Thrombosis, thrombophilia and antithrombotic therapy

1. Thrombosis, thrombophilia and antithrombotic therapy By Carl Peters

2. Definitions • Thrombi – solid masses or plugs formed in the circulation from blood constituents • Both arterial and venous thrombosis can occur. • Thrombophilia – term used to describe the inherited or acquired disorders of the haemostatic mechanism which predispose to thrombosis. • Hypofibrinolysis – can’t lyse clots normally.

3. Pathogenesis • Virchow’s triad suggests three components important in thrombus formation: • Slowing down of blood flow (venous) • Hypercoagulability of the blood (venous) • Vessel wall damage (arterial)

4. Remember… • Risk stratification represents a spectrum • Overall aim to decide: • What interventions can be implemented? • Specifically, what anticoagulants to use? • How long to use them? • And, when to use them (e.g. surgery, pregnancy)?

5. Arterial thrombosis

6. Common locations • Coronary arteriesÞ myocardial infarction • Carotid / vertebral / intracerebral arteries Þ ischaemic stroke • Peripheral arteries (usually leg) Þ ischemia and (eventually) gangrene • Mesenteric arteries Þ bowel infarction • Retinal arteries Þ loss of vision • Placental arteries Þ placental insufficiency, miscarriage, and major complications of pregnancy including eclampsia and pre-eclampsia, abruptio placentae, and intrauterine growth retardation.

7. Pathogenesis • Atherosclerosis of arterial wall, plaque rupture and endothelial injury expose blood to subendothelial collagen and tissue factor.

8. Haemostasis pathway Injury Collagen exposure Tissue Factor Platelet adhesion Coagulation Cascade Release reaction Platelet aggregation Fibrin Primary haemostatic plug Secondary haemostatic plug

9. Coagulation cascade

10. Pathogenesis cont… • Platelet deposition and thrombus formation are important in the pathogenesis of atherosclerosis • Platelet-derived growth factor (PDGF) stimulates the migration and proliferation of smooth cells and fibroblasts in the arterial intima. Regrowth of endothelium and repair at the site of arterial damage and incorporated thrombus result in thickening of vessel wall.

11. Risk factors (arterial) • Related to the development of atherosclerosis • Northwick Park heart study (Lancet. 1986 Sep 6;2(8506):533-7) showed that elevated plasma levels of factor VII and fibrinogen are the strongest independent predictors of coronary events.

12. Risk factors (arterial) • Positive family history • Male sex • Hyperlipidaemia • Hyperhomocysteinaemia • Low serum folate, vitamin B12, vitamin B6 • Hypertension • Diabetes mellitus • Gout • Polycythaemia • Cigarette smoking • ECG abnormalities • Elevated factor VII and fibrinogen (elevations of one std. deviation associated with increases of risk of 62% and 84% respectively of IHD) • Lupus anticoagulant • Collagen vascular diseases • Bechet’s disease (chronic, multisystem inflammatory disease) • F VIII

13. Hyperhomocysteinaemia • Mild hyperhomocysteinemia in approx. 5 to 7 percent of the general population • Severe hyperhomocysteinemia is rare • Patients with mild hyperhomocysteinemia are asymptomatic until the third or fourth decade of life when premature coronary artery disease may develop, as well as recurrent arterial and venous thrombosis

14. Dietary protein Remethylation Trans-sulphuration Folic acid

15. Hyperhomocysteinaemia • Homocyteine – derived from dietary methionine. • Removed by either remethylation or trans-sulphuration • Classic homocystinuria • rare autosomal dominant disorder • caused by deficiency of cystathione beta-synthetase, enzyme responsible for trans-sulphuration • Vascular disease and thrombosis major features of disease • Heterozygous CBS deficiency • present in around 0.5% of popn. • leads to moderate increase in homocysteine. • Methylene tetrahydrofolate reductase (MTHFR) • involved in the remethylation pathway • a common thermolabile variant may be responsible for mild homocysteinaemia, although may only be seen in presence of folate deficiency. • Acquired risk factors for homocysteinaemia include deficiencies of folate, B12 or B6, drugs (e.g. cyclosporine), renal damage and smoking. Levels also increase with age and higher in men and post-menopausal women. • NB also a risk factor for venous thrombosis

16. Hyperhomocysteinaemia • Diagnosis: plasma homocysteine measured on a morning specimen after an overnight fast.  Because homocysteine is continuously released by blood cells, the specimen must be centrifuged and the plasma separated immediately to avoid falsely elevated values • Elevations in plasma homocysteine are typically caused either by genetic defects in the enzymes involved in homocysteine metabolism or by nutritional deficiencies in vitamin cofactors • Homocysteine >13.1 umol/L- Folic acid 5 mg, Vitamin B6 100 mg, Vitamin B12, 2000 ug/day

17. Venous thrombosis

18. Common locations • most often occurs in the leg, either in the superficial veins (superficial phlebitis), or in the deep veins (deep venous thrombosis) • cerebral sinus • retinal veins/and or arteries • veins in the arms or upper thorax • veins in the mesentery • veins in bones, usually the hip or jaw, causing bone death (osteonecrosis) • veins and/or arteries in the placenta, causing placental insufficiency, miscarriage, and major complications of pregnancy including eclampsia and pre-eclampsia, abruptio placentae, and intrauterine growth retardation.

19. Pathogenesis • Increased systemic coagulability and stasis are most important • Vessel wall damage being less important than in arterial thrombosis, although may be important in patients with sepsis and in-dwelling catheters • Stasis allows the completion of blood coagulation at the site of initiation of the thrombus (e.g. behind the valve pockets of the leg veins in immobile patients)

20. Risk factors Related to coagulation abnormality: • Hereditary hemostatic disorders • Factor V Leiden • Prothrombin G20210A variant • Protein C deficiency • Protein S deficiency • Antithrombin III deficiency • Abnormal fibrinogen • Abnormal plasminogen

21. Risk factors cont… • Acquired haemostatic disorders (some of these may also occur as inherited disorders) • Raised plasma levels of factor VII, VIII, IX or XI • Raised plasma levels of fibrinogen • Raised plasma levels of homocysteine, possibly B12 defriciency • Glucosylceramide deficiency • Coagulation factor IX concentrates • Lupus anticoagulant • Oestrogen therapy (oral contraceptive and HRT) • Heparin-induced thrombocytopenia • Pregnancy and puerperium • Surgery, especially abdominal and hip • Major trauma • Malignancy • Myocardial infarct • Thrombocythaemia

22. Risk factors cont… Related to stasis • Cardiac failure • Stroke • Prolonged immobility • Pelvic obstruction • Nephrotic syndrome • Dehydration • Hyperviscosity, polycythaemia • Varicose veins

23. Risk factors cont… Related to unknown factors • Age • Obesity • unfitness • Sepsis • Paroxysmal nocturnal haemoglobinuria • Behcet’s disease

24. Risk factors Related to coagulation abnormality: • Hereditary hemostatic disorders • Factor V Leiden • Prothrombin G20210A variant • Protein C deficiency • Protein S deficiency • Antithrombin III deficiency • Abnormal fibrinogen • Abnormal plasminogen

25. Factor V Leiden • Activated protein C resistance • Characterized by a poor anticoagulant response to activated protein C (APC) • Specific G-to-A substitution at nucleotide 1691 in gene for factor V. Predicts a single amino acid replacement (Arg506Gln) at one of three APC cleavage sites in the factor Va molecule • Factor V Leiden is inactivated approx. ten times slower than normal factor V and persists longer in the circulation, resulting in increased thrombin generation and a mild hypercoagulable state • Most common genetic defect for inherited thrombosis • 15% of Causasians are carriers • Rare in Asians & Africans

27. Factor V Leiden cont… • Diagnosis made either by the APC resistance assay as a coagulation screening test or by DNA analysis of the factor V gene • Heterozygotes 5-10x  risk of clotting • Heterozygotes + OCP 30 to 35 fold  risk • Homozygotes 50-100x  risk • Heterozygotes who are also heterozygous for the G20210A prothrombin mutation have eg 30x  risk • Homozygotes + OCP ???>100 • Hyperhomocysteinemia combined with Factor V Leiden, heterozygous: 20 fold  risk

28. Risk factors Related to coagulation abnormality: • Hereditary hemostatic disorders • Factor V Leiden • Prothrombin G20210A variant • Protein C deficiency • Protein S deficiency • Antithrombin III deficiency • Abnormal fibrinogen • Abnormal plasminogen

29. Protein C deficiency • Protein Cinactivates factor Va and factor VIIIa • Two classifications of protein C deficiency: type I and type II • Type I protein C deficiency: inadequate amount of protein C present • Type II protein C deficiency: normal amount but defective protein C molecules. Numerous defects in the protein C molecule have been described that alter its interactions with thrombomodulin, phospholipids, factor Va and factor VIIIa as well as others.

32. Protein C deficiency • Present in approximately 0.2% of the general population • Heterozygous – 7 fold incr. risk • Homozygous – serious thrombosis at birth (?FFP use, Protein C use) • Both protein C and protein S are Vit. K dependent • Diagnosis: Protein C assays

33. Risk factors Related to coagulation abnormality: • Hereditary hemostatic disorders • Factor V Leiden • Prothrombin G20210A variant • Protein C deficiency • Protein S deficiency • Antithrombin III deficiency • Abnormal fibrinogen • Abnormal plasminogen

34. Protein S deficiency • Function is carried out directly by protein C, and protein S serves as a cofactor. Activated protein C combines with protein S on the surface of a platelet. • Protein S exists in two primary forms. One form is free and the other is bound to an additional protein. Only the free form of protein S is able to interact with protein C in the manner described above • Three classifications of protein S deficiency: type I, type II and type III • Type I: inadequate amount of protein S present in both free and bound forms. Protein S that is present functions normally. • Type II: characterized by defective protein S molecules. The amount of protein S present is normal, but it is unable to interact normally with the other molecules involved in coagulation to perform its function. • Type III: characterized by a low amount of free protein S, but an overall normal amount of total protein S.

36. Protein S deficiency cont… • Incidence in general population is not known. • In the Caucasian population, protein S deficiency has been found in between 1 to 5% of persons who have a venous thrombotic event • Heterozygous: 6 fold increase in risk • Homozygous: severe thrombosis at birth

37. Risk factors Related to coagulation abnormality: • Hereditary hemostatic disorders • Factor V Leiden • Prothrombin G20210A variant • Protein C deficiency • Protein S deficiency • Antithrombin III deficiency • Abnormal fibrinogen • Abnormal plasminogen

38. Antithrombin III deficiency • Antithrombin is a potent inhibitor of the reactions of the coagulation cascade. • Although the name, antithrombin, implies that it works only on thrombin, it actually serves to inhibit virtually all of the coagulation enzymes to at least some extent. • The primary enzymes it inhibits are factor Xa, factor IXa and thrombin (factor IIa). Also has inhibitory actions on factor XIIa, factor XIa and the complex of factor VIIa and tissue factor. • Its ability to limit coagulation through multiple interactions makes it one of the primary natural anticoagulant proteins.

40. Antithrombin deficiency cont… • Antithrombin acts as a relatively inefficient inhibitor on its own. • However, when it is able to bind with heparin, the speed with which the reaction that causes inhibition occurs is greatly accelerated • Two primary types: type I and type II. • Type I: characterized by an inadequate amount of normal antithrombin present. • Type II: normal amount of antithrombin present, but does not function properly and is thus unable to carry out its normal functions. • In many cases, the antithrombin in type I deficiencies has a problem binding to heparin, although there have been multiple other changes to the antithrombin molecule described. • Heterozygous: 5 fold increase risk clots • Homozygous type I: thought to be lethal prior to birth • Homozygous type II: both arterial and venous thrombotic disease. Often have severe complications. • ?Antithrombin concentrates

41. Risk factors Related to coagulation abnormality: • Hereditary hemostatic disorders • Factor V Leiden • Prothrombin G20210A variant • Protein C deficiency • Protein S deficiency • Antithrombin III deficiency • Abnormal fibrinogen • Abnormal plasminogen

42. Prothrombin (Factor II) G20210A variant • Second most common genetic defect for inherited thrombosis • Autosomal dominant disorder • Heterozygotes 3- to 11-fold  risk for thrombosis in both men and women and for all age groups. Homozygosity is rare • Frequently co-inherited in Factor V Leiden carriers • Nucleotide substitution of G to A at position 20210 in the untranslated portion of the prothrombin gene on chromosome 11 causes an elevation of the level of functional prothrombin in plasma which is associated with an increased risk of thrombosis • Diagnosed by a PCR test on WBC’s

43. Risk factor • Hereditary or acquired haemostatic disorders • Raised plasma levels of factor VII, VIII, IX or XI • Raised plasma levels of fibrinogen • Raised plasma levels of homocysteine • Glucosylceramide deficiency • Coagulation factor IX concentrates • Lupus anticoagulant • Oestrogen therapy (oral contraceptive and HRT) • Heparin-induced thrombocytopenia • Pregnancy and puerperium • Surgery, especially abdominal and hip • Major trauma • Malignancy • Myocardial infarct • Thrombocythaemia

44. Antiphospholipids • LA’s have traditionally been classified as anti-phospholipid antibodies, but a more correct view is that they are antibodies directed against plasma proteins, which also bind to phospholipid surfaces, a type of fat molecule that is part of the normal cell membrane. • There are three test in which antiphospholipid antibodies may show up: • 1) anticardiolipin antibodies, • 2) the lupus anticoagulant and • 3) antibodies directed against specific molecules including a molecule known as beta-2-glycoprotein 1. (also anti-prothrombin and the "false-positive" test for syphilis)

45. Antiphospholipids cont… • They are usually IgG, IgM, or mixtures of both, and frequently interfere with standard phospholipid-dependent coagulation tests, often being discovered accidentally such as when a prolonged activated partial thromboplastin time (APTT) is found during a pre-operative evaluation. Importantly, the clotting test abnormalities caused by LA are in vitro phenomena; the antiphospholipid antibodies of the LA react with the phospholipid preparations used to initiate clotting reactions. In vivo clotting factor activities are not diminished and, except in extremely rare cases where there are specific antibodies directed against clotting factor II, there is no danger of a bleeding diathesis.

46. Antiphospholipids cont… • Often, there are no clinical consequences other than the need to explain the reason for the long APTT. A minority of patients with LA have a hypercoagulable state manifested by recurrent thromboses, multiple spontaneous miscarriages, migraine headaches, or stroke. Very rarely, patients may have bleeding. NB important cause of strokes in people under age 40. • Two main classifications of antiphospholipid antibody syndrome: If patient has an underlying autoimmune disorder (e.g. SLE), patient said to have secondary antiphospholipid antibody syndrome. If patient has no known underlying autoimmune disorder, it is termed primary antiphospholipid antibody syndrome

47. Lupus Anticoagulant • Suspect LA in patients with prolonged PTT and no history of bleeding, in women with recurrent miscarriages, and in those with recurrent DVT. • LA not always associated with SLE, although about 50% of SLE patients test positive for LA. However, many patients with LA do not have SLE or any other autoimmune disorder. • The presence of LA does not necessarily indicate the diagnosis of antiphospholipid syndrome. The syndrome requires a positive test of an anti-phospholipid antibody (LA or anticardiolipin antibody), repeatably positive at least 6 weeks later (ie may be transient) plus a clinical history of pregnancy loss or thrombosis—venous more often than arterial.