in the US and EU Clinical Trials Register in Europe for access to information on clinical studies for a wide range of diseases and conditions.

Other

Vitamin K does not prevent or control bleeding in hemophilia B.

Fresh frozen plasma is no longer recommended to treat hemophilia B because it is not treated with a virucidal agent.

Mode of Inheritance

Hemophilia B is inherited in an X-linked manner.

Risk to Family Members

Parents of a male proband

• The father of an affected male will not have the disease, nor will he be hemizygous for the F9 pathogenic variant; therefore, he does not require further evaluation/testing.
• In a family with more than one affected individual, the mother of an affected male is an obligate heterozygote. Note: If a female has more than one affected child and no other affected relatives and if the familial pathogenic variant cannot be detected in her leukocyte DNA, she most likely has germline mosaicism.
• Approximately 30% of affected males have no family history of hemophilia B. If a male is the only affected family member (i.e., a simplex case), it is possible that:
  • The mother is heterozygous for an F9 pathogenic variant.
  • The mother has somatic/germline mosaicism. Somatic mosaicism is reported in ≤11% of families [Ketterling et al 1999, Miller 2021].
    Note: Testing of maternal leukocyte DNA may not detect all instances of somatic mosaicism and will not detect a pathogenic variant that is present in the germ cells only.
  • The mother is not heterozygous for an F9 pathogenic variant, and the affected male has a de novo pathogenic variant.
• Molecular genetic testing of the mother is recommended to assess her genetic status and to allow reliable recurrence risk assessment.

Parents of a female proband

• A heterozygous female proband may have inherited the F9 pathogenic variant from either her mother or her father, or the pathogenic variant may be de novo.
• Detailed evaluation of the parents and review of the extended family history may help distinguish probands with a de novo pathogenic variant from those with an inherited pathogenic variant. Molecular genetic testing of the mother and the father can help determine if the pathogenic variant was inherited.

Sibs of a male proband. The risk to the sibs depends on the genetic status of the mother:

• If the mother of the proband has an F9 pathogenic variant, the chance of transmitting it in each pregnancy is 50%.
  • Males who inherit the pathogenic variant will be affected.
  • Females who inherit the pathogenic variant will be heterozygotes. Heterozygous females with a factor IX clotting activity level lower than 40% are at risk for bleeding that is usually comparable to that seen in males with mild hemophilia. However, more subtle abnormal bleeding may occur with baseline factor IX clotting activity levels between 30% and 60% [Plug et al 2006].
• All sibs should have factor IX clotting activity assayed unless molecular genetic testing confirms that they have not inherited the F9 pathogenic variant present in the family.

Sibs of a female proband. The risk to sibs depends on the genetic status of the mother and father:

• If the mother of the proband has an F9 pathogenic variant, the chance of the mother transmitting it in each pregnancy is 50% (see Sibs of a male proband).
• If the father of the proband has an F9 pathogenic variant, he will transmit it to all his daughters and none of his sons.

Offspring of a male proband. Affected males transmit the F9 pathogenic variant to all of their daughters and none of their sons.

Offspring of a female proband. Females with an F9 pathogenic variant have a 50% chance of transmitting the pathogenic variant to each child.

Other family members

• The maternal aunts and maternal cousins of a male proband may be at risk of having an F9 pathogenic variant.
• The risk to other family members of a female proband depends on the status of the proband's mother and father: if a parent has the F9 pathogenic variant, the parent's family members may be at risk.

Note: Molecular genetic testing can often determine the point of origin of a de novo pathogenic variant. Determining the point of origin of a de novo pathogenic variant is important for determining which branches of the family are at risk for hemophilia B.

Heterozygote Detection

Molecular genetic testing for identification of female heterozygotes is most informative if the F9 pathogenic variant has been identified in an affected family member. If an affected family member is not available for testing, molecular genetic testing can be performed first by sequence analysis, and if no pathogenic variant is identified, then by gene-targeted deletion/duplication analysis.

See Management, Evaluation of Relatives at Risk, At-risk females for information on evaluating at-risk female relatives for the purpose of early diagnosis and treatment.

Note: Factor IX clotting activity does not reliably identify heterozygous females, as only approximately 30% of females heterozygous for an F9 pathogenic variant have factor IX clotting activity lower than 40% [Plug et al 2006].

Related Genetic Counseling Issues

See Management, Evaluation of Relatives at Risk for information on evaluating at-risk relatives for the purpose of early diagnosis and treatment.

See the World Federation of Hemophilia treatment guidelines for recommendations regarding psychosocial support for individuals with hemophilia and their families.

Family planning

• The optimal time for determination of genetic risk and discussion of the availability of prenatal/preimplantation genetic testing is before pregnancy.
• It is recommended that the genetic status of a female at risk be established prior to pregnancy or as early in a pregnancy as possible (see Management, Pregnancy Management).
• It is appropriate to offer genetic counseling (including discussion of potential risks to offspring and reproductive options) to young adults who are affected, are heterozygous, or are at risk of being heterozygous.

Prenatal Testing and Preimplantation Genetic Testing

Once the F9 pathogenic variant has been identified in an affected family member, or if it cannot be identified but linkage can be established in the family, prenatal and preimplantation genetic testing are possible [Laurie et al 2010].

Differences in perspective may exist among medical professionals and within families regarding the use of prenatal testing. While most centers would consider use of prenatal testing to be a personal decision, discussion of these issues may be helpful.

Molecular Pathogenesis

Factor IX is synthesized in hepatocytes and circulates as a zymogen at 90 nmol/L (5 µg/mL). During coagulation initiation in vivo, it is activated by factor VIIa / tissue factor, and in coagulation amplification and propagation by factor IXa, in a reaction in which the activation peptide is cleaved. Activated factor IX is the intrinsic factor X activator, requiring its cofactor, activated factor VIII, a lipid surface, and calcium. Molecular interactions across multiple regions of the factor IXa molecule are involved in factor Xa activation [Kristensen et al 2016]. This activation is a critical early step that can regulate the overall rate of thrombin generation in coagulation.

Factor IX includes several distinct domains [Kanagasabai et al 2013, Rallapalli et al 2013]. From the 5' end, these domains are:

• Signal peptide and propeptide domain: cleaved to yield the mature protein, a secreted 415-amino acid peptide;
• GLA domain;
• Two domains homologous with epidermal growth factor;
• Connecting sequence: includes the activation peptide;
• Catalytic domain: typical of serine proteases.

Post-translational modifications include glycosylation, sulfation, phosphorylation, beta-hydroxylation, and gamma-carboxylation. A gamma-carboxylase binds to the propeptide before cleavage and, in a vitamin K-dependent step, converts the first 12 glutamic acid residues (near the amino-terminus) to gamma-carboxyglutamic residues (GLA domain). The GLA domain then binds calcium ions and adopts a conformation capable of binding to a phospholipid surface, where the clot initiation and propagation occurs.

Mechanism of disease causation. Loss of function

F9-specific laboratory technical considerations. Somatic mosaicism of F9 pathogenic variants have been described in males with hemophilia B [Ketterling et al 1999].

Author Notes

Washington Center for Bleeding Disorders provides comprehensive care for individuals with bleeding disorders across the state of Washington and through research and diagnostics works to advance the care of individuals with bleeding disorders.

Bloodworks Northwest laboratories provide specialty laboratory services including in hemostasis and hemostasis genomics to support hemophilia diagnosis and care. The laboratory served as the core laboratory for the My Life, Our Future program.

Barbara A Konkle (gro.dbcaw@elknok.arabrab) is actively involved in clinical research regarding individuals with hemophilia and would be happy to communicate with persons who have any questions regarding diagnosis of hemophilia or other considerations. Dr Konkle is also interested in hearing from clinicians treating families affected by hemophilia in whom no causative variant has been identified through molecular genetic testing of the genes known to be involved in this group of disorders.

Author History

Cheryl L Brower, RN, MSPH; Puget Sound Blood Center (2008-2011)
Frank K Fujimura, PhD, FACMG; GMP Genetics, Inc (2000-2003)
Haley Huston, BS; Bloodworks Northwest (2017-2023)
Maribel J Johnson, RN, MA; Puget Sound Blood Center (2000-2008)
Neil C Josephson, MD; Seattle Genetics (2011-2017)
Barbara A Konkle, MD (2011-present)
Shelley Nakaya Fletcher, BS (2011-present)
Arthur R Thompson, MD, PhD; University of Washington (2000-2014)

Revision History

• 9 February 2023 (sw) Comprehensive update posted live
• 15 June 2017 (sw) Comprehensive update posted live
• 5 June 2014 (me) Comprehensive update posted live
• 22 September 2011 (me) Comprehensive update posted live
• 8 April 2008 (me) Comprehensive update posted live
• 17 August 2005 (me) Comprehensive update posted live
• 11 May 2004 (cd) Revision: molecular genetic testing table
• 8 May 2003 (me) Comprehensive update posted live
• 2 October 2000 (me) Review posted live
• August 2000 (at) Original submission

Published Guidelines / Consensus Statements

Guidelines regarding genetic testing for this disorder have been published for the UK:

• Ludlam CA, Pasi KJ, Bolton-Maggs P, Collins PW, Cumming AM, Dolan G, Fryer A, Harrington C, Hill FG, Peake IR, Perry DJ, Skirton H, Smith M; UK Haemophilia Centre Doctors' Organisation. A framework for genetic service provision for haemophilia and other inherited bleeding disorders. Available online. 2005. Accessed 1-30-23.
• Mitchell M, Keeney S, Goodeve A. Practice guidelines for the molecular diagnosis of haemophilia B. UK Haemophilia Centre Doctors' Organisation, the Haemophilia Genetics Laboratory Network and the Clinical Molecular Genetics Society. Available online. 2010. Accessed 1-30-23.

Literature Cited

• Andersson NG, Chalmers EA, Kenet G, Ljung R, Makipernaa A, Chambost H, et al. Mode of delivery in hemophilia: vaginal delivery and Cesarean section carry similar risks for intracranial hemorrhages and other major bleeds. Haematologica. 2019;104:2100-6. [PMC free article: PMC6886417] [PubMed: 30792204]
• Angelini D, Konkle BA, Sood SL. Aging among persons with hemophilia: contemporary concerns. Semin Hematol. 2016;53:35-9. [PubMed: 26805905]
• Astermark J, Holstein K, Abajas YL, Kearney S, Croteau SE, Liesner R, Funding E, Kempton CL, Acharya S, Lethagen S, LeBeau P, Bowen J, Berntrop E, Shapiro AD. The C-Natural study – the outcome of immune tolerance induction therapy in patients with severe haemophilia B. Haemophilia. 2021;27:802-13. [PubMed: 34118102]
• Berntorp E, Fischer K, Hart DP, Mancuso ME, Stephensen D, Shapiro AD, Blanchette V. Haemophilia. Nature Reviews. 2021;7:45. [PubMed: 34168126]
• Caughey AB, Krist AH, Wolff TA, Barry MJ, Henderson JT, Owens DK, Davidson KW, Simon MA, Mangione CM. USPSTF approach to addressing sex and gender when making recommendations for clinical preventive services. JAMA. 2021;326:1953-61. [PubMed: 34694343]
• Chalmers E, Williams M, Brennand J, Liesner R, Collins P, Richards M, et al. Guideline on the management of haemophilia in the fetus and neonate. Br J Haematol. 2011;154:208-15. [PubMed: 21554256]
• Chitlur M, Warrier I, Rajpurkar M, Lusher JM. Inhibitors in factor IX deficiency a report of O the ISTH-SSC international FIX inhibitor registry (1997-2006). Haemophilia. 2009;15:1027-31. [PubMed: 19515028]
• Darby SC, Kan SW, Spooner RJ, Giangrande PL, Hill FG, Hay CR, Lee CA, Ludlam CA, Williams M. Mortality rates, life expectancy, and causes of death in people with hemophilia A or B in the United Kingdom who were not infected with HIV. Blood. 2007;110:815-25. [PubMed: 17446349]
• DiMichele D. Inhibitor development in haemophilia B: an orphan disease in need of attention. Br J Haematol. 2007;138:305-15. [PubMed: 17614818]
• Duga S, Salomon O. Congenital factor XI deficiency: an update. Semin Thromb Hemost. 2013;39:621-31. [PubMed: 23929304]
• Funnell APW, Crossley M. Hemophilia B Leyden and once mysterious cis-regulatory mutations. Trends Genet. 2014;30:18-23. [PubMed: 24138812]
• Goodeve AC. Hemophilia B: molecular pathogenesis and mutation analysis. J Thromb Haemost. 2015;13:1184-95. [PMC free article: PMC4496316] [PubMed: 25851415]
• Hart DP, Matino D, Astermark J, Dolan G, d’Oiron R, Hermans C, Jimenez-Yuste V, Linares A, Matsushita T, McRae S, Ozelo MC, Platton S, Staffor D, Sidonio Jr RF, Tiede A. International consensus recommendations on the management of people with haemophilia B. Ther Adv Hematol. 2022;13:20406207221085202. [PMC free article: PMC8980430] [PubMed: 35392437]
• Hassan S, Monahan RC, Mauser-Bunschoten EP, van Vulpen LFD, Eikenboom J, Beckers EAM, Hooimeiger L, Ympa PF, Nieuwenhuizen L, Coppens M, Schols SEM, Leebeek FWG, Smit C, Driessens MH, Le Cessie S, van Balen EC, Rosendaal FR, van der Bom JG, Gouw SC. Mortality, life expectancy, and causes of death of persons with hemophilia in the Netherlands 2001–2018. J Thromb Haemost. 2021;19:645-53. [PMC free article: PMC7986360] [PubMed: 33217158]
• Hay CR, Brown S, Collins PW, Keeling DM, Liesner R. The diagnosis and management of factor VIII and IX inhibitors: a guideline from the United Kingdom Haemophilia Centre Doctors Organisation. Br J Haematol. 2006;133:591-605. [PubMed: 16704433]
• Iorio A, Stonebraker JS, Chambost H, Makris M, Coffin D, Herr C, Germini F. Establishing the prevalence and prevalence at birth of hemophilia in males: a meta-analytic approach using national registries. Ann Int Med. 2019;171:540-6. [PubMed: 31499529]
• Johnsen JM, Fletcher SN, Dove A, McCracken H, Martin BK, Kircher M, Josephson NC, Shendure J, Ruuska SE, Valentino LA, Pierce FG, Watson C, Cheng D, Recht M, Konkle BA. Results of genetic analysis of 11,341 participants enrolled in the My Life, Our Future hemophilia genotyping initiative in the United States. J Thromb Haemost. 2022;20:2022-34. [PubMed: 35770352]
• Kanagasabai V, Schmidt AE, Marder VJ, Krishnaswamy S, Bajaj SP. Structure and function of vitamin K-dependent coagulant and anticoagulant proteins. In: Marder VJ, Aird WC, Bennett JS, Schulman S, White GC II, eds. Hemostasis and Thrombosis: Basic Principles and Clinical Practice. 6 ed. Philadelphia, PA: Lippincott-Williams & Wilkins; 2013:208-32.
• Ketterling RP, Bottema CD, Koeberl DD, Ii S, Sommer SS. T296----M, a common mutation causing mild hemophilia B in the Amish and others: founder effect, variability in factor IX activity assays, and rapid carrier detection. Hum Genet. 1991;87:333-7. [PubMed: 1864609]
• Ketterling RP, Vielhaber E, Li X, Drost J, Schaid DJ, Kasper CK, Phillips JA 3rd, Koerper MA, Kim H, Sexauer C, Gruppo R, Ambriz R, Paredes R, Sommer SS. Germline origins in the human F9 gene: frequent G:C-->A:T mosaicism and increased mutations with advanced maternal age. Hum Genet. 1999;105:629-40. [PubMed: 10647899]
• Khachidze M, Buil A, Viel KR, Porter S, Warren D, Machiah DK, Soria JM, Souto JC, Ameri A, Lathrop M, Blangero J, Fontcuberta J, Warren ST, Almasy L, Howard TE. Genetic determinants of normal variation in coagulation factor (F) IX levels: genome-wide scan and examination of the FIX structural gene. J Thromb Haemost. 2006;4:1537-45. [PubMed: 16839351]
• Kihlberg K, Baghaei F, Bruzelius M, Funding E, Holme PA, Lassila R, Martin M, Nummi V, Ranta S, Strandberg K, Andersson NG, Berntorp E, Astermark J. Factor IX antibodies and tolerance in hemophilia B in the Nordic countries – the impact of F9 variants and complications. Thromb Res. 2022;217:22-32. [PubMed: 35842956]
• Kristensen LH, Olsen OH, Blouse GE, Brandstetter H. Releasing the brakes in coagulation factor IXa by co-operative maturation of the substrate-binding site. Biochem J. 2016;473:2395-411. [PubMed: 27208168]
• Kulkarni R, Soucie JM, Lusher J, Presley R, Shapiro A, Gill J, Manco-Johnson M, Koerper M, Mathew P, Abshire T, Dimichele D, Hoots K, Janco R, Nugent D, Geraghty S, Evatt B, et al. Sites of initial bleeding episodes, mode of delivery and age of diagnosis in babies with haemophilia diagnosed before the age of 2 years: a report from the Centers for Disease Control and Prevention's (CDC) Universal Data Collection (UDC) project. Haemophilia. 2009;15:1281-90. [PubMed: 19637999]
• Laurie AD, Hill AM, Harraway JR, Fellowes AP, Phillipson GT, Benny PS, Smith MP, George PM. Preimplantation genetic diagnosis for hemophilia A using indirect linkage analysis and direct genotyping approaches. J Thromb Haemost. 2010;8:783-9. [PubMed: 20102489]
• Leebeek FWG, Duvekot J, Kruip MJHA. How I manage pregnancy in carriers of hemophilia and patients with von Willebrand disease. Blood. 2020;136:2143-50. [PubMed: 32797211]
• Leebeek FWG, Miesbach W. Gene therapy for hemophilia: a review on clinical benefit, limitations, and remaining issues. Blood. 2021;138:923-31. [PubMed: 34232980]
• Male C, Andersson NG, Rafowicz A, Liesner R, Kurnik K, Fischer K, Platokouki H, Santagostino E, Chambost H, Nolan B, Konigs C, Kenet G, Ljung R, van den Berg HM. Inhibitor incidence in an unselected cohort of previously untreated patients with severe hemophilia B: a PedNet study. Haematologica. 2021;106:123-9. [PMC free article: PMC7776246] [PubMed: 31919092]
• Manco-Johnson MJ, Abshire TC, Shapiro AD, Riske B, Hacker MR, Kilcoyne R, Ingram JD, Manco-Johnson ML, Funk S, Jacobson L, Valentino LA, Hoots WK, Buchanan GR, DiMichele D, Recht M, Brown D, Leissinger C, Bleak S, Cohen A, Mathew P, Matsunaga A, Medeiros D, Nugent D, Thomas GA, Thompson AA, McRedmond K, Soucie JM, Austin H, Evatt BL. Prophylaxis versus episodic treatment to prevent joint disease in boys with severe hemophilia. N Engl J Med. 2007;357:535-44. [PubMed: 17687129]
• Mancuso ME, Mahlangu JN, Pipe SW. The changing treatment landscape in haemophilia: from standard half-life clotting factor concentrates to gene editing. Lancet. 2021;397:630-40. [PubMed: 33460559]
• Menegatti M, Palla R, Boscarino M, Bucciareli P, Muszbek L, Katona E, Makris M, Peyvandi F. Minimal factor XIII activity level to prevent major spontaneous bleeds. J Thromb Haemost. 2017;15:1728-36. [PubMed: 28688221]
• Miller CH. The clinical genetics of hemophilia B (factor IX deficiency). Appl Clin Genet. 2021;14:445-54. [PMC free article: PMC8627312] [PubMed: 34848993]
• Mitchell M, Keeney S, Goodeve A. Practical guidelines for the molecular diagnosis of haemophilia B. UK Haemophilia Centre Doctors' Organisation, the Haemophilia Genetics Laboratory Network and the Clinical Molecular Genetics Society. Available online. 2010. Accessed 1-30-23.
• Monahan PE, Di Paola J. Recombinant factor IX for clinical and research use. Semin Thromb Hemost. 2010;36:498-509. [PubMed: 20632248]
• Nathwani AC. Gene therapy for hemophilia. Hematology Am Soc Hematol Educ Program. 2019;2019:1-8. [PMC free article: PMC6913446] [PubMed: 31808868]
• Oldenburg J, Kriz K, Wuillemin WA, Maly FE, von Welten A, Siegemun A, Keeling DM, Baker P, Chu K, Konkle BA, Lammie B, Albert T, et al. Genetic predisposition to bleeding during oral anticoagulant therapy: evidence for common founder mutations (FIXVal-10 and FIXThr-10) and an independent CpG hotspot mutation. Thromb Haemost. 2001;85:454-7. [PubMed: 11307814]
• Pai M, Key NS, Skinner M, Curtis R, Feinstein M, Kessler C, Lane SJ, Makris M, Riker E, Santesso N, Soucie JM, Yeung CHT, Iorio A, Schunemann HJ. NHF-McMaster guideline on care models for haemophilia management. Haemophilia. 2016;22:6-16. [PubMed: 27348396]
• Plug I, Mauser-Bunschoten EP, Brocker-Vriends AH, van Amstel HK, van der Bom JG, van Diemen-Homan JE, Willemse J, Rosendaal FR. Bleeding in carriers of hemophilia. Blood. 2006;108:52-6. [PubMed: 16551972]
• Puetz J, Soucie JM, Kempton CL, Monahan PE, et al. Prevalent inhibitors in haemophilia B subjects enrolled in the Universal Data Collection database. Haemophilia. 2014;20:25-31. [PMC free article: PMC4520536] [PubMed: 23855900]
• Rallapalli PM, Kemball-Cook G, Tuddenham EG, Gomez K, Perkins SJ. An interactive mutation database for human coagulation factor IX provides novel insights into the phenotypes and genetics of hemophilia B. J Thromb Haemost. 2013;11:1329-40. [PubMed: 23617593]
• Richards S, Aziz N, Bale S, Bick D, Das S, Gastier-Foster J, Grody WW, Hegde M, Lyon E, Spector E, Voelkerding K, Rehm HL, et al. Standards and guidelines for the interpretation of sequence variants: a joint consensus recommendation of the American College of Medical Genetics and Genomics and the Association for Molecular Pathology. Genet Med. 2015;17:405-24. [PMC free article: PMC4544753] [PubMed: 25741868]
• Row C, Chamouni P, Berger C, Lienhart A, Meunier S, Fretigny M, Dalibard V, Viprey M, Chambost H, Barbay V, Bovet J. Abnormal bleeding phenotype for mild haemophilia B patients with the p.Ile112Thr variation on the gene for factor IX. Haemophilia. 2021;27:e462-e465. [PubMed: 32996663]
• Saini S, Hamasaki-Katagiri N, Pandey GS, Yanover C, Guelcher C, Simhadri VL, Dandekar S, Guerrera MF, Kimchi-Sarfaty C, Sauna ZE. Genetic determinants of immunogenicity to factor IX during the treatment of haemophilia B. Haemophilia. 2015;21:210-8. [PubMed: 25470321]
• Sidonio RF, Mili FD, Li T, Miller CH, Hooper WC, DeBaun MR, Soucie M, et al. Females with FVIII and FIX deficiency have reduced joint range of motion. Am J Hematol. 2014;89:831-6. [PMC free article: PMC4477796] [PubMed: 24838518]
• Simioni P, Tormene D, Tognin G, Gavasso S, Bulato C, Iacobelli NP, Finn JD, Spiezia L, Radu C, Arruda VR. X-linked thrombophilia with a mutant factor IX (factor IX Padua). N Engl J Med. 2009;361:1671. [PubMed: 19846852]
• Soucie JM, Nuss R, Evatt B, Abdelhak A, Cowan L, Hill H, Kolakoski M, Wilber N. Mortality among males with hemophilia: relations with source of medical care. The Hemophilia Surveillance System Project Investigators. Blood. 2000;96:437-42. [PubMed: 10887103]
• Srivastava A, Santagostino E, Dougall A, Kitchen S, Sutherland M, Pipe SW, Carcao M, Mahlangu J, Ragni MV, Windyga J, Llinás A, Goddard NJ, Mohan R, Poonnoose PM, Feldman BM, Lewis SZ, van den Berg HM, Pierce GF, et al. WFH guidelines for the management of hemophilia, 3rd edition. World Federation of Hemophilia. Haemophilia. 2020;26:1-158.
• van Galen KPM, d’Orion R, James P, Abdul-Kadir R, Kouides PA, Kulkarni R, Mahlangu JN, Othman M, Peyvandi F, Rotellini D, Winikoff R, Sidonio RF. A new hemophilia carrier nomenclature to define hemophilia in women and girls: Communication from the SSC of the ISTH. J Thromb Haemost. 2021;19:1883-7. [PMC free article: PMC8361713] [PubMed: 34327828]
• Yang MY, Ragni MV. Clinical manifestations and management of labor and delivery in women with factor IX deficiency. Haemophilia. 2004;10:483-90. [PubMed: 15357775]
• Zwagemaker AF, Gouw SC, Jansen JS, Vuong C, Coppens M, Hu Q, Feng X, Kim SK, Van der Bom JG, Fijnvandraat K. Incidence and mortality rates of intracranial hemorrhage in hemophilia: a systematic review and meta-analysis. Blood. 2021;138:2853-73. [PubMed: 34411236]