ABSTRACT
Venous thrombosis is a multicausal disease due to the interaction of genetic and environmental risk factors. Some of the recently discovered genetic risk factors, such as factor V Leiden and prothrombin G20210A mutations, are quite common in the population. Accordingly, laboratory investigation of thrombophilic disorders has expanded to incorporate molecular assays, which unlike functional assays, are unequivocal with no borderline values. When testing for these mutations, specific issues of patient management need to be addressed, such as the duration of anticoagulant therapy, risk stratification for primary or secondary prophylaxis, and family studies. Criteria used to select specific DNA methodologies will center on the issues of cost, automation, speed, reliability, and simplicity. A variety of molecular methods fulfill many but not all of these criteria, whereas the new, semiautomated methodologies of real-time polymerase chain reaction and DNA microarrays offer the potential for widespread application and utility in the future.
KEYWORDS
Thrombophilia - DNA - genetic testing - PCR
REFERENCES
-
1
Rosendaal F R.
Venous thrombosis: a multicausal disease.
Lancet.
1999;
353
1167-1173
-
2
Bertina R.
Molecular risk factors for thrombosis.
Thromb Haemost.
1999;
82
601-609
-
3
De Stefano V, Finazzi G, Manucci P M.
Inherited thrombophilia: pathogenesis, clinical syndromes, and management.
Blood.
1996;
87
3531-3544
-
4
Seligsohn U, Lubetsky A.
Genetic susceptibility to venous thrombosis.
N Engl J Med.
2001;
344
1222-1231
-
5
Lane D A, Bayston T, Olds R J et al..
Antithrombin mutation database; 2nd (1997) update. For the plasma coagulation inhibitors Subcommittee of the Scientific and Standardization Committee of the International Society on Thrombosis and Haemostasis.
Thromb Haemost.
1997;
77
197-211
-
6
Reitsma P H, Bernardi F, Doig R G et al..
Protein C deficiency: a database of mutations, 1995 update. On behalf of the Plasma Coagulation Inhibitors Subcommittee of the Scientific and Standardization Committee of the International Society on Thrombosis and Haemostasis.
Thromb Haemost.
1995;
73
876-889
-
7
Gandrille S, Borgel D, Sala N et al..
Protein S deficiency: a database of mutations-summary of the first update.
Thromb Haemost.
2000;
84
918
-
8
Zivelin A, Grittin J H, Xu X et al..
A single genetic defect for a common Caucasian risk factor for venous thrombosis.
Blood.
1997;
89
397-402
-
9
Zivelin A, Rosenberg N, Faier S et al..
A single genetic origin for the common prothrombotic G20210 polymorphism in the prothrombin gene.
Blood.
1998;
92
1119-1124
-
10
Rees D C, Cox M, Clegg J B.
World distributions of factor V Leiden.
Lancet.
1995;
346
1133-1134
-
11
Rosendaal F R, Doggen C JM, Zivelin A et al..
Geographical distribution of the 20210 G to A prothrombin variant.
Thromb Haemost.
1998;
79
706-708
-
12
Manucci P M, Tripodi A, Botasso B et al..
Markers of procoagulant imbalance in patients with inherited thrombophilic syndromes.
Thromb Haemost.
1992;
67
200-202
-
13
Bertina R M, Koeleman BP C, Koster T et al..
Mutation in blood coagulation factor V associated with resistance to activated protein C.
Nature.
1994;
369
64-67
-
14
Kalafatis M, Bertina R M, Rand M D, Mann K G.
Characterization of the molecular defect in factor VR506Q.
J Biol Chem.
1995;
270
4053-4057
-
15
Heeb M J, Kojima Y, Greengard J S, Griffin J H.
Activated protein C resistance: molecular mechanisms based on studies using purified Gln506-factor V.
Blood.
1995;
85
3405-3411
-
16
Koster T, Rosendaal F R, De Ronde H et al..
Venous thrombosis due to a poor anticoagulant response to activated protein C. Leiden Thrombophilia Study.
Lancet.
1993;
342
1503-1506
-
17
Press R D, Bauer K A, Kukovich J L, Heit J A.
Clinical utility of factor V Leiden (R506Q) testing for the diagnosis and management of thromboembolic disorders.
Arch Pathol Lab Med.
2002;
126
1304-1318
-
18
Middledorp S, Meinardi J R, Koopman M M et al..
A prospective study of asymptomatic carriers of the factor V Leiden mutation to determine the incidence of venous thromboembolism.
Ann Intern Med.
2001;
135
322-327
-
19
Simioni P, Tormene D, Prandoni P et al..
Incidence of venous thromboembolism in asymptomatic family members who are carriers of factor V Leiden: a prospective cohort study.
Blood.
2002;
99
1938-1942
-
20
Poort S R, Rosendaal F R, Reitsma P H, Bertina R M.
A common genetic variation in the 3′ untranslated region of the prothrombin gene is associated with elevated plasma prothrombin levels and an increase in venous thrombosis.
Blood.
1996;
88
3698-3703
-
21
Gehring N H, Frede U, Neu-Yilik G et al..
Increased efficiency of mRNA 3′ end formation: a new genetic mechanism contributing to hereditary thrombophilia.
Nat Genet.
2001;
28
389-392
-
22
Butenas S, van't Veer C, Mann K G.
“Normal” thrombin generation.
Blood.
1999;
94
2169-2178
-
23
Arruda V R, Annichino-Bizzacchi J M, Gonclaves M S, Costa F F.
Prevalence of the prothrombin gene variant (nt20210) in venous thrombosis and arterial disease.
Thromb Haemost.
1997;
78
1430-1433
-
24
McGlennen R C, Key N S.
Clinical and laboratory management of the prothrombin G20210A mutation.
Arch Pathol Lab Med.
2002;
126
1319-1325
-
25
Frosst P, Blom H J, Milos R et al..
A candidate genetic risk factor for vascular disease: a common mutation in methylenetetrahydrofolate reductase.
Nat Genet.
1995;
10
111-113
-
26
Franco R F, Araujo A G, Guerreiro J F, Elion J, Zago M A.
Analysis of the 677 C→T mutation of the methylenetetrahydrofolate reductase gene in different ethnic groups.
Thromb Haemost.
1998;
79
119-121
-
27
Key N S, McGlennen R C.
Hyperhomocysteinemia and thrombophilia.
Arch Pathol Lab Med.
2002;
126
1367-1375
-
28
Kluijtmans L A, den Heijer M, Reitsma P H, Heil S G, Blom H J, Rosendaal F R.
Thermolabile methylenetetrahydrofolate reductase and factor V Leiden in the risk of deep-vein thrombosis.
Thromb Haemost.
1998;
79
254-258
-
29
Williamson D, Brown K, Luddington R, Baglin C, Baglin T.
Factor V Cambridge: a new mutation (Arg306→Thr) associated with resistance to acquired protein C.
Blood.
1998;
91
1140-1144
-
30
Faioni E M, Franchi F, Bucciarelli P et al..
Co-inheritance of the HR2 haplotype in the factor V gene confers an increased risk of venous thromboembolism to carriers of factor V R506Q (factor V Leiden).
Blood.
1999;
94
3062-3066
-
31
Bernardi F, Faioni E M, Castoldi E et al..
A factor V genetic component differing from factor V R506Q contributes to the activated protein C resistance phenotype.
Blood.
1998;
90
1552-1557
-
32
Castaman G, Faioni E M, Tosetto A, Bernardi F.
The factor V HR2 haplotype and the risk of venous thrombosis: a meta-analysis.
Haematologica.
2003;
88
1182-1189
-
33
Faioni E M, Castaman G, Asti D et al..
Association of factor V deficiency with factor V HR2.
Haematologica.
2004;
89
195-200
-
34
Tripodi A, Manucci P M.
Laboratory investigation of thrombophilia.
Clin Chem.
2001;
47
1597-1606
-
35
Pecheniuk N M, Walsh T P, Marsh N A.
DNA technology for the detection of common genetic variants that predispose to thrombophilia.
Blood Coagul Fibrinolysis.
2000;
11
683-700
-
36
Gomez E, van der Poel S CPAM, Jansen J H, van der Reijden L M, Lowenberg R.
Rapid simultaneous screening of factor V Leiden and G20210A prothrombin variant by multiplex polymerase chain reaction on whole blood.
Blood.
1998;
91
2208-2211
-
37
Muriel G, Christine B, Philippe J, Christophe M, Francois S J, Patricia A M.
An even easier method for one-step detection of both FV Leiden and FII G20210A transition.
Blood.
1998;
92
3478-3479
-
38
Hezard N, Cornillet P, Droulle C, Gillot L, Potron G, Nguyen P.
Factor V Leiden: detection in whole blood by ASA PCR using an additional mismatch in antepenultimate position.
Thromb Res.
1997;
88
59-66
-
39
Poort S R, Bertina R M, Vos H L.
Rapid detection of the prothrombin 20210A variation by allele-specific PCR.
Thromb Haemost.
1997;
78
1157-1158
-
40
Mitterer M, Lanthaler A J, Mair W, Giacomuzzi K, Coser P.
Simultaneous detection of FV Q506and prothrombin 20210 A variation by allele-specific PCR.
Haematologica.
1999;
84
204-207
-
41
Van D Locht L TF, Kuypers A WHM, Verbruggen B W, Linssen P CM, Novakova I RO, Mensink E JBM.
Semi-automated detection of factor V mutation by allele-specific amplification and capillary electrophoresis.
Thromb Haemost.
1995;
74
1276-1279
-
42
Margaglione M, D'Andrea G, Capucci G et al..
Detection of the factor V Leiden using SSCP.
Thromb Haemost.
1996;
76
814-815
-
43
Hezard N, Cornillet P, Vallade P, Nguyen P.
Detection of factor V Leiden using ASO (allele specific oligonucleotide).
Thromb Haemost.
1997;
78
1296
-
44
Zotz R B, Marhun-Debowski B, Sharf R E.
Mutation in the gene coding for coagulation factor V and resistance to activated protein C: detection of the genetic mutation by oligonucleotide ligation assay using a semi-automated system.
Thromb Haemost.
1996;
76
53-55
-
45
Pecheniuk N M, Marsh N A, Walsh T P.
Multiple analysis of three common genetic alterations associated with thrombophilia.
Blood Coagul Fibrinolysis.
2000;
11
183-189
-
46
Ameziane N, Lamotte M, Lamoril J et al..
Combined factor V Leiden (G1691A) and prothrombin (G20210A) genotyping by multiplex real-time polymerase chain reaction using fluorescent resonance energy transfer hybridization probes on the Rotor-Gene 2000. Blood Coagul Fibrinolysis.
2003;
14
421-424
-
47
Cooper P C, Cooper S M, Smith J M, Kitchen S, Makris M.
Evaluation of the Roche LightCycler: a simple and rapid method for direct detection of factor V Leiden and prothrombin G20210A genotypes from blood samples without the need for DNA extraction.
Blood Coagul Fibrinolysis.
2003;
14
499-503
-
48
Schrijver I, Lay M J, Zehnder J L.
Diagnostic single nucleotide polymorphism analysis of factor V Leiden and prothrombin 20210G>A. A comparison of the Nanogen Electronic Microarray with restriction enzyme digestion and the Roche LightCycler.
Am J Clin Pathol.
2003;
119
490-496
-
49
Green D.
Genetic hypercoagulability: screening should be an informed choice.
Blood.
2001;
98
20
-
50
Machin S J.
Pros and cons of thrombophilia testing: cons.
J Thromb Haemost.
2003;
1
412-413
-
51
Manucci P M.
Genetic hypercoagulability: prevention suggests testing family members.
Blood.
2001;
98
21-22
-
52
Baglin T, Luddington R, Brown K, Baglin C.
Incidence of recurrent venous thromboembolism in relation to clinical and thrombophilic risk factors: prospective cohort study.
Lancet.
2003;
362
523-526
Mark Hertzberg M.D. Ph.D.
Department of Haematology, Westmead Hospital, Westmead
New South Wales 2145, Australia
eMail: markh@westgate.wh.usyd.edu.au
