PDF herunterladen
Thromb Haemost 2012; 108(02): 247-257
DOI: 10.1160/TH11-12-0875
Blood Coagulation, Fibrinolysis and Cellular Haemostasis
Schattauer GmbH

Deficiencies of antithrombin, protein C and protein S – Practical experience in genetic analysis of a large patient cohort

Michael Caspers
1   Institute of Experimental Haematology and Transfusion Medicine, University Clinic Bonn, Bonn, Germany
,
Anna Pavlova
1   Institute of Experimental Haematology and Transfusion Medicine, University Clinic Bonn, Bonn, Germany
,
Julia Driesen
1   Institute of Experimental Haematology and Transfusion Medicine, University Clinic Bonn, Bonn, Germany
,
Ursula Harbrecht
1   Institute of Experimental Haematology and Transfusion Medicine, University Clinic Bonn, Bonn, Germany
,
Robert Klamroth
2   Vivantes – Klinikum in Friedrichhain, Department of Angiology and Clotting Disorders, Haemophilia Treatment Center, Berlin, Germany
,
Janos Kadar
3   Laboratory and Medical practice for Transfusion Medicine, Köln, Germany
,
Ronald Fischer
4   Department of Internal Medicine, Justus Liebig University Giessen, Giessen, Germany
,
Bettina Kemkes- Matthes
4   Department of Internal Medicine, Justus Liebig University Giessen, Giessen, Germany
,
Johannes Oldenburg
1   Institute of Experimental Haematology and Transfusion Medicine, University Clinic Bonn, Bonn, Germany
› Institutsangaben
Weitere Informationen

Publikationsverlauf

Received: 20. Dezember 2011

Accepted after major revision: 22. April 2012

Publikationsdatum:
25. November 2017 (online)

Auch verfügbar auf
    Lizenzen und Reprints

Summary

Deficiencies of natural anticoagulant proteins including antithrombin (AT), protein C (PC) and protein S (PS) are important causes of inherited thrombophilia. This study aimed to report on the practical experience gained in performing genetic analyses of a large cohort of patients with AT, PC and PS deficiencies and to relate this knowledge to clinical application. We genotyped a large cohort of 709 unrelated patients with AT (231), PC (234) and PS (244) deficiencies referred to us by physicians throughout Germany. Mutations were detected by direct sequencing and multiplex ligation-dependent probe amplification (MLPA). The highest mutation detection rate (MDR) was found for the SERPINC1 gene (83.5%), followed by the PROC (69%) and PROS1 (43%) genes. Even at AT activities close to the normal range (75%), the MDR was 70%. Contrastingly, for PC and PS deficiencies, the MDR dropped significantly and mildly lowered to subnormal values. At PS activities >55% for PS no mutations were detected. Mutation profiles of all three genes were similar with the highest prevalence for missense mutations (63–78%), followed by nonsense (7–11%), splice-site mutations (7–13%), small deletions (1–8%), small insertions/duplications (1–4%) and large deletions (3–6%). In conclusion, genetic testing is a useful diagnostic tool for diagnosing thrombophilia. Based on our data, genetic analysis for patients with AT deficiency is indicated for all subnormal activities. In contrast, genotyping is not advisable for PC activities >70% and for PS activities >55%.

Keywords

Protein C - protein S - antithrombin - thrombophilia - genetic analysis
 

  • References

  • 1 Bafunno V, Margaglione M. Genetic basis of thrombosis. Clin Chem Lab Med 2010; 48 (Suppl. 01) S41-51.
    PubMedGoogle Scholar
  • 2 Naess IA, Christiansen SC, Romundstad P. et al. Incidence and mortality of venous thrombosis: a population-based study. J Thromb Haemost 2007; 05: 692-699.
    CrossrefPubMedGoogle Scholar
  • 3 Kyrle PA, Minar E, Bialonczyk C. et al. The risk of recurrent venous thromboembolism in men and women. N Engl J Med 2004; 350: 2558-2563.
    CrossrefPubMedGoogle Scholar
  • 4 Lane DA, Mannucci PM, Bauer KA. et al. Inherited thrombophilia: Part 1. Thromb Haemost 1996; 76: 651-662.
    Artikel in Thieme ConnectPubMedGoogle Scholar
  • 5 Olds RJ, Lane DA, Chowdhury V. et al. Complete nucleotide sequence of the antithrombin gene: evidence for homologous recombination causing thrombophilia. Biochemistry 1993; 32: 4216-4224.
    CrossrefPubMedGoogle Scholar
  • 6 Hirsh J, Piovella F, Pini M. Congenital antithrombin III deficiency. Incidence and clinical features. Am J Med 1989; 87 Suppl 3B 34S-8S.
    PubMedGoogle Scholar
  • 7 Foster DC, Yoshitake S, Davie EW. The nucleotide sequence of the gene for human protein C. Proc Natl Acad Sci USA 1985; 82: 4673-4677.
    CrossrefPubMedGoogle Scholar
  • 8 Whitlatch NL, Ortel TL. Thrombophilias: when should we test and how does it help?. Semin Respir Crit Care Med 2008; 29: 25-39.
    Artikel in Thieme ConnectPubMedGoogle Scholar
  • 9 Ploos van Amstel JK, van der Zanden AL, Bakker E. et al. Two genes homologous with human protein S cDNA are located on chromosome 3. Thromb Haemost 1987; 58: 982-987.
    Artikel in Thieme ConnectPubMedGoogle Scholar
  • 10 Beauchamp NJ, Dykes AC, Parikh N. et al. The prevalence of, and molecular defects underlying, inherited protein S deficiency in the general population. Br J Haematol 2004; 125: 647-654.
    CrossrefPubMedGoogle Scholar
  • 11 de Bruijn SF, Stam J, Koopman MM. et al. Case-control study of risk of cerebral sinus thrombosis in oral contraceptive users and in [correction of who are] carriers of hereditary prothrombotic conditions. The Cerebral Venous Sinus Thrombosis Study Group. Br Med J 1998; 316: 589-592.
    CrossrefPubMedGoogle Scholar
  • 12 Miller SA, Dykes DD, Polesky HF. A simple salting out procedure for extracting DNA from human nucleated cells. Nucleic Acids Res 1988; 16: 1215.
    CrossrefPubMedGoogle Scholar
  • 13 Goodeve AC, Reitsma PH, McVey JH. Nomenclature of genetic variants in hemostasis. J Thromb Haemost 2011; 09: 852-855.
    CrossrefPubMedGoogle Scholar
  • 14 Perry DJ, Daly M, Harper PL. et al. Antithrombin Cambridge II, 384 Ala to Ser. Further evidence of the role of the reactive centre loop in the inhibitory function of the serpins. FEBS Lett 1991; 285: 248-250.
    CrossrefPubMedGoogle Scholar
  • 15 Olds RJ, Lane DA, Boisclair M. et al. Antithrombin Budapest 3. An antithrombin variant with reduced heparin affinity resulting from the substitution L99F. FEBS Lett 1992; 300: 241-246.
    CrossrefPubMedGoogle Scholar
  • 16 Thein SL, Lane DA. Use of synthetic oligonucleotides in the characterization of antithrombin III Northwick Park (393 CGT----TGT) and antithrombin III Glasgow (393 CGT----CAT). Blood 1988; 72: 1817-1821.
    PubMedGoogle Scholar
  • 17 Duchemin J, Gandrille S, Borgel D. et al. The Ser 460 to Pro substitution of the protein S alpha (PROS1) gene is a frequent mutation associated with free protein S (type IIa) deficiency. Blood 1995; 86: 3436-3443.
    PubMedGoogle Scholar
  • 18 Cunningham MT, Olson JD, Chandler WL. et al. External quality assurance of antithrombin, protein C, and protein s assays: results of the College of American Pathologists proficiency testing program in thrombophilia. Arch Pathol Lab Med 2011; 135: 227-232.
    PubMedGoogle Scholar
  • 19 Bereczky Z, Kovacs KB, Muszbek L. Protein C and protein S deficiencies: similarities and differences between two brothers playing in the same game. Clin Chem Lab Med 2010; 48 (Suppl. 01) S53-66.
    PubMedGoogle Scholar
  • 20 Kottke-Marchant K, Comp P. Laboratory issues in diagnosing abnormalities of protein C, thrombomodulin, and endothelial cell protein C receptor. Arch Pathol Lab Med 2002; 126: 1337-1348.
    PubMedGoogle Scholar
  • 21 Koeleman BP, Reitsma PH, Bertina RM. Familial thrombophilia: a complex genetic disorder. Semin Hematol 1997; 34: 256-264.
    PubMedGoogle Scholar
  • 22 Spek CA, Koster T, Rosendaal FR. et al. Genotypic variation in the promoter region of the protein C gene is associated with plasma protein C levels and thrombotic risk. Arterioscler Thromb Vasc Biol 1995; 15: 214-218.
    CrossrefPubMedGoogle Scholar
  • 23 Johansson AM, Hillarp A, Sall T. et al. Large deletions of the PROS1 gene in a large fraction of mutation-negative patients with protein S deficiency. Thromb Haemost 2005; 94: 951-957.
    Artikel in Thieme ConnectPubMedGoogle Scholar
  • 24 Lanke E, Johansson AM, Hillarp A. et al. Co-segregation of the PROS1 locus and protein S deficiency in families having no detectable mutations in PROS1. J Thromb Haemost 2004; 02: 1918-1923.
    CrossrefPubMedGoogle Scholar
  • 25 Persson KE, Dahlback B, Hillarp A. Diagnosing protein S deficiency: analytical considerations. Clin Lab 2003; 49: 103-110.
    PubMedGoogle Scholar
  • 26 ten Kate MK, van der Meer J. Protein S deficiency: a clinical perspective. Haemophilia 2008; 14: 1222-1228.
    PubMedGoogle Scholar
  • 27 Almasy L, Soria JM, Souto JC. et al. A quantitative trait locus influencing free plasma protein S levels on human chromosome 1q: results from the Genetic Analysis of Idiopathic Thrombophilia (GAIT) project. Arterioscler Thromb Vasc Biol 2003; 23: 508-511.
    CrossrefPubMedGoogle Scholar
  • 28 Mulder R, Ten Kate MK, Kluin-Nelemans HC. et al. Low cut-off values increase diagnostic performance of protein S assays. Thromb Haemost 2010; 104: 618-625.
    Artikel in Thieme ConnectPubMedGoogle Scholar
  • 29 Lijfering WM, Mulder R, ten Kate MK. et al. Clinical relevance of decreased free protein S levels: results from a retrospective family cohort study involving 1143 relatives. Blood 2009; 113: 1225-1230.
    CrossrefPubMedGoogle Scholar
  • 30 Luxembourg B, Delev D, Geisen C. et al. Molecular basis of antithrombin deficiency. Thromb Haemost 2011; 105: 635-646.
    Artikel in Thieme ConnectPubMedGoogle Scholar
  • 31 Pintao MC, Garcia AA, Borgel D. et al. Gross deletions/duplications in PROS1 are relatively common in point mutation negative hereditary protein S deficiency. Hum Genet 2009; 126: 449-456.
    CrossrefPubMedGoogle Scholar
  • 32 Pavlova A, El-Maarri O, Luxembourg B. et al. Detection of heterozygous large deletions in the antithrombin gene using multiplex polymerase chain reaction and denatured high performance liquid chromatography. Haematologica 2006; 91: 1264-1267.
    PubMedGoogle Scholar
  • 33 Yin T, Takeshita S, Sato Y. et al. A large deletion of the PROS1 gene in a deep vein thrombosis patient with protein S deficiency. Thromb Haemost 2007; 98: 783-789.
    Artikel in Thieme ConnectPubMedGoogle Scholar
  • 34 Shaw CJ, Lupski JR. Implications of human genome architecture for rearrangement-based disorders: the genomic basis of disease. Hum Mol Genet 2004; 13 (Suppl. 01) R57-64.
    CrossrefPubMedGoogle Scholar
  • 35 Aiach M, Nicaud V, Alhenc-Gelas M. et al. Complex association of protein C gene promoter polymorphism with circulating protein C levels and thrombotic risk. Arterioscler Thromb Vasc Biol 1999; 19: 1573-1576.
    CrossrefPubMedGoogle Scholar
  • 36 Pomp ER, Doggen CJ, Vos HL. et al. Polymorphisms in the protein C gene as risk factor for venous thrombosis. Thromb Haemost 2009; 101: 62-67.
    Artikel in Thieme ConnectPubMedGoogle Scholar
  • 37 Kimura R, Kokubo Y, Miyashita K. et al. Polymorphisms in vitamin K-dependent gamma-carboxylation-related genes influence interindividual variability in plasma protein C and protein S activities in the general population. Int J Hematol 2006; 84: 387-397.
    CrossrefPubMedGoogle Scholar
  • 38 Leroy-Matheron C, Duchemin J, Levent M. et al. Genetic modulation of plasma protein S levels by two frequent dimorphisms in the PROS1 gene. Thromb Haemost 1999; 82: 1088-1092.
    Artikel in Thieme ConnectPubMedGoogle Scholar
  • 39 Castaman G, Biguzzi E, Razzari C. et al. Association of protein S p.Pro667Pro dimorphism with plasma protein S levels in normal individuals and patients with inherited protein S deficiency. Thromb Res 2007; 120: 421-426.
    CrossrefPubMedGoogle Scholar
  • 40 Buil A, Soria JM, Souto JC. et al. Protein C levels are regulated by a quantitative trait locus on chromosome 16: results from the Genetic Analysis of Idiopathic Thrombophilia (GAIT) Project. Arterioscler Thromb Vasc Biol 2004; 24: 1321-1325.
    CrossrefPubMedGoogle Scholar