Hemophilia A, Molecular Analysis for Inversion, Diagnosis and Carrier Detection
Clinical Information Discusses physiology, pathophysiology, and general clinical aspects, as they relate to a laboratory test
Hemophilia A is one of the most common inherited bleeding disorders in humans, affecting approximately 1 in 5,000 males. Common clinical symptoms for individuals with hemophilia A include prolonged oozing after tooth extraction, surgery or injuries, and renewed bleeding at wound sites. It is an X-linked condition in which carrier females are at risk to have affected sons but are typically asymptomatic themselves.
The clinical severity of hemophilia A correlates closely with the plasma concentration of factor VIII, an important factor in the coagulation process. Factor VIII activity levels of <1% are associated with severe disease, 1% to 5% activity with moderate disease, and 5% to 40% with mild hemophilia A. In males with severe deficiency, spontaneous bleeding into the joints can occur, resulting in a progressive arthropathy if untreated. Males with mild to moderate disease have variable onset of prolonged oozing and other bleeding issues but typically do not have spontaneous bleeding. Although rare, some carrier females may have mild symptoms. Approximately 10% of carrier females have a factor VIII clotting activity levels lower than 35% and are at risk for bleeding.
Factor VIII is encoded by the factor VIII gene (F8). Approximately 98% of patients with a diagnosis of hemophilia A are found to have a mutation in the F8 gene (ie, intron 1 and 22 inversions, point mutations, insertions, and deletions). Literature suggests that intron 22 inversions account for approximately 45% and intron 1 inversions account for approximately 2% to 3%, of mutations associated with severe hemophilia A. These inversions are typically not identified in patients with mild or moderate hemophilia A. Inversion analysis is the recommended first-tier molecular test for individuals with severe hemophilia A.
It is recommended that the F8 mutation be confirmed in the affected male or obligate carrier female prior to testing at-risk individuals. Obligate carrier females are identified by family history assessment and affected males by factor VIII activity and clinical evaluation. If this assay does not detect an inversion in these individuals, additional analysis (ie, F8 gene sequencing) may be able to identify the familial mutation. Of note, not all females with an affected son are germline carriers of an F8 mutation as de novo mutations in the F8 gene do occur. Approximately 20% of mothers of isolated cases do not have an identifiable germline F8 mutation. Importantly, there is a small risk for recurrence even when the familial F8 mutation is not identified in the mother of the affected patient due to the possibility of mosaicism.
Identifying the disease-causing mutation in males with severe hemophilia A
Determining hemophilia A-carrier status for individuals with a family history of F8 intron 1 or 22 inversions
Prenatal testing for hemophilia A when there is a high risk of inheriting an F8 intron 1 or 22 inversion and the fetus is male
An interpretive report will be provided that includes specimen information, pedigree information (when appropriate), assay information, whether test results are consistent with a diagnosis or positive carrier status, and an estimate of residual carrier risk, when appropriate.
Cautions Discusses conditions that may cause diagnostic confusion, including improper specimen collection and handling, inappropriate test selection, and interfering substances
The technology used in this assay detects only intron 1 and 22 inversion mutations. Thus, a negative result does not exclude the presence of other mutations in the F8 gene.
The intron 22 inversion detected in this direct analysis is found in about 45%, and intron 1 inversions account for approximately 2% to 3%, of individuals with severe hemophilia A; the assay may be uninformative for a number of families.
Any error in the diagnosis or in the pedigree provided will lead to an erroneous segregation pattern and an incorrect interpretation of the results.
Medical genetic or hemophilia center consultation is available for all possible hemophilia A cases and is particularly indicated in complex cases or in situations in which the diagnosis is atypical or uncertain.
Reference Values Describes reference intervals and additional information for interpretation of test results. May include intervals based on age and sex when appropriate. Intervals are Mayo-derived, unless otherwise designated. If an interpretive report is provided, the reference value field will state this.
An interpretive report will be provided which will include a risk analysis (probability of being a carrier).
Clinical References Provides recommendations for further in-depth reading of a clinical nature
1. Castaldo G, D'Argenio V, Nardiello P, et al: Haemophilia A: molecular insights. Clin Chem Lab Med 2007;45(4):450-461
2. Lakich D, Kazazian Jr HH, et al: Inversions disrupting the factor VIII gene are a common cause of severe hemophilia A. Nature Genetics 1993;5(3):236-241
3. Antonarakis SE, Rossiter JP, Young M, et al: Factor VIII gene inversions in severe hemophilia A: results of an international consortium study. Blood 1995;86(6):2206-2212