Hypercoagulable States - Thrombophilia

Key Points

The widespread use of numerous anticoagulants for a variety of diseases, including the newest direct oral anticoagulant agents, has created clinical dilemmas for both anticoagulant monitoring and the interpretation of coagulation testing. Evaluations for thrombophilia may demonstrate interference once heparin, warfarin, or one of the new direct agents is administered, making interpretation of results difficult. For more information, refer to Impacts of Common Anticoagulants on Coagulation Testing.

Diagnosis

Indications for Testing

• New or recurrent venous thromboembolism (VTE) without obvious risk factors (see Differential Diagnosis)
  • Abnormalities may be identified in a significant number of patients; however, identification of an abnormality may not predict risk of recurrence or alter therapeutic plan
  • Use when the results will impact management of the patient or patient family members
  • Do not order testing for first deep vein thrombosis (DVT) in the setting of known etiology (Five Things Physicians and Patients Should Question, 2015; Society for Vascular Medicine)
  • Do not test during acute episode – results do not alter therapy and some results may be inaccurate
• Any of the following situations (College of American Pathologists [CAP], 2002; Grody, American College of Medical Genetics and Genomics [ACMG], 2001)
  • Idiopathic thrombosis in patient ≤50 years
  • Recurrent thrombosis
  • Unusual sites of thrombosis in the absence of risk factors
  • First-degree relatives with thromboses
  • Thrombotic event during pregnancy or while taking oral contraceptives or hormone replacement therapy

Laboratory Testing

• Based on family and patient history – nongenetic tests may be altered by anticoagulant therapy or acute phase reaction due to clot
• Consider acquired disorders such as antiphospholipid syndrome
• Children – very low risk for thrombophilia and should only have testing performed in consultation with hematologist (ACMG)
• If testing pursued
  • Do not test for protein C, protein S, or antithrombin (AT) levels during an active clotting event to diagnose a hereditary deficiency because these tests are not analytically accurate during an active clotting event (Choosing Wisely: 20 Things Physicians and Patients Should Question, 2017; American Society for Clinical Pathology)
  • Consider the following combination (ACMG)
    • Activated protein C resistance (with or without reflex to factor V Leiden [FVL] mutation); factor V R2 A4070G mutation
    • Prothrombin mutation
    • AT activity
    • Protein C activity
    • Free protein S
  • Less common disorders – testing may be considered if results are uninformative and additional testing is considered necessary
• The following tests are not recommended
  • Factor VIII activity – testing other factor activities such as factor VIII and factor IX is controversial and not currently recommended
  • Factor XIII (F13A1) V34L variant – presence of variant associated with decreased/reduced risk for DVT, myocardial infarction, and coronary artery disease in Whites
• Before making a definitive diagnosis of an inherited thrombophilia, consider repeating abnormal functional or antigenic testing
  • False positives and false negatives may encourage inappropriate therapies or follow-up planning
  • Low results can be obtained due to patient condition/biologic variability, medications, and assay variability or interference
  • Normal ranges vary by age and gender and must be considered when interpreting results

Differential Diagnosis

• Acquired thrombophilia is more common than inherited thrombophilia and should be considered when evaluating patients with thrombosis
  • Antiphospholipid antibodies/lupus anticoagulant
  • Malignancy
  • Long-distance travel
  • Trauma
  • Surgery
  • Immobilization
  • Presence of a central venous catheter
  • Pregnancy/postpartum
  • Therapy-related thrombophilia
    • Hormone replacement therapy
    • Oral contraceptives
    • Tamoxifen/raloxifene
    • Chemotherapy
    • Heparin-induced thrombocytopenia

Screening

Population testing is not recommended for unselected patients.

Background

Epidemiology

• Adults
  • Incidence – 30-500/10,000 for all disorders
  • Ethnicity
    • Slightly higher among African Americans
    • Lower in Asian and Native Americans

    • Inherited Thrombophilic Disorders

      Disorder	Prevalence in Normals (%)	Frequency in Patients with VTE (%)	Relative Risk of First Episode of DVT
      Factor V Leiden (heterozygous)	0.05-4.8a	18.8	7
      Factor V Leiden (homozygous)	0.02	1.5	80
      Factor V with R2 mutation	0.06-0.12	10.0	7 (heterozygote) 10 (compound heterozygote)
      Prothrombin G20210A allele	0.06-2.7a	7.1	2.8
      Protein C deficiency	0.2-0.4	3.7	6.5
      Protein S deficiency	0.16-0.21	2.3	5.0
      Antithrombin deficiency	0.02	1.9	20
      Dysfibrinogenemia	<0.01	0.8	Unknown
      Hyperhomocysteinemiab	5-7	10	2.95
      Elevated factor VIII level	11	25	4.8
      Elevated factor IX level	10	20	2.8
      Elevated factor XI level	10	19	2.2
      Elevated lipoprotein (a) level	7	20	3.2
      Elevated TAFI	9	14	1.7
      aPercent lowest in Asian or African descent; highest in White descent b>18.5 µmol/L DVT, deep vein thrombosis; TAFI, thrombin-activatable fibrinolysis inhibitor; VTE, venous thromboembolism Source: Whitlatch, 2008

• Pediatrics
  • Incidence – 0.05-14/100,000
  • Sex – M<F
  • Age
    • Peak in neonates and infants <1 year
    • Second peak in puberty

Etiologies

Most Common Thrombophilias

• Factor V Leiden (FVL)
  • Genetics and pathophysiology
    • FVL mutation of the F5 gene is the most common inherited thrombophilia
      • Accounts for more than 90% of patients with activated protein C resistance (APC-R)
        • During normal hemostasis, APC limits clot formation by proteolytic inactivation of factors Va and VIIIa
        • FVL prevents inactivation of factor Va by APC at the normal rate, increasing the risk for thrombosis
      • Functional tests for APC-R are generally used to screen for FVL
        • DNA tests are used to confirm positive screening tests and to differentiate between heterozygotes and homozygotes
    • Incomplete autosomal dominant inheritance
      • Heterozygotes have a 5- to 10-fold increased risk
      • Homozygotes have a 50- to 100-fold increased risk
  • Clinical presentation
    • Venous thromboembolism (VTE) is the most common type of thrombotic event
      • Recurrent VTE (pulmonary embolism, deep vein thrombosis [DVT]) is uncommon in heterozygotes unless additional risk factors are present
      • Increased risk of recurrent VTE in homozygotes
    • Pregnancy complications – recurrent miscarriage in the second trimester
  • Additional risk factors
    • Presence of F5 R2 (A4070G) mutation with FVL increases risk of thrombotic event 10-fold
    • ≥1 genetic risk factor – concomitant prothrombin G20210A mutation of F2, protein C deficiency, homocysteinemia
    • Acquired thrombophilic disorders (eg, malignancy, hyperhomocysteinemia, high factor VIII levels)
    • Nongenetic factors such as pregnancy, oral contraceptives, hormone replacement therapy, selective estrogen-receptor modulators, travel, immobilization, central venous catheters, surgery, transplantation, and advanced age
• ​Prothrombin G20210A
  • ​Genetics and pathophysiology
    • Prothrombin G20210A mutation of the F2 gene – second most common inherited thrombophilia
      • Results in elevated levels of plasma prothrombin leading to hypercoagulability (gain of function)
      • Detected using DNA tests – factor II (prothrombin) activity testing may not identify the disease and should not be used for diagnosis
    • Autosomal dominant inheritance
      • Variable penetrance – many patients who are either heterozygous or homozygous for G20210A do not experience VTE
      • Heterozygosity causes a 2- to 4-fold increase in thrombotic risk
      • Homozygosity is rare and increases thrombotic risk above that observed in G20210A heterozygotes
  • Clinical presentation
    • VTE
    • Pregnancy complications – preeclampsia, placental abruption
  • Additional risk factors
    • Combined heterozygosity for both prothrombin G20210A and FVL mutations leads to thrombophilia with earlier onset, higher rate of recurrence, and more severe thrombotic events than either mutation alone
    • Increased risk of thrombosis associated with oral contraceptive use and pregnancy

Less Common Thrombophilias

• Increased clotting factors – elevated factor VIII (FVIII) levels are often found in patients with venous thrombosis, but routine testing is controversial
• Protein C deficiency
  • Genetics and pathophysiology
    • Protein C is a vitamin K-dependent plasma anticoagulant activated to APC by thrombin-thrombomodulin, which then inactivates factors Va and VIIIa
    • Inherited protein C deficiency is uncommon – 2 forms
      • Type I – quantitative
      • Type II – qualitative
    • Autosomal dominant inheritance – highly variable phenotypic expression
    • Functional assays preferred over antigenic assays for diagnosis – protein C levels vary with age
    • Decreased levels in acute thrombotic states, disseminated intravascular coagulation (DIC), liver disease, malnutrition (vitamin K deficiency), and with warfarin therapy
    • Increased levels in diabetes, nephrotic syndrome, pregnancy, and with oral contraceptive use
      • Elevated FVIII levels may result in falsely decreased values for some functional assays
      • Heparin and direct thrombin inhibitors may result in falsely elevated values for some functional assays
  • Clinical presentation
    • Additional risk factors likely necessary to provoke thrombosis (eg, infection – varicella-zoster virus [VZV], meningococcal)
    • VTE in heterozygotes
    • Neonatal purpura fulminans (DIC) in homozygous infants – widespread thromboses with hemorrhagic skin necroses
    • Warfa​rin-induced skin necrosis is rarely seen
• Protein S deficiency
  • Genetics and pathophysiology
    • Protein S is a vitamin K-dependent plasma anticoagulant that acts as a cofactor for activated protein C and exists in 2 forms
      • Free protein S
        • 40% of the total
        • Physiologically active
      • Bound protein S (attached to C4b-binding protein)
        • 60% of the total
        • No anticoagulant activity
    • Inherited protein S deficiency is uncommon
      • 3 forms
        • Type I – quantitative
        • Type II – qualitative
        • Type III (also called type IIa) – quantitative with normal levels of total protein S
      • Autosomal dominant inheritance
      • Antigenic tests for free protein S preferred for diagnosis – free protein S values are higher in males than in females
    • Decreased levels in acute thrombotic states, nephrotic syndrome, inflammatory syndromes (due to increased C4b-binding protein), DIC, liver disease, malnutrition (vitamin K deficiency), pregnancy, and with estrogen and warfarin therapy
      • Elevated FVIII levels and/or APC resistance may result in falsely decreased values in some functional assays
      • Increased levels in heparin and direct thrombin inhibitors for some functional assays
  • Clinical presentation
    • Additional risk factors likely necessary to provoke thrombosis
    • VTE most common; arterial thrombosis may occur
    • Neonatal purpura fulminans (DIC) in homozygous infants
    • Warfarin-induced skin necrosis (rare)
• Hyperhomocysteinemia (acquired or inherited)
  • Acquired
    • Independent risk factor for thromboembolic events
    • Most patients with hyperhomocysteinemia have no genetic variants or polymorphisms
    • Result of defective homocysteine metabolism and may be the result of vitamin B6, B12, or folic acid deficiency
    • Thrombotic risk most closely associated with increased fasting plasma homocysteine levels regardless of underlying etiology – plasma homocysteine testing is recommended over DNA-based tests
  • Inherited
    • Homocysteine metabolism defects caused by MTHFR gene variants
      • Most common variants – c.665C>T (p.Ala222Val) (previously designated C677T) and c.1286A>C (p.Glu429la) (previously designated A1298C)
      • Homozygosity for the c.665C>T variant is a risk factor for hyperhomocysteinemia
    • Autosomal recessive inheritance
    • Elevated plasma homocysteine level may be associated with a mild increase in risk for VTE independent of MTHFR status, but meta-analyses have cast doubt on the association between moderate homocysteine elevation and increased risk for coronary heart disease and VTE (Hickey, 2013)
    • The American College of Medical Genetics recommends MTHFR genotyping not be ordered as part of routine workup for thrombophilia due to questionable clinical significance (Hickey, 2013)
    • Inherited hyperhomocysteinemia rare
      • 

• Antithrombin deficiency
  • Genetics and pathophysiology
    • Antithrombin (AT) – plasma anticoagulant; inactivates thrombin, factor Xa, and other activated clotting factors
      • AT activity enhanced by heparin-like glycosaminoglycans on the endothelial surface and by pharmaceutical heparin
      • Synthesized in the liver
    • 2 forms of inherited antithrombin deficiency
      • Type I – quantitative
      • Type II – qualitative
    • Autosomal dominant inheritance
      • Functional assays preferred for diagnosis
      • Homozygous state is embryonic lethal
    • Decreased levels occur in acute thrombotic states, liver disease, DIC, nephrotic syndrome, and heparin therapy; mild decreases may be seen in pregnancy or with oral contraceptive use
    • Increased levels may occur with long-term warfarin therapy
  • Clinical presentation
    • VTE
    • Recurrent thrombosis may occur even in the absence of additional risk factors
    • Some deficient patients are resistant to heparin therapy
• Impaired clot lysis (dysfibrinogenemia, abnormal fibrinolysis)​

Acquired Thrombophilic State

Antiphospholipid syndrome