Factor II (Prothrombin)

Clinical Usage

  • Determine risk of venous thromboembolism, especially for patients that will receive medications (for example, oral contraceptives or other hormone treatments for women) or other treatments that will increase their risk of thromboembolism
  • Determine cause of venous thromboembolism

Background Information

Venous thromboembolism (VTE) is a common chronic disease with a risk of recurrence. One of the risk factors for VTE is a genetic variant of Factor II, or prothrombin.

Clotting factor II, or prothrombin, is a vitamin K–dependent proenzyme that functions in the blood coagulation cascade. Alterations in the Factor II (prothrombin) gene, (G20210A or A19922G) result in an increased expression of prothrombin which is linked to an increased risk of thrombosis. Prothrombin is synthesized in the liver and is a precursor to the serine protease thrombin. Increased blood concentration of prothrombin leads to increased rates of thrombin generation. Thrombin cleaves fibrinogen to fibrin, which then activates platelets and other blood clotting factors and results in excessive growth of fibrin clots.

The frequency of the G20210A mutation (A substitution) of Factor II is higher in patients with thromboembolic disease. The A19911G mutation (G substitution) is also associated with a slightly increased risk of thrombosis when accompanied by the G20210A mutation. Individuals with the A19911G mutation of Factor II plus Factor V Leiden mutation also have an elevated risk of developing thrombosis. Prothrombin-related thrombophilia is inherited in an autosomal dominant manner and the risk for thrombosis is more severe for those who have 2 copies of the mutant gene (homozygous) than for those carrying only a single gene copy (heterozygous). Multiple genetic risk factors for thrombosis are additive.

Gene Information

The Factor II gene is located on the short (p) arm of chromosome 11 at position 11. More precisely, the F2 gene is located from base pair 46,740,742 to base pair 46,761,055 on chromosome 11.

The G20210A variant is in the 3′-untranslated region (3’UT) of the prothrombin gene, G to A substitution at nucleotide position 20210.  
The A1991G variation is an A to G substitution at nucleotide position 1991 of the prothrombin gene.

Population Information

Approximately 95 – 99% of the American population is homozygous wild type for the G20210A  variant of Factor II.
Approximately 2-5 percent of Caucasians and 0.3 percent of African Americans are heterozygous for G20210A.
Approximately 0.01% of the population is homozygous for the G20210 mutant form of Factor II.
The G20210A variant is rare in Asians and native Americans.

The homozygous wild type of A19911G is found in approximately 20 – 25% of the American population.
Approximately 50 – 55% of the population are heterozygous for A19911G.
Approximately 15 – 20% of the population are homozygous for the A19911G mutant form of Factor II. 

Test Method

This test was developed using CLSI guidelines.  Control DNA samples of known genotype are tested together with each patient sample to ensure correct results.  Genomic DNA is extracted from the submitted buccal swab or blood sample and subjected to polymerase chain reaction (PCR).  The  G20210A and A1991G variants were detected by fluorescence monitoring.  The normal or wild type allele, *1, was assigned by default if none of the variant alleles were detected.

Specimen

Buccal swab or whole blood

Collection

• Buccal swab
• Whole blood,  2-5 mL, into:
i. EDTA-containing tube (purple or lavender top), or
ii. ACD-containing tube (yellow top), or
iii. Citrate-containing tube (blue top).
• Store at 2-8°C.  Ship by overnight carrier at ambient temperature.

Rejection Criteria

• Buccal swab:
i. Physical damage
ii. Specimen appears to have microbial contamination or other visible  contamination
iii. The name on the tube does not match the name on the paperwork.
iv. It is older than 10 days.
• Blood specimen:
i. It is collected in a heparin-containing tube because heparin can  inhibit the PCR reaction.
ii. It leaked in the shipping container.
iii. The name on the tube does not match the name on the paperwork.
iv. It is older than 10 days.

Interpretation

Note that genotypes for both the G20201A and A19911G variants are determined for each patient.  Other genetic and clinical factors should be considered in determining risk, and multiple risk factors are additive.

CPT code

81240
Test Limitations
Other genetic factors, medications and clinical factors (pregnancy, central venous catheters, immobility due to surgery or travel, smoking, transplantation) are also risk factors for thrombosis.  These risk factors tend to be additive.  The detection of Factor II genetic variants does not replace the need for therapeutic drug monitoring or other appropriate clinical monitoring by the health care provider. Additional mutations for Factor II that are not described in the methodology section will not be detected. 
The Factor II genotyping  test was developed and performance characteristics were determined by MDL. The MDL tests have not been cleared or approved by the Food and Drug Administration.  The FDA has determined that such approval is not necessary.

This test is approved for use on New York state residents.
References

1. Blondon M, Hwang M, Smith NL.  Genotyping in Prothrombotic States:  Implications for the Clinician.  Curr Cardiovasc Risk Rep.  5: 525-532.  2011.
2. Chinthammitre Y, Vos HL, Rosendaal FR, Doggen CJM.  The Association of Prothrombin A19911G Polymorphism with Plasma Prothrombin Activity and Venous Thrombosis:  Results of the MEGA Study, a Large Population-Based Case-Control Study.  J Thrombosis and Haemostasis 4: 2587 – 2592.  2006
3. Gohil R, Peck G, Sharma P.  The genetics of venous thromboembolism.  A meta-analysis involving ~120,000 cases and 180,000 controls.  Thromb Haemost 102: 360-370.  2009.
4. Kujovich JL.  Prothrombin-Related Thrombophilia.  In   GeneReviews.  Pagon RA, Bird TD, Dolan CR et al, eds.  pp 1 – 47.  2011.
5. Martinelli I, Battaglioli T.Tosetto A, Legnani C, Sottile L, Ghiotto R, Mannucci PM.  Prothrombin A19911G polymorphism and the risk of venous thromboembolism.  Journal of Thrombosis and Haemostasis 4: 2582-2586.  2006.
6. Perez-Ceballos, Elena, et al.  Prothrombin A11911G and G20210A Polymorphisms’ Role in Thrombosis.  Journal of Thrombosis and Haemostasis, 118: 610-614.  2002
7. Reiner AP, Lange LA, Smith NL, Zakai NA, Cushman M, Folsom AR.  Common hemostasis and inflammation gene variants and venous thrombosis in older adults from the Cardiovascular Health Study.  J Thromb Haemost.  7(9): 1499-1505.  2009
8. Yıldız Z, Deda G, Akar N.  Prothrombin G20210A and A19911G mutations in Turkish pediatric stroke patients.  Turk J Hematol 23:197-199.  (2006) 
9. Zee YL, Glynn RJ, Cheng S, Steiner L, Rose L, Ridker PM.  An Evaluation of Candidate Genes of Inflammation and Thrombosis in Relation to the Risk of Venous Thromboembolism. Circulation Cardiovascular Genetics.  2: 57-62.  2009