Anti-Remed with MECO MMSA: Scenario 5 (English) Tutorial 7th Block : Hemophilia A

Scenario 5 (English) Tutorial 7^th Block versi II

Author : Didit

Hemophilia A is characterized by deficiency in factor VIII 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 VIII clotting activity. In severe hemophilia A, spontaneous joint or deep-muscle bleeding is the most frequent symptom. Individuals with severe hemophilia A 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.

Individuals with moderate hemophilia A seldom have spontaneous bleeding; 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, usually from once a month to once a year.

Individuals with mild hemophilia A do not have spontaneous bleeding episodes; however, without pre- and postoperative treatment, abnormal bleeding occurs with surgery or tooth extractions; the frequency of bleeding episodes varies widely, typically from once a year to once every ten years. Individuals with mild hemophilia A are often not diagnosed until later in life.

Other. In any individual with hemophilia A, bleeding episodes may be more frequent in childhood and adolescence than in adulthood. Approximately 10% of carrier females are at risk for bleeding (even if the affected family member is mildly affected) and are thus symptomatic carriers, 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 A is established in individuals with low factor VIII clotting activity in the presence of a normal von Willebrand factor (VWF) level. Molecular genetic testing of F8, the gene encoding factor VIII, identifies disease-causing mutations in as many as 98% of individuals with hemophilia A. Such testing is available clinically.

Management.

Treatment of manifestations: Referral to one of the approximately 140 federally funded hemophilia treatment centers (HTCs) in the USA or worldwide for assessment, education, and genetic counseling and to facilitate management. Training and home infusions administered by parents followed by patient self-infusion are critical components of comprehensive care; especially for those with severe disease, intravenous infusion of plasma-derived factor VIII concentrate is most effective when infused within one hour of the onset of bleeding. For those with mild disease, including most symptomatic carriers, immediate treatment of bleeding or prophylaxis with intravenous or nasal desmopressin (DDAVP [1-deamino-8-D-arginine vasopressin]) or factor VIII concentrate.

Prevention of primary manifestations: For those with severe disease, prophylactic infusions of factor VIII concentrate three times a week or every other day usually maintain a “trough” factor VIII clotting activity higher than 1% and prevent spontaneous bleeding.

Prevention of secondary complications: Reduction of chronic joint disease by prompt effective treatment of bleeding, including home therapy.

Surveillance: For individuals with severe or moderate hemophilia A, annual assessments at an HTC are recommended; for individuals with mild hemophilia A, every two to three years; monitor carrier mothers for delayed bleeding post partum unless it is known that their baseline factor VIII clotting activity is normal.

Agents/circumstances to avoid: Circumcision of at-risk males until hemophilia A is either excluded or treated with factor VIII concentrate regardless of severity; intramuscular injections; activities with a high risk of trauma, particularly head injury; aspirin and all aspirin-containing products.

Testing of relatives at risk: To clarify genetic status of females at risk before pregnancy or early in pregnancy and to facilitate management.

Therapies under investigation: Ongoing clinical trials for a longer-acting factor VIII concentrate.

Other: Vitamin K does not prevent or control bleeding in hemophilia A; cryoprecipitate contains factor VIII but does not undergo viral inactivation so is no longer used to treat hemophilia A; no clinical trials for gene therapy in hemophilia A are currently in progress although several improved approaches are in pre-clinical testing.

Genetic counseling.

Hemophilia A is inherited in an X-linked manner. The risk to sibs of a proband depends on the carrier status of the mother. Carrier females have a 50% chance of transmitting the F8 mutation in each pregnancy: sons who inherit the mutation will be affected; daughters who inherit the mutation are carriers. Affected males transmit the mutation to all of their daughters and none of their sons. Carrier testing for at-risk family members and prenatal testing for pregnancies at increased risk are possible if the F8 disease-causing mutation has been identified in a family member or if informative intragenic linked markers have been identified.

Diagnosis

Clinical Diagnosis

A specific diagnosis of hemophilia A cannot be made on clinical findings. A coagulation disorder is suspected in individuals with any of the following:

· 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 GI bleeding or hematuria *

· Menorrhagia, especially with onset at menarche (in symptomatic carriers) *

· Prolonged nosebleeds, especially recurrent and bilateral *

· Excessive bruising, especially with firm, subcutaneous hematomas

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

Testing

Coagulation screening tests. Evaluation of an individual with a suspected bleeding disorder includes: platelet count and platelet function analysis (PFA closure times) or bleeding time; activated partial thromboplastin time (APTT); and prothrombin time (PT), a screen for the extrinsic clotting system. Thrombin time and/or plasma concentration of fibrinogen can be useful for rare disorders.

In individuals with hemophilia A, the above screening tests are normal, with the following exceptions:

· The APTT is prolonged in severe and moderate hemophilia A. Prolongations in APTT that correct on mixing with an equal volume of normal plasma indicate an intrinsic system clotting factor deficiency, including factor VIII, without an inhibitor.

Note: It is important to confirm the diagnosis of hemophilia A and to exclude other deficiencies with a specific factor VIII clotting activity assay, which is available in most hospital laboratories or coagulation reference laboratories.

· The APTT may be normal but is usually mildly prolonged in mild hemophilia A.

· The prothrombin time (PT) should be normal unless another hemostatic defect such as liver disease is present.

Note: In some clinical laboratories, the APTT is not sensitive enough to diagnose mild hemophilia A.

Coagulation factor assays. Individuals with a history of a lifelong bleeding tendency should have specific coagulation factor assays performed even if all the coagulation screening tests are in the normal range:

· The normal range for factor VIII clotting activity is approximately 50% to 150%.

· Individuals with factor VIII clotting activity higher than 30% usually do not have bleeding [Kaufman et al 2006]. However, a mild bleeding tendency can occur with low to low-normal factor VIII clotting activity in hemophilia A carrier females [Plug et al 2006] or in those with mild von Willebrand disease. The risk of having a bleeding tendency appears to be higher in carriers of alleles associated with severe hemophilia A, regardless of the baseline factor VIII clotting activity [Miesbach et al 2011].

· In hemophilia A, the factor VIII clotting activity is usually lower than 30%-35% with a normal, functional von Willebrand factor level.

· Classification of hemophilia A based on in vitro clotting activity:

o Severe hemophilia A. <1% factor VIII

o Moderate hemophilia A. 1%-5% factor VIII

o Mild hemophilia A. 6%-35% factor VIII

Note: Rarely, in individuals with mild hemophilia A, a standard "one-stage" factor VIII clotting activity assay shows near-normal or low-normal factor VIII clotting activity (40%-80%), whereas in a "two-stage" or chromogenic assay, factor VIII activity is low. Thus, low-normal in vitro clotting activity does not always exclude the presence of mild hemophilia A.

Carrier females

Coagulation factor assays. Approximately 10% of hemophilia A carrier females have factor VIII clotting activity lower than 35% regardless of the severity of hemophilia A in the family. Bleeding may also be more severe in those with low-normal factor VIII activity [Plug et al 2006].

Factor VIII clotting activity is unreliable in the detection of hemophilia A carriers:

· Factor VIII clotting activity in plasma is increased with pregnancy, oral contraceptive use, aerobic exercise, and chronic inflammation.

· Factor VIII clotting activity in plasma is approximately 25% lower in individuals of blood group O than in individuals of blood groups A, B, or AB.

· The majority of obligate carriers, even of severe hemophilia A, have normal factor VIII clotting activities.

Molecular Genetic Testing

Gene. F8 is the only gene in which mutations are known to cause hemophilia A.

Clinical testing

· Targeted mutation analysis

o An F8 intron 22-A inversion is identified in nearly half of families with severe hemophilia A [Kaufman et al 2006]. This inversion can be detected by Southern blotting or, more recently, by long-range PCR [Bagnall et al 2006] or inverse PCR [Rossetti et al 2008].

o An F8 intron 1 inversion is identified in 2%-3% of individuals with severe hemophilia A. This inversion is typically detected by PCR [Bagnall et al 2002].

· Sequence analysis

o The mutation detection rate in individuals with hemophilia A who do not have one of the two common inversions varies from 75% to 98%, depending on the testing methods used.

o In severe hemophilia A, gross gene alterations (including large deletions or insertions, frameshift and splice junction changes, and nonsense and missense mutations) of F8 account for approximately 50% of mutations detected [Kemball-Cook et al 1998, El-Maarri et al 2005, Kaufman et al 2006].

o In mild to moderate hemophilia A, missense mutations within the exons coding for the three A domains or the two C domains account for most of the mutations detected [Kemball-Cook et al 1998, Kaufman et al 2006].

· Deletion/duplication analysis

o In affected males deletion/duplication analysis can confirm the present of exonic, multiexonic, or larger deletions suspected on sequence analysis.

o In carrier females deletion/duplication analysis can detect gene deletions and rearrangements not detectable by sequence analysis [Santacroce et al 2009].

Table 1. Summary of Molecular Genetic Testing Used in Hemophilia A

Gene Symbol	Test Method	Mutations Detected	Mutation Detection Frequency by Test Method ¹				Test Availability
			Severe Hemophilia A		Moderate or Mild Hemophilia A
			Affected Males	Carrier Females	Affected Males	Carrier Females
F8	Targeted mutation analysis	Intron 22-A inversion ²	48% ³	48% ³	0% ³	0% ³	Clinical
	Targeted mutation analysis	Intron 1 inversion	2-3% ⁴	2-3% ⁴	0% ⁴	0% ⁴
	Sequence analysis / mutation scanning ⁵	Sequence variants ⁶	49% ^7,8	43% ⁸	76%-99% ^7,8	76%-98% ⁹
	Deletion / duplication analysis ¹⁰	Deletion / duplication of one or more exons or the whole gene	6%	6%	<1%	<1%

Test Availability refers to availability in the GeneTests^TM Laboratory Directory. GeneReviews designates a molecular genetic test as clinically available only if the test is listed in the GeneTests^TM Laboratory Directory by either a US CLIA-licensed laboratory or a non-US clinical laboratory. GeneTests does not verify laboratory-submitted information or warrant any aspect of a laboratory's licensure or performance. Clinicians must communicate directly with the laboratories to verify information.

1. The ability of the test method used to detect a mutation that is present in the indicated gene;

2. Intron 22 inversions can be accompanied by adjacent partial-gene deletions or duplication/insertions [Andrikovics et al 2003].;p

3. An intron 22-A inversion is identified in nearly half of families with severe hemophilia A [Kaufman et al 2006] and not identified in families with moderate or mild hemophilia A. Note: An uncommon exception occurs when severe hemophilia A is misdiagnosed as moderate hemophilia A, given the phenotypic variability among persons with null mutations;

4. An intron 1 inversion is identified in 2%-3% of individuals with severe hemophilia A [Bagnall et al 2002] and has not been described in families with moderate or mild hemophilia A;

5. Sequence analysis and mutation scanning of the entire gene can have similar detection frequencies; however, detection rates for mutation scanning may vary considerably among laboratories based on specific protocol used;

6. Examples of mutations detected by sequence analysis may include small intragenic deletions/insertions and missense, nonsense, and splice site mutations;

7. Lack of amplification by PCRs prior to sequence analysis suggests a deletion of one or more exons or the entire X-linked gene in a male; confirmation may require additional testing by using alternative amplification primers or deletion/duplication analysis;

8. Includes the mutation detection frequency using deletion/duplication analysis;

9. Sequence analysis of genomic DNA cannot detect deletion or duplication of one or more exons or the entire X-linked gene in a heterozygous female;

10. Testing that identifies deletions/duplications not readily detectable by sequence analysis of the coding and flanking intronic regions of genomic DNA; a variety of methods including quantitative PCR, long-range PCR, multiplex ligation-dependent probe amplification (MLPA), or targeted array GH (gene/segment-specific) may be used. A full array GH analysis that detects deletions/duplications across the genome may also include this gene/segment. See array GH.

Interpretation of test results. For issues to consider in interpretation of sequence analysis results, click here.

Linkage analysis is used to track an unidentified F8 disease-causing allele in a family and to identify the origin of de novo mutations:

· Tracking an unidentified F8 mutation. When a disease-causing mutation of F8 is not identified in an affected family member by direct DNA testing, linkage analysis can be considered to obtain information for genetic counseling in families in which more than one family member has the unequivocal diagnosis of hemophilia A. Linkage studies are always based on accurate clinical diagnosis of hemophilia A in the affected family members and accurate understanding of the genetic relationships in the family. In addition, linkage analysis depends on the availability and willingness of family members to be tested and on the presence of informative heterozygous polymorphic markers. Use of up to five intragenic variants and one extragenic variant is informative in approximately 80%-90% of families. Recombination events between F8 and the extragenic site occur in up to 5% of meioses, but have not been observed between hemophilia-causing mutations and intragenic sites.

· Identifying the origin of a de novo mutation. Among the nearly 50% of families with a simplex case of hemophilia A (i.e., occurrence in one family member only), the origin of a de novo mutation can often be identified by performing molecular genetic testing in conjunction with linkage analysis. The presence of the mutation on the affected individual's factor VIII haplotype is tracked back through the parents and, if necessary, through maternal grandparents to identify the individual in whom the mutation originated.

Testing Strategy

To confirm/establish the diagnosis of hemophilia A in a proband requires measurement of factor VIII clotting activity.

Molecular genetic testing is performed on a proband to detect the family-specific mutation in F8 in order to obtain information for genetic counseling of at-risk family members. If an affected individual is not available, an obligate carrier female can be tested.

In an individual who represents a simplex case, identification of the specific F8 mutation can help predict the clinical phenotype and assess the risk of developing a factor VIII inhibitor.

For (a) individuals with severe hemophilia A, (b) females with a family history of severe hemophilia A, or (c) females with a family history of hemophilia A of unknown severity in whom the family-specific mutation is not known, molecular genetic testing is generally performed in the following sequence until a mutation (or linkage) is identified:

· Targeted mutation analysis to identify the intron 22 or intron 1 inversion

· Sequence analysis of the 26 exons in F8

· Deletion/duplication analysis

· Linkage analysis

For (a) individuals with moderate or mild hemophilia A or (b) females with a family history of moderate or mild hemophilia A in whom the family-specific mutation is not known, molecular genetic testing is generally performed in the following sequence until a mutation is identified:

· Sequence analysis of F8

· Deletion/duplication analysis

· Linkage analysis

Note: When carrier testing is performed without previous identification of the F8 mutation in the family, a negative result in an at-risk relative is not informative.

Carrier testing for at-risk relatives is most informative after identification of the disease-causing mutation in the family. See above for testing of at-risk females when the family-specific mutation is not known.

Note: Carriers are heterozygotes for an X-linked disorder and may develop clinical findings related to the disorder.

Prenatal diagnosis and preimplantation genetic diagnosis (PGD) for at-risk pregnancies require prior identification of the disease-causing mutations in the family.

Note: It is the policy of GeneReviews to include clinical uses of testing available from laboratories listed in the GeneTests^TM Laboratory Directory; inclusion does not necessarily reflect the endorsement of such uses by the author(s), editor(s), or reviewer(s).

Genetically Related (Allelic) Disorders

No other phenotypes are associated with mutations in F8.

Clinical Description

Natural History

Hemophilia A in the untreated individual is characterized by delayed bleeding or prolonged oozing after injuries, tooth extractions, or surgery, or renewed bleeding after initial bleeding has stopped [Kessler & Mariani 2006]. Muscle hematomas or intracranial bleeding can occur four or five days after the original injury. 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 A of any severity may have prolonged oozing; but they can also heal normally without treatment. In severe hemophilia A, spontaneous joint bleeding is the most frequent symptom.

The age of diagnosis and frequency of bleeding episodes in the untreated individual are related to the factor VIII 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 A are usually diagnosed during the first year of life. On rare occasions, infants with severe hemophilia A have extra- or intracranial bleeding following birth. In untreated toddlers, bleeding from minor mouth injuries and large "goose eggs" from minor head bumps are common and are the most frequent presenting symptoms of severe hemophilia A. Intracranial bleeding may also result from head injuries. The untreated child almost always has subcutaneous hematomas; some have been referred for evaluation of possible non-accidental 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 adults with severe hemophilia A 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, but other sites include the kidneys, gastrointestinal tract, and brain. Without prophylactic treatment, individuals with hemophilia A have prolonged bleeding or excessive pain and swelling from minor injuries, surgery, and tooth extractions.

Individuals with moderate hemophilia A seldom have spontaneous bleeding but 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 VIII concentrates varies from once a month to once a year. Signs and symptoms of bleeding are otherwise similar to those found in severe hemophilia A.

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

Carrier females with a factor VIII clotting activity level lower than 35% 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 a baseline factor VIII clotting activity between 35% and 60% [Plug et al 2006].

Table 2. Symptoms Related to Severity of Untreated Hemophilia A

Severity	Factor VIII Clotting Activity ¹	Symptoms	Usual Age of Diagnosis
Severe	<1%	Frequent spontaneous bleeding; abnormal bleeding after minor injuries, surgery, or tooth extractions	Age ≤2 years ²
Moderately severe	1%-5%	Spontaneous bleeding is rare; abnormal bleeding after minor injuries, surgery, or tooth extractions	Age <5-6 years
Mild	>5%-35%	No spontaneous bleeding; abnormal bleeding after major injuries, surgery, or tooth extractions	Often later in life, depending on hemostatic challenges

1. Clinical severity does not always correlate with the in vitro assay result.

2. Kulkarni et al [2009]

Complications of untreated bleeding. The leading cause of death related to bleeding is intracranial hemorrhage. The major cause of disability from bleeding is chronic joint disease [Luck et al 2004]. Currently available treatment with clotting factor concentrates is normalizing life expectancy and reducing chronic joint disease for children with hemophilia A. Prior to the availability of such treatment, the median life expectancy for individuals with severe hemophilia A was 11 years (the current life expectancy for affected individuals in several developing countries). Excluding death from HIV, life expectancy for those severely affected individuals receiving adequate treatment is 63 years [Darby et al 2007].

Other. Since the mid-1960s, the mainstay of treatment of bleeding episodes has been factor VIII concentrates that initially were derived solely from donor plasma. Viral inactivation methods and donor screening of plasmas were introduced by the mid-1980s and recombinant factor VIII concentrates were introduced in the early 1990s, essentially ending the risk of HIV transmission. Many individuals who received plasma-derived factor VIII concentrates from 1979 to 1985 contracted HIV. Approximately half of these individuals died of AIDS prior to the advent of effective HIV therapy.

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

Approximately 30% of individuals with severe hemophilia A develop alloimmune inhibitors to factor VIII, usually within the first 20 exposures to infused factor VIII [Hay et al 2011] and, infrequently, in those who have received more than 50 exposures [Kempton 2010] (see Management, Treatment of Manifestations). Among individuals with hemophilia A, more blacks than whites develop the inhibitors, possibly as a result of differences in F8 haplotypes [Viel et al 2009].

Genotype-Phenotype Correlations

Disease severity

· F8 inversions are associated with severe hemophilia A and account for 45% of the severe cases [Kaufman et al 2006]. Of these, 20% to 30% develop alloimmune inhibitors. Occasionally, individuals considered to have moderate hemophilia A have been found to have F8 inversions. Often their assays have contained either some residual factor VIII clotting activity from a prior transfusion or the assay methods used were inaccurate at low levels.

· An inversion between a 1-kb sequence in intron 1 and an inverted repeat 5' to F8 [Bagnall et al 2002] is also associated with a severe phenotype, and some individuals have developed inhibitors.

· Point mutations leading to new stop codons are essentially all associated with a severe phenotype, as are most frameshift mutations. (An exception is the insertion or deletion of adenosine bases resulting in a sequence of eight to ten adenosines, which may result in moderate hemophilia A [Nakaya et al 2001].)

· Splice site mutations are often severe but may be mild, depending on the specific change and location.

· Missense mutations occur in fewer than 20% of individuals with severe hemophilia A but nearly all of those with mild or moderately severe bleeding

· A single base change in the 5’ promoter region of F8 has been associated with mild hemophilia A [Riccardi et al 2009].

Penetrance

All males with a F8 disease-causing mutation will be affected and will have approximately the same severity of disease as other affected males in the family. However, other genetic and environmental effects may modify the clinical severity somewhat.

Approximately 10% of females with one F8 disease-causing mutation and one normal allele have a factor VIII clotting activity lower than 30% and a bleeding disorder; mild bleeding can occur in carriers with low-normal factor VIII activity [Plug et al 2006].

Anticipation

Anticipation is not observed.

Prevalence

The birth prevalence of hemophilia A is approximately 1:4,000 to 1:5,000 live male births worldwide.

The birth prevalence is the same in all countries and all races, presumably because of a high spontaneous mutation rate in F8 and its presence on the X chromosome.

Prevalence is approximately 1:10,000 in the US and other countries in which optimum treatment with clotting factor concentrates is available [Kessler & Mariani 2006]; however, reporting varies widely [Stonebraker et al 2010].

Differential Diagnosis

When an individual presents with bleeding or the history of being a "bleeder," the first task is to determine if he/she truly has abnormal bleeding. "Bleeding a lot" during or immediately after major trauma, after a tonsillectomy, or for a few hours following tooth extraction may not be significant. In contrast, prolonged or intermittent oozing that lasts several days following tooth extraction or mouth injury, renewed bleeding or increased pain and swelling several days after an injury, or developing a wound hematoma several days after surgery almost always indicates a coagulation problem. A detailed history of bleeding episodes can help determine if the individual has a lifelong, inherited bleeding disorder or an acquired (often transient) bleeding disorder.

Physical examination provides few specific diagnostic clues. An older individual with severe or moderate hemophilia A may have joint deformities and muscle contractures. Large bruises and subcutaneous hematomas for which no trauma can be identified may be present, but individuals with a mild bleeding disorder have no outward signs except during an acute bleeding episode. Petechial hemorrhages indicate severe thrombocytopenia and are not a feature of hemophilia A.

A family history with a pattern of autosomal dominant, autosomal recessive, or X-linked inheritance provides clues to the diagnosis of the bleeding disorder but is not definitive. Hemophilia A and hemophilia B are both inherited in an X-linked manner. De novo F8 mutations occur and their origin can be documented in up to half of the families with newly diagnosed, affected members. Some families with mild hemophilia A are mistakenly diagnosed as having von Willebrand disease because both men and women have abnormal bleeding. With improved testing for von Willebrand disease, it is now possible to determine that women in such families often do not have von Willebrand disease, but rather are symptomatic carriers of hemophilia A.

Hemophilia A is only one of several lifelong bleeding disorders, and coagulation factor assays are the main tools for determining the specific diagnosis. Other inherited bleeding disorders associated with a low factor VIII clotting activity include the following:

· Mild (type 1) von Willebrand disease (VWD) accounts for 80% of individuals with VWD and is characterized by a quantitative deficiency of von Willebrand factor (low VWF antigen, factor VIII clotting activity, and ristocetin cofactor activity). Mucous membrane bleeding and prolonged oozing after surgery or tooth extractions are the predominant symptoms; laboratory testing is needed to differentiate mild hemophilia from VWD. Essentially all individuals with hemophilia A have a normal VWF level. Inheritance of VWD is autosomal dominant; penetrance varies.

· Type 2A or 2B VWD is characterized by a qualitative deficiency of VWF with a decrease of the high molecular weight multimers. VWF antigen and factor VIII clotting activity may be low-normal to mildly decreased. Functional VWF level is low in a ristocetin cofactor assay. Inheritance is autosomal dominant. Type 2B VWD is caused by a gain of function in platelet binding and is often accompanied by thrombocytopenia. Molecular genetic testing can aid in diagnosis.

· Type 2M VWD is also characterized by a qualitative deficiency of VWF with a similar gain of function in platelet binding as with type 2B; however, it is associated with a normal multimer pattern. Inheritance is autosomal dominant. Molecular genetic testing can aid in the diagnosis and distinction of subtypes of VWD type 2.

· Type 2N VWD is an uncommon variant caused by one of several missense mutations in the amino terminus of the circulating VWF protein, resulting in defective binding of factor VIII to VWF. Platelet function is completely normal. Clinically and biochemically, type 2N VWD is indistinguishable from mild hemophilia A; however, mild hemophilia A can be distinguished from type 2N VWD by molecular genetic testing of F8, molecular genetic testing of VWF, or measuring binding of factor VIII to VWF using ELISA or column chromatography. The low factor VIII clotting activity usually shows autosomal recessive inheritance.

· Severe, type 3 VWD is characterized by frequent episodes of mucous membrane bleeding and joint and muscle bleeding similar to that seen in individuals with hemophilia A. The VWF level is lower than 1% and the factor VIII clotting activity is 2%-8%. Inheritance is autosomal recessive. Parents may have type 1 VWD but more often are asymptomatic.

· Mild combined factor V and factor VIII deficiencies are usually caused by rare autosomal recessive inheritance of a deficiency of one of two intracellular chaperone proteins encoded by LMAN1 or MCFD2 [Zhang et al 2008].

The following are other bleeding disorders with normal factor VIII clotting activity:

· Hemophilia B is clinically indistinguishable from hemophilia A. Diagnosis is based on a factor IX clotting activity lower than 30%. Inheritance is X-linked.

· Factor XI deficiency is inherited in an autosomal recessive manner with heterozygotes showing a factor XI coagulant activity of 25% to 75% of normal, while homozygotes have activity of lower than 1% to 15% [Thompson 2006]. Two mutations are common among individuals of Ashkenazi Jewish descent. Both compound heterozygotes and homozygotes may exhibit bleeding similar to that seen in mild or moderate hemophilia A. A specific factor XI clotting assay establishes the diagnosis.

· Factor XII, prekallekrein, or high molecular weight kininogen deficiencies do not cause clinical bleeding but can cause a long activated partial thromboplastin time (APTT).

· Prothrombin (factor II), factor V, factor X, and factor VII deficiencies are rare bleeding disorders inherited in an autosomal recessive manner [Thompson 2006]. Individuals may display easy bruising and hematoma formation, epistaxis, menorrhagia, and bleeding after trauma and surgery. Hemarthroses are uncommon. Spontaneous intracranial bleeding can occur. Factor VII deficiency should be suspected if the PT is prolonged and APTT normal. Individuals with deficiency of factors II, V, or X usually have prolonged PT and APTT, but specific coagulation factor assays establish the diagnosis. Combined (multiple) deficiencies are usually acquired disorders, although a few families have hereditary deficits of the vitamin K-dependent factors, often resulting from deficiency of gamma-carboxylase.

· Fibrinogen disorders can be severe, mild, or asymptomatic [Thompson 2006]:

o Congenital afibrinogenemia is a rare disorder inherited in an autosomal recessive manner with manifestations similar to hemophilia A except that bleeding from minor cuts is prolonged because of the lack of fibrinogen to support platelet aggregation.

o Hypofibrinogenemia can be inherited either in an autosomal dominant or autosomal recessive manner and is usually asymptomatic but may be combined with dysfibrinogenemia.

o Dysfibrinogenemia is inherited in an autosomal dominant manner. Individuals with hypofibrinogenemia or dysfibrinogenemia have mild to moderate bleeding symptoms or may be asymptomatic; some individuals with dysfibrinogenemia are at risk for thrombosis. Diagnosis is based on kinetic being lower than antigenic protein levels, although the thrombin time is usually prolonged and is a simple screening test.

· Factor XIII deficiency is a rare autosomal recessive disorder [Thompson 2006]. Umbilical stump bleeding occurs in more than 80% of individuals. Intracranial bleeding that occurs spontaneously or following minor trauma is seen in 30% of individuals. Subcutaneous hematomas, muscle hematomas, defective wound healing, and recurrent spontaneous abortion are also seen. Joint bleeding is rare. All kinetic coagulation screening tests are normal; a specific test for clot solubility must be performed.

· Platelet function disorders cause bleeding problems similar to those seen in individuals with thrombocytopenia. Individuals have skin and mucous membrane bleeding, recurring epistaxis, gastrointestinal bleeding, menorrhagia, and excessive bleeding during or immediately after trauma and surgery. Joint, muscle, and intracranial bleeding is rare. Diagnosis is made utilizing platelet aggregation assays and flow cytometry.

o Bernard-Soulier syndrome is inherited in an autosomal recessive manner and involves the VWF receptor, the platelet membrane GPIb-IX complex.

o Glanzmann's thrombasthenia, also autosomal recessive, involves the GPIIb-IIIa receptor necessary for platelet aggregation. Abnormal platelet function is usually associated with a prolonged bleeding time or prolonged closure time on platelet function analysis.

· Hemophilia A: males

· Hemophilia A: female heterozygotes

Management

Evaluations Following Initial Diagnosis

To establish the extent of disease in an individual diagnosed with hemophilia A, the following evaluations are recommended:

· Identification of the specific F8 disease-causing mutation in an individual to aid in determining disease severity, the likelihood of inhibitor development, and the chance that immune tolerance will be successful if an inhibitor does develop

· A personal and family history of bleeding to help predict disease severity

· A joint and muscle evaluation, particularly if the individual describes a history of hemarthrosis or deep-muscle hematomas

· Screening for hepatitis A, B, and C, as well as HIV, particularly if blood products or plasma-derived clotting factor concentrates were administered prior to 1985

· Baseline CBC and platelet count, especially if there is a history of nose bleeds, GI bleeding, mouth bleeding; or in women, menorrhagia or postpartum hemorrhage

Treatment of Manifestations

In developed countries, life expectancy for individuals with hemophilia A has greatly increased over the past four decades [Darby et al 2007]; disability has decreased with the intravenous infusion of factor VIII concentrate, home infusion programs, prophylactic treatment, and improved patient education.

Individuals with hemophilia A benefit from referral for assessment, education, and genetic counseling at one of the approximately 140 federally funded hemophilia treatment centers (HTCs) in the USA that can be located through the National Hemophilia Foundation. Worldwide, treatment centers can be found through the World Federation of Haemophilia. The treatment centers establish appropriate treatment plans and provide referrals or direct care for individuals with inherited bleeding disorders. They also are a resource for current information on new treatment modalities for hemophilia. An assessment at one of these centers usually includes extensive patient education, genetic counseling, and laboratory testing.

Intravenous infusion of factor VIII concentrate. Recombinant factor VIII concentrates have been available for more than 15 years; some recombinant products now contain no human- or animal-derived proteins. Virucidal treatment of plasma-derived concentrates has eliminated the risk of HIV transmission since 1985, and of hepatitis B and C viruses since 1990.

Bleeding episodes are prevented or controlled quickly with intravenous infusions of either plasma-derived or recombinant factor VIII concentrate. Fast, effective treatment of bleeding episodes decreases pain and disability and reduces the risk of chronic joint disease. Ideally, the affected individual should receive clotting factor within an hour of noticing symptoms [Kessler & Mariani 2006]. Doses vary among individuals, but knowledge of a single in vivo recovery value does not always help in determining the appropriate dose [Bjorkman et al 2007]:

· Arranging efficient, effective treatment for infants and toddlers is especially challenging. Because frequent venipunctures may be necessary, it is important to identify staff members who are expert in performing venipunctures in small children.

· It is recommended that the parents of children age two to five years with severe hemophilia A be trained to administer the infusions as soon as it is feasible. Home treatment allows for prompt treatment after symptoms occur and facilitates prophylactic therapy.

DDAVP (1-deamino-8-D-arginine vasopressin). For many individuals with mild hemophilia A, including most symptomatic carriers, immediate treatment of bleeding or prophylaxis can be achieved with desmopressin (DDAVP) [Castaman et al 2009]. A single intravenous dose often doubles or triples factor VIII clotting activity. Alternatively, a multi-use, nasal formulation of desmopressin (Stimate^®) is more convenient and available.

Pediatric issues. Special considerations for care of infants and children with hemophilia A include the following [Chalmers et al 2005]:

· Infant males with a family history of hemophilia A should not be circumcised unless hemophilia A is either excluded or, if present, is treated with factor VIII concentrate directly before and after the procedure to prevent delayed oozing and poor wound healing.

· Intramuscular injections should be avoided; immunizations should be administered subcutaneously.

· Effective dosing of factor VIII requires an understanding of different pharmacokinetics in young children.

Inhibitors. Alloimmune inhibitors to factor VIII greatly compromise the ability to manage bleeding episodes [Hay et al 2006, Kessler & Mariani 2006]. High titer inhibitors can often be eliminated by immune tolerance therapy. Individuals with large gene deletions are less likely to respond to immune tolerance than individuals with other types of mutations [Peyvandi et al 2006, Coppola et al 2009].

Prevention of Primary Manifestations

Children with severe hemophilia A are often given "primary" prophylactic infusions of factor VIII concentrate three times a week or every other day to maintain factor VIII clotting activity above 1%; these infusions prevent spontaneous bleeding and decrease the number of bleeding episodes. Prophylactic infusions almost completely eliminate joint bleeding and greatly decrease chronic joint disease.

Prevention of Secondary Complications

Prevention of chronic joint disease is a major concern. It is agreed that most individuals with severe hemophilia A benefit from primary prophylaxis, but controversy still exists about when these regular infusions should begin. The age at which a child experiences the first joint bleed can vary greatly. Prophylactic infusions almost completely eliminate spontaneous joint bleeding, decreasing chronic joint disease, although complications of venous access ports in young children can occur [Feldman et al 2006, Manco-Johnson et al 2007].

"Secondary" prophylaxis is often used for several weeks if recurrent bleeding in a "target" joint or synovitis occurs, or for longer periods in adults with frequent bleeding.

Surveillance

Persons with hemophilia who are followed at hemophilia treatment centers (HTCs) (see Resources) have lower mortality than those who are not [Soucie et al 2000].

It is recommended that young children with severe or moderate hemophilia A have assessments at an HTC (accompanied by their parents) every six to 12 months to review their history of bleeding episodes and to adjust treatment plans as needed. 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 problems related to the individual's hemophilia and family and community support.

Screening for alloimmune inhibitors is usually done in individuals with severe hemophilia A every three to six months after treatment with factor VIII concentrates has been initiated either for bleeding or prophylaxis. After 50 to 100 exposure days, annual screening is sufficient; in adults, it is usually performed only prior to any elective surgery. Testing for inhibitors should also be performed in any individual with hemophilia whenever a sub-optimal clinical response to treatment is suspected, regardless of disease severity.

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

Individuals with mild hemophilia A can benefit from maintaining a relationship with an HTC and having regular assessments every two to three years.

Agents/Circumstances to Avoid

Avoid the following:

· Activities that involve 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 is indicated.

Testing of Relatives at Risk

Identification of at-risk relatives. A thorough family history may identify other male relatives who are at risk but have not been tested (particularly in families with mild hemophilia A).

Early determination of the genetic status of males at risk. Either assay of factor VIII clotting activity from a cord blood sample obtained by venipuncture of the umbilical vein (to avoid contamination by amniotic fluid or placenta tissue) or molecular genetic testing for the family-specific F8 mutation can establish or exclude the diagnosis of hemophilia A in newborn males at risk. Infants with a family history of hemophilia A should not be circumcised unless hemophilia A is either excluded or, if present, factor VIII concentrate is administered immediately before and after the procedure to prevent delayed oozing and poor wound healing.

Note: The cord blood for factor VIII 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.

Determination of genetic status of females at risk. Approximately 10% of carriers have factor VIII activity lower than 30%-35% and may have abnormal bleeding themselves. In a survey of Dutch hemophilia carriers, 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 VIII activity [Plug et al 2006]. Therefore, all daughters and mothers of an affected male and other at-risk females should have a baseline factor VIII clotting activity assay to determine if they are at increased risk for bleeding (unless they are known to be non-carriers based on molecular genetic testing). Very occasionally, a woman will have particularly low factor VIII clotting activity that may result from heterozygosity for an F8 mutation associated with skewed X-chromosome inactivation or, on rare occasion, compound heterozygosity for two F8 mutations [Pavlova et al 2009].

It is recommended that the carrier status of a woman at risk be established prior to pregnancy or as early in a pregnancy as possible.

Pregnancy Management

Obstetric issues. It is recommended that the carrier status of a woman at risk be established prior to pregnancy or as early in a pregnancy as possible.

If the mother is a symptomatic carrier (i.e., has baseline factor VIII clotting activity <35%), she will be somewhat protected by the natural rise of factor VIII clotting activity during pregnancy, which may even double by the end of the third trimester. However, postpartum factor VIII clotting activity can return to baseline within 48 hours, and delayed bleeding may ensue [Lee et al 2006].

Newborn males. Controversy remains as to indications for Cesarean section versus vaginal delivery [James & Hoots 2010, Ljung 2010]. For elective deliveries, the relative risks of Cesarean section versus vaginal delivery should be considered, especially if a male has been diagnosed with hemophilia A prenatally.

At birth or in the early neonatal period, intracranial hemorrhage in affected males is uncommon (1%-2%), even in males with severe hemophilia A who are delivered vaginally.

Therapies Under Investigation

Longer-acting factor VIII concentrates are undergoing clinical trials. The hope is that one infusion a week rather than three to four infusions a week will provide prophylaxis against spontaneous bleeding [Spira et al 2010].

Attempts are being made to learn more about the immunology of inhibitors and ways to prevent them or improve the success rate of immune tolerance [Lollar 2006, Zakarija et al 2011].

All clinical trials for gene therapy in hemophilia A have been discontinued because of complications and failure to achieve significant factor VIII expression in humans with hemophilia A. Although the hemophilia community remains hopeful, several obstacles must be overcome before new trials can begin with factor VIII [Pierce et al 2007].

Other

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

Cryoprecipitate is no longer recommended to treat hemophilia A because it is not treated with a virucidal agent.

Genetics clinics, staffed by genetics professionals, provide information for individuals and families regarding the natural history, treatment, mode of inheritance, and genetic risks to other family members as well as information about available consumer-oriented resources. See the GeneTests Clinic Directory.

Mode of Inheritance

Hemophilia A 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 a carrier of the mutation.

· Women who have an affected son and one other affected relative in the maternal line are obligate carriers.

· If a woman has more than one affected son and the disease-causing mutation cannot be detected in her DNA, she has germline mosaicism.

· One third to one half of affected males have no family history of hemophilia A. If an affected male represents a simplex case (an affected male with no known family history of hemophilia), several possibilities regarding his mother's carrier status and the carrier risks of extended family members need to be considered:

o The mother is not a carrier and the affected male has a de novo disease-causing mutation. Somatic mosaicism may occur in as many as 15% of probands with a point mutation and no known family history of hemophilia A [Leuer et al 2001]; germline mosaicism is rare.

o The mother is a carrier of a de novo, disease-causing mutation that occurred in one of the following ways:

§ As a germline mutation (i.e., in the egg or sperm at the time of her conception and thus present in every cell of her body and detectable in her DNA). Ninety-eight percent of mothers of a simplex case with an intron 22 inversion are carriers because most of these mutations occur in spermatogenesis.

§ As a somatic mutation (i.e., a change that occurred very early in embryogenesis, resulting in somatic mosaicism in which the mutation is present in some but not all cells and may or may not be detectable in DNA).

§ As germline mosaicism (in which some germ cells have the mutation and some do not, and in which the mutation is not detectable in DNA from her leukocytes).

o The mother is a carrier and has inherited the disease-causing mutation either from her mother who has a de novo disease-causing mutation or from her asymptomatic father who is mosaic for the mutation.

o The mother is a carrier of a mutation arising in a previous generation, which has been passed on through the family without manifesting symptoms in female carriers.

Overall, the mother has an approximately 80% chance of being a carrier when her son is the first affected individual in the family; however, the mother of a severely affected male with an intron 22 inversion has a 98% chance of being a carrier.

· Molecular genetic testing combined with linkage analysis can often determine the point of origin of a de novo mutation. Determining the point of origin of a de novo mutation is important for determining which branches of the family are at risk for hemophilia A.

Sibs of a male proband

· The risk to the sibs depends on the mother's carrier status. If the proband's mother is a carrier, each male sib is at a 50% risk of having hemophilia A and each female sib is at a 50% risk of being a carrier.

· Germline mosaicism is possible, albeit uncommon. Thus, if an affected male represents a simplex case and if his mother has a normal factor VIII clotting activity and no evidence of her son's F8 disease-causing mutation in DNA from her leukocytes, she is still at a theoretically increased (but low) risk of having additional affected children.

· All sibs should have factor VIII clotting activity assayed unless mutation analysis confirms that they have not inherited the F8 mutation in their family.

Offspring of a male proband

· All daughters will be carriers of the F8 mutation causing hemophilia A of the same severity as their father's hemophilia.

· No sons will inherit the mutant F8 allele, have hemophilia A, or pass it on to their offspring.

Other family members of the proband. The proband's maternal aunts and their offspring may be at risk of being carriers or being affected (depending on their gender, family relationship, and the carrier status of the proband's mother).

Carrier Detection

Carrier testing by molecular genetic testing is clinically available for most at-risk females if the mutation has been identified in the family.

Factor VIII clotting activity, or its ratio to von Willebrand factor level, is not a reliable test for determining carrier status: it can only be suggestive if low.

Prenatal Testing

Molecular genetic testing. Prenatal testing is available for pregnancies of women who are carriers if the mutation has been identified in a family member or if linkage has been established in the family. Of note, prenatal diagnosis of recurrent F8 inversions has been simplified using a PCR-based “inverse shifting” procedure [Radic et al 2009]. The usual procedure is to determine fetal sex by performing chromosome analysis of fetal cells obtained by chorionic villus sampling (CVS) at approximately ten to 12 weeks' gestation or by amniocentesis usually performed at approximately 15 to 18 weeks' gestation. If the karyotype is 46,XY, DNA extracted from fetal cells can be analyzed for the known F8 disease-causing mutation or for the informative markers.

Note: Gestational age is expressed as menstrual weeks calculated either from the first day of the last normal menstrual period or by ultrasound measurements.

Percutaneous umbilical blood sampling (PUBS). If the disease-causing F8 mutation is not known and linkage is not informative, prenatal diagnosis is possible using a fetal blood sample obtained by PUBS at approximately 18 to 21 weeks' gestation for assay of factor VIII clotting activity.

Requests for prenatal testing for conditions which (like hemophilia A) do not affect intellect and have treatment available are not common. Differences in perspective may exist among medical professionals and within families regarding the use of prenatal testing, particularly if the testing is being considered for the purpose of pregnancy termination rather than early diagnosis. Although most centers would consider decisions about prenatal testing to be the choice of the parents, discussion of these issues is appropriate.

Preimplantation genetic diagnosis (PGD) may be available for families in which the disease-causing mutation has been identified. Successful PGD has been reported in hemophilia A [Laurie et al 2010].