1. Thrombophilia

2. Now considered a multicausal disease, with an interplay of acquired and genetic thrombotic risk factors Approximately half of venous thromboembolic episodes in patients with inherited thrombophilias occur in relation to events that are generally recognized as a predisposing states, such surgery, pregnancy, and immobilization

3. Inherited thrombophilic states (1) Antithrombin deficiency Abnormalities in protein C and protein S system - protein C deficiency - protein S deficiency - abnormal thrombomodulin Resistance to activated protein C (FV Leiden, FV Cambridge)

4. Inherited thrombophilic states (2) Hyperprothrombinemia (prothrombin variant G20210A) Dysfibrynogeneimia Abnormalities in fibrinolytic system - hypo- or dysplasminogenemia - elevated plasminogen activator inhibitor - decreased tissue plasminogen activator Hyperhomocysteinemia Heparin cofactor II defciency Elevated histidine-rich glycoprotein Factor XII deficiency

5. Frequency (%) of inherited thrombophilic syndromes in the general population and in patients with venous thrombosis (VT) Syndrome General population Unselected patients with VT Selected patients with VT* AT deficiency PC deficiency PS deficiency APC-resistance 0.02-0.17 0.14-0.5 - 3.6-21 1.1 3.2 2.2 21 0.5-4.9 1.4-8.6 1.4-7.5 10-64 *- age < 45 years and/or recurrent thrombosis

6. Molecular basis of inherited thrombophilia caused by impaired anticoagulant mechanisms (1) Genetic defect No. of different mutations Most frequent mutations AT deficiency PC deficiency >79 >160 Type I: whole or partial gene deletions (<10% of cases) Short insertions or deletions Single nucleotide changes Type II: missense mutations (leading to amino acids substitutions ) Type I: frameshift mutations, nonsense, missense mutations Type II: missense mutations

7. Molecular basis of inherited thrombophilia caused by impaired anticoagulant mechanisms (2) Genetic defect No. of different mutations Most frequent mutations PS deficiency APC-resistance >13 2 Type I: gene deletions, frameshift mutations, nonsense mutation, missense mutations Type II: missense mutations Missence mutation in the factor V molecule

8. Clinical features of patients with inherited deficiencies of AT, PC, PS, and APC- resistance Venous thrombosis (>90% of cases) Deep vein thrombosis of the lower limbs (common) Pulmonary embolism (common) Superficial thrombophlebitis Mesenteric vein thrombosis (rare but characteristic) Cerebral vein thrombosis (rare but characteristic) Frequent family history of thrombosis First thrombosis usually at young age (<40yr*) Frequent recurrences* Neonatal purpura fulminans (homozygous PC or PS deficiency) *- all these features are less evident in patients with APC-resistance, who appear to be less severely affected clinically

9. Laboratory diagnosis of inherited thrombophilia (1) First stepSecond step AT: Heparin cofactor synthetic substrate-based assays PC: Synthetic substrate-based assays (venoms as a PC activators) AT: Immunoassays, crossed immunoelectrophoresis DNA analysis PC: Immunoassays, crossed immunoelectrophoresis DNA analysis

10. Laboratory diagnosis of inherited thrombophilia (2) First stepSecond step PS: Immunoassay of total PS Immunoassay of free PS APC-resistance: APTT-based functional assays (using FV-deficient plasma) PS: crossed immunoelectrophoresis DNA analysis APC-resistance: DNA analysis (mutant factor V)

11. Guidelines for prophylaxis and treatment of thrombosis in patients with inherited thrombophilia (1) IndicationStandard treatmentSpecial cases Primary prophylaxis surgery pregnancy puerperium (up to 4wk after delivery) UFH, sc 5000 IU3xd or OAT, INR 2.0-3.0 Orthopedic or cancer surgery: consider AT or PC concentrates AT deficiency: UFH sc, adjusted doses (APTT 1.3- 1.5), or sequential UFH/OAT AT deficiency: consider AT concentrates at delivery

12. Guidelines for prophylaxis and treatment of thrombosis in patients with inherited thrombophilia (2) IndicationStandard treatmentSpecial cases Secondary prophylaxis first thrombosis recurrent : acute : OAT, INR 2-3 for 6mo lifelong OAT, INR 2-3 UFH iv or sc, APTT ratio 1.5-2.5 followed by OAT, INR 2-3 Life-threatening thrombosis and/or multiple defects: lifelong OAT No postheparin prolongation of APTT or life-threatening events in AT deficiency: add AT concentrates

13. Crossed immunoelectrophoresis of antithrombin in the presence of heparin in 1 st dimension and AT antibody in the 2 nd dimension 1st 2nd

14. Characteristics of AT deficiency Autosomal dominant inheritance Quantitative and qualitative defects Thrombotic phenomena in adolescence or even earlier Frequently pulmonary embolism as first clinical manifestation

15. Characteristics of PC deficiency Autosomal dominant inheritance Quantitative and qualitative defects Homozygotes die because of thrombosis in infancy Thrombotic phenomena in adolescence Skin necrosis when warfarin therapy introduced

16. Characteristics of PS deficiency Autosomal dominant inheritance Quantitative and qualitative defects Homozygotes die because of thrombosis „in utero” or in the early infancy Thrombotic phenomena in adolescence Skin necrosis when warfarin therapy introduced

17. Odds ratios (95% CI) for fetal loss and type of thrombophilia, with control group as reference, adijusted for number of pregnancies and centre (Preston et al., Lancet 1996) Type All spontaneous Miscarriage Stillbirth fetal losses Antitrombin 2.1 (1.2 - 3.6) 1.7 (1.0 - 2.8) 5.2 (1.5 - 18.1) Protein C 1.4 (0.9 - 2.2) 1.4 (0.9 - 2.2) 2.3 (0.6 - 8.3) Protein S 1.3 (0.8 - 2.1) 1.2 (0.7 - 1.9) 3.3 (1.0 - 11.3) Factor V Leiden 1.0 (0.6 - 1.7) 0.9 (0.5 - 1.5) 2.0 (0.5 - 7.7) Combined defects 2.0 (0.5 - 8.1) 0.8 (0.2 - 3.6) 14.3 (2.4 - 86.0)