Hypercoagulable State

1 1 4 Hypercoagulable State Andrew I. Schafer The Concept of Hypercoagulability................ 2423 Primary Hypercoagulable States.................. 2426 Secondary Hypercoagulable States................ 2428 Gene Gene and Gene Environment Interactions in the Hypercoagulable States................... 2431 Clinical Approach to Patients with Suspected Hypercoagulable State........................ 2431 Summary..................................... 2433 Key Points The term hypercoagulable states is used interchangeably with thrombophilia or prethrombotic state to refer to hereditary or acquired conditions that predispose individuals to thrombosis. The coagulation system, also known as the coagulation cascade, is a highly coordinated and tightly linked series of enzymatic reactions, involving the sequential conversion of plasma clotting factors that circulate in their inactive zymogen forms to enzymatically active serine proteases, culminating in the formation of fibrin. The coagulation system is normally kept in check by several physiological anticoagulant (antithrombotic) mechanisms. These include antithrombin III, the protein C/protein S/thrombomodulin system, tissue factor pathway inhibitor, the protein Z/protein Z-dependent protease inhibitor system, and the fibrinolytic system. Primary hypercoagulable states are mostly inherited thrombophilias caused by (1) quantitative deficiencies or qualitative defects of the physiologic anticoagulants, or (2) increased levels or function of the coagulation factors. Secondary hypercoagulable states encompass a variety of diseases and conditions, most of which are acquired, which predispose individuals to thrombosis by complex and multifactorial mechanisms. In gene gene interactions, the inheritance of more than one thrombophilic defect leads to an increased lifelong risk of thrombosis. In gene environment interactions, many thrombotic complications in individuals with inherited (primary) hypercoagulable states are precipitated by acquired, acute thrombogenic insults, pointing to complex interactions between genetic and environmental factors in the pathogenesis of clinical thrombotic events. The hypercoagulable workup is a laboratory evaluation of individuals with suspected hypercoagulable states. It is guided by the history, physical examination, and implications for management decisions. The Concept of Hypercoagulability Definitions The hypercoagulable state or hypercoagulability is a term used interchangeably with thrombophilia and prothrombotic state to refer broadly to hereditary or acquired conditions that predispose individuals to thrombosis. 1,2 The concept of hypercoagulability was proposed 160 years ago by Rudolf Virchow, who postulated three interrelated pathophysiologic causes of thrombosis: (1) changes in the vessel wall, (2) changes in blood flow, and (3) changes in the composition of blood that make blood clot under conditions in which it normally remains fluid. 3 It was the third of Virchow s triad that suggested that systemic alterations in the coagulability of blood is a critical factor in thrombogenesis, a prescient conceptual leap validated in recent years by advances in molecular biology. Normal Coagulation and Physiologic Anticoagulant Mechanisms Blood fluidity throughout the circulation is maintained by the actions of endothelial cells that line the intimal surface of the entire circulatory tree. Under normal conditions these endothelial cells produce or support a complex of physiologic anticoagulant substances that prevent blood clotting and preserve vascular patency. The thromboresistant properties of endothelium are locally disrupted at a site of injury to the vessel wall. This leads to instantaneous activation of the coagulation system and platelets, resulting in the formation of a protective hemostatic plug composed of fibrin and platelets that is localized precisely to the area of vascular damage. 2423

2424 chapter 114 The activation of platelets and the activation of the coagulation system (the so-called coagulation cascade) of plasma proteins occur essentially simultaneously and synergistically to create a platelet-fibrin clot at the site of vascular injury. Arterial thrombi tend to be preferentially composed of platelets, whereas venous thrombi are composed of predominately fibrin, but all thrombi have both platelet and fibrin constituents. In addition, there is growing appreciation of the role of inflammation in many forms of thrombosis, reflected by the presence of leukocytes in thrombi. The Coagulation Cascade Coagulation proteins ( clotting factors ) normally circulate in plasma mostly in their biologically inactive forms, known as zymogens or proenzymes. Focal vascular injury triggers activation of the coagulation system, resulting in the formation of a fibrin clot. Fibrin is rapidly generated through a cascade of coagulation protein reactions. The coagulation cascade is a highly coordinated and tightly linked series of enzymatic reactions, involving the sequential conversion of plasma clotting factors that circulate as zymogens to their enzymatically active serine protease forms. Each protease then catalyzes the downstream zymogen-protease transition by cleavage of specific peptide bonds. Thus, the coagulation cascade is a biochemical amplifier in which a relatively small initiating stimulus, typically caused by local damage or perturbation of endothelium, explosively generates high levels of the end-product fibrin, which is deposited at that site. As shown in simplified form in Figure 114.1, fibrin is formed from fibrinogen by the action of the serine protease thrombin. Thrombin, in turn, is generated from its inactive zymogen proenzyme prothrombin in a reaction catalyzed by factor X a, the activated, enzyme-active form of factor X. Factor V a serves as a cofactor for this reaction. The conversion of factor X to X a occurs primarily by the action of the factor VII a /tissue factor (VII a /TF) complex, which is produced upon exposure of tissue factor (e.g., in a damaged vessel wall) to circulating blood. This is known as the extrinsic or tissue factor pathway of coagulation. Alternatively, factor X can be activated to factor X a by the intrinsic or contact activation pathway of coagulation; here, a sequence of linked zymogen-protease conversions involving factors XII, XI, and VIII generates factor X a -activating factor IX a from factor IX. The fibrin monomers that are formed at the end of the coagulation cascade are stabilized by covalent cross-linking through the action of a transglutaminase, factor XIII a (not shown in Fig. 114.1). Comprehensive reviews provide further details of the complex interactions and kinetics of the coagulation cascade that culminates in the formation of a fibrin clot. 4,5 Physiologic Anticoagulant Mechanisms In normal individuals, the coagulation system is controlled and kept in check by several physiologic anticoagulant mechanisms. Each of these mechanisms depends on the integrity of vascular endothelium for optimal activity. As shown in Figure 114.1, these natural antithrombotic systems essentially blanket the entire coagulation cascade, acting at different strategic steps to limit the amount of fibrin that can VII TFPI PC PS VII a VIII a X TF VII a /TF V a PT PZ-ZPI X a Fibrinogen Th FIBRIN AT III FDP PI PA Plasminogen accumulate. Inherited deficiencies of one or more of the physiologic anticoagulants can produce a lifelong hypercoagulable state. Antithrombin III (or simply antithrombin ) is a serine protease inhibitor (or serpin ), which is the major plasma inhibitor of thrombin and other activated clotting factors. 6,7 The ability of antithrombin to neutralize thrombin and other serine proteases in the coagulation cascade is catalyzed by heparin. This is the major mechanism of action of heparin as a pharmacologic anticoagulant. However, heparin-line substances also exist naturally within the vessel wall. Therefore, the physiologic antithrombin-mediated inactivation of clotting factors probably occurs preferentially on vascular surfaces, where heparins are available to catalyze these reactions, rather than in fluid-phase plasma. Protein C is a vitamin K dependent plasma glycoprotein that becomes a physiologic anticoagulant only after it is converted to activated protein C (APC). 7 10 The major activator of protein C is thrombin itself; thus, thrombin actually stimulates an inhibitor of its own generation. Thrombininduced protein C activation occurs physiologically at specific endothelial cell surface binding sites for thrombin, called thrombomodulin. 7,11 Protein C binds to its own receptors on endothelium (endothelial cell protein C receptor), 9,10 facilitating its concentration in proximity to the thrombinthrombomodulin complex and thereby greatly enhancing IX IX a XI XI a XII XII a FIGURE 114.1. The coagulation cascade and sites of action of physiologic antithrombotic (anticoagulant) mechanisms. Inactive coagulation factors (zymogens) are indicated by roman numerals; their activated forms are indicated by the subscript a. AT III, antithrombin III; FDP, fibrin(ogen) degradation products; PA, plasminogen activator; Pl, plasmin; PC, protein C; PS, protein S; PT, prothrombin; PZ, protein Z; TF, tissue factor; TFPI, tissue factor pathway inhibitor; Th, thrombin; ZPI, protein Z dependent protease inhibitor.

hypercoagulable state 2425 the efficiency of its activation. Activated protein C acts as an anticoagulant by proteolytically digesting factors V a and VIII a in the coagulation cascade. This reaction is accelerated by protein S, another vitamin K dependent plasma protein, which serves as a cofactor for APC. 7,10,12 Protein Z is a more recently discovered vitamin K dependent plasma protein. It circulates in plasma in a complex with the serpin, the protein Z dependent protease inhibitor (ZPI). In the presence of protein Z, the ability of ZPI to inhibit factor X a (in a heparinindependent manner) is dramatically enhanced. Tissue factor pathway inhibitor (TFPI) is a multivalent Kunitz-type serine protease inhibitor that blocks the tissue factor-induced extrinsic pathway of coagulation. 13 It inhibits not only the factor VII a /tissue factor (VII a /TF) complex, as shown in Figure 114.1, but also factor X a. Tissue factor pathway inhibitor circulates bound to lipoproteins. It can also be released by heparin from endothelial cells and platelets, representing a potential additional mechanism of heparin s anticoagulant actions. Finally, any thrombin that is generated despite the coordinated actions of these physiologic anticoagulant mechanisms to quench its formation and activity can stimulate the endogenous fibrinolytic system to dispose of intravascular fibrin. 14,15 Plasmin is the major protease of the fibrinolytic system, mirroring the role of thrombin as the major protease of the coagulation system. Like thrombin, plasmin is a serine protease; it is formed from its inactive plasma zymogen precursor, plasminogen, by the action of the plasminogen activators, tissue-type plasminogen activator (t-pa) and urokinase-type plasminogen activator (u-pa). Both t-pa and u-pa are released from endothelial cells in response to a variety of humoral stimuli (e.g., cytokines, hormones, growth factors) and hemodynamic forces. Plasmin is a two-chain molecule; its light chain (or B chain) contains its enzymeactive site, while its heavy chain (or A chain) contains lysinebinding sites that attach it to fibrin, thereby permitting physiologic fibrinolysis to be localized to fibrin clots. Plasmin cleaves fibrin, as well as fibrinogen, in a process of sequential proteolysis, to yield fibrin(ogen) degradation products (FDPs). When plasmin acts on covalently cross-linked fibrin specifically, D-diners are released; therefore, D-dimers can be measured in plasma as a relatively specific test of fibrin (rather than fibrinogen) degradation. The fibrinolytic system is itself subject to physiologic controls, including plasminogen activator inhibitors (PAIs), which block plasminogen activators, and α 2 -antiplasmin, which neutralizes plasmin. In addition, thrombin itself can downregulate fibrinolysis via thrombinactivatable fibrinolysis inhibitor (TAFI). 16,17 The Prethrombotic State While the physiologic anticoagulant systems described above act in concert to continuously quench thrombin generation and fibrin formation, some baseline systemic state of lowlevel activation of coagulation does occur under normal circumstances. Thus, the hemostatic system is constantly primed to respond instantaneously to injury with a burst of thrombin generation and fibrin deposition at the site of vascular damage. 18 This normal, low-level, baseline state of systemic activation of coagulation can be detected by specific and sensitive assays of the activation of blood coagulation, including plasma levels of prothrombin fragments 1 + 2 (F1 + 2), fibrinopeptides, soluble fibrin monomers, thrombinantithrombin complexes, and D-dimers. Using these tests, it has been demonstrated that individuals with specific thrombophilias have a generalized heightened baseline activation of the coagulation system even when they are asymptomatic. 19,20 This represents biochemical validation of the prethrombotic state, which is presumably a lifelong condition in individuals with inherited thrombophilias. Indeed, even in the absence of a specific identifiable hypercoagulable state, increased levels of activation markers of coagulation and fibrinolysis in twins have provided evidence for a striking genetic basis to the prethrombotic state in otherwise healthy individuals. 21 Heritability of Venous Function Not all hereditary factors that contribute to increased risk of venous thromboembolism involve hypercoagulability per se. For example, genetic factors influence the responsiveness of veins to adrenergic stimulation. 22 Twin studies have likewise demonstrated a strong genetic influence on intrinsic venous function, including compliance and capacitance. 23 This could involve heritable changes in venous structure or the mechanical properties of surrounding tissue, which may contribute to the pathogenesis of thrombosis. Classification of Hypercoagulable States The primary hypercoagulable states are mostly inherited thrombophilias. They can be broadly classified into two categories: (1) quantitative deficiencies or qualitative defects of the physiologic anticoagulants, and (2) increased levels or function of the coagulation factors. 18,24,25 (Table 114.1). In general, the risk of thrombosis is higher in individuals with decreased levels of antithrombotic proteins (antithrombin, protein C and protein S deficiency) than in those with increased levels of prothrombotic proteins (factor V Leiden, prothrombin gene mutation, elevated levels of specific coagulation factors) (Table 114.2). One cohort family study found the overall incidence of venous thromboembolism (per 100 patient-years) to be 1.07 for antithrombin deficiency, 0.54 for protein C deficiency, 0.50 for protein S deficiency, and 0.30 for activated protein C resistance or factor V Leiden. 26 The secondary hypercoagulable states encompass a variety of diseases and conditions, most of which are acquired. These disorders predispose individuals to thrombosis by mostly complex and multifactorial mechanisms (Table 114.3). TABLE 114.1. The major primary hypercoagulable states Decreased antithrombotic proteins Antithrombin deficiency Protein C deficiency Protein S deficiency Increased prothrombotic proteins Factor V Leiden (activated protein C resistance) Prothrombin gene mutation G20210A Increased levels of factors VII, XI, IX, VIII, von Willebrand factor, fibrinogen

2426 chapter 114 TABLE 114.2. Estimated relative risks for first and recurrent episodes of venous thromboembolism in individuals with a thrombophilic defect as compared to individuals without a defect a Estimated relative risk Thrombophilic defect First episode Recurrent episode Normal 1 1 Antithrombin deficiency 8 10 2.5 b Protein C deficiency 7 10 2.5 b Protein S deficiency 8 10 2.5 b Factor V Leiden/APC resistance Heterozygote 3 7 1.3 b Homozygote 80 Prothrombin 20210A mutation 3 1.4 b Hyperhomocystinemia c 2.5 2.6 3.1 MTHFR C677T mutation 1 Elevated factor VIII:C levels d 2 11 6 11 Antiphospholipid antibodies 1.6 3.2 2 9 Lupus anticoagulant 11 a Data reviewed elsewhere. 156,162 b Pooled using the Mantel-Haenzel method. 161 c Mild hyperhomocystinemia, determined fasting or post-methionine loading. d Plasma level (dose)-dependent. Primary Hypercoagulable States Major clinical characteristics of the primary hypercoagulable states are listed in Table 114.4. These disorders are associated with venous rather than arterial thrombosis. However, some considerations qualify this generalization. First, arterial thromboembolism may originate in deep vein thrombi by paradoxical embolism across a patent foramen ovale. This presentation of venous thromboembolism may be underrecognized in view of the high incidence of patent foramen ovale in patients presenting with cryptogenic stroke. 27 31 Second, there is an association between venous thrombosis and atherosclerotic vascular disease. 32 This link has been attributed to a systemic prothrombotic state in atherosclerosis, which promotes venous thrombosis, as well as the sharing of common risk factors between the two diseases. 33 Interestingly, statins reduce not only arterial complications but also the risk of venous thromboembolism. 34 TABLE 114.3. Secondary hypercoagulable states Antiphospholipid syndrome Malignancy Pregnancy Hormonal therapy Trauma Postoperative state Immobility Hyperhomocystinemia Inflammatory bowel disease Nephrotic syndrome Myeloproliferative disorders Hemolytic anemias TABLE 114.4. Clinical characteristics of inherited hypercoagulable states Venous thromboembolism: deep vein thrombosis, pulmonary embolism, superficial thrombophlebitis, intraabdominal or cerebral vein thrombosis First thrombotic event at young age (<40 years) Positive family history of thrombosis Recurrent thrombosis Neonatal purpura fulminans (in homozygous protein C and protein S deficiency) Antithrombin III Deficiency As antithrombin is the major physiologic regulator of thrombin and other activated clotting factors, its deficiency leads to unregulated protease activity and increased fibrin formation. 35 Antithrombin deficiency is inherited in an autosomal dominant fashion, affecting both sexes equally. Over 250 different mutations have been described in inherited antithrombin deficiency. Patients with type I deficiency have proportionately reduced plasma levels of antithrombin antigen and functional activity, resulting from a quantitative deficiency of the normal protein. Major gene deletions, single nucleotide changes, or short insertions or deletions in the antithrombin gene cause impaired synthesis, defective secretion, or instability of antithrombin in type I deficiency states. 36 Patients with type II antithrombin deficiency have normal or near-normal plasma antigen levels accompanied by low functional activity, indicating the presence of a qualitatively defective antithrombin molecule. Type II antithrombin deficiency is further subdivided into subtypes in which abnormalities affect the protease inhibitory activity, or the catalytic heparin binding site (also sometimes referred to as type III deficiency), or both. Type II deficiency is usually caused by point mutations leading to specific amino acid substitutions that produce a dysfunctional antithrombin protein. 35,36 Heterozygosity for antithrombin deficiency, even when causing only a mild deficiency state, is associated with clinical thrombophilia. Homozygous antithrombin deficiency is considered to be incompatible with life, except for individuals with the type II state associated with impaired heparin binding, who have a severe thrombotic phenotype. 37 Patients with antithrombin deficiency are at higher risk for thrombosis than those with any of the currently known primary hypercoagulable states 24 ; about 60% of patients with heterozygous antithrombin deficiency have an episode of venous thromboembolism by age 60 years. 38 The frequency of heterozygous antithrombin deficiency in the general population has been estimated to be between 1 in 250 and 1 in 500. 36 Among unselected patients who present with venous thromboembolism, antithrombin deficiency is found in only about 1%; this rate increases to about 2.5% in selected patients who have recurrent thrombosis or thrombosis before the age of 45 years. Protein C Deficiency Protein C deficiency causes impaired inactivation of factors V a and VIII a, leading to increased thrombin generation and

hypercoagulable state 2427 fibrin formation. It is inherited in an autosomal dominant fashion. More than 160 mutations have been described. Like antithrombin deficiency, protein C deficiency is divided into two general forms: individuals with type I deficiency have a quantitative deficiency of protein C and have proportionate reductions in plasma antigen and functional activity, while those with type II deficiency have a qualitative abnormality of protein C and therefore have disproportionately reduced activity relative to antigen. The more common type I protein C deficiency is associated with a variety of frameshift, nonsense, or missense mutations in the protein C gene, which cause premature termination of synthesis or loss of stability of the protein. Heterozygous protein C deficiency can cause clinical thrombophilia. It is found in about 1 per 200 to 500 in the general population and in 0.5% to 6% of patients with venous thromboembolism. 24,25 Homozygous protein C deficiency associated with absence of protein C production causes the syndrome of neonatal purpura fulminans, which can be fatal unless protein C replacement is administered. 39 Rarely in adults with protein C or protein S deficiency, particularly in individuals with more severe states (e.g., double heterozygosity), or in individuals with APC resistance, warfarin-induced skin necrosis can occur within the first few days of starting therapy. This complication is typically seen in individuals with unsuspected underlying protein C or S deficiency or APC resistance who are started on large loading doses of warfarin, particularly when this is done in the absence of simultaneous heparin anticoagulation. The skin lesions develop predominantly over the extremities, breasts, trunk, or penis, and progress from purpura to necrosis. Skin biopsies characteristically show fibrin thrombi in the cutaneous vessels, similar to the findings in neonatal purpura fulminans. This thrombotic complication of protein C or S deficiency is caused by a temporary prothrombotic imbalance induced by initiation of warfarin, which further reduces the preexisting low level of protein C or S (both of which are vitamin K dependent proteins) to near zero before the long-term anticoagulant actions of warfarin are realized. Because of the rarity of this complication, routine screening for protein C or S deficiency or APC resistance is not recommended for individuals who are to be started on warfarin. The occurrence of skin necrosis is not a contraindication to long-term oral anticoagulation with warfarin in protein C or S deficient or APC-resistant individuals. Related Disorders A fatal thrombotic disorder has been described in a patient with an acquired inhibitor of protein C. 40 A condition mimicking protein C deficiency has been associated with a reduced level of endothelial receptors for protein C, which leads to impaired activation of protein C. 41 Several mutations in the thrombomodulin gene have been reported in patients with venous thromboembolism, but these are rare even in highly selected thrombophilic patients. 42 Protein S Deficiency As a cofactor of protein C, deficiency of protein S causes impaired inactivation of factors V a and VIII a, leading to increased thrombin generation and fibrin formation. Like most of the other inherited hypercoagulable states, protein S deficiency is an autosomal dominant disorder. Unlike protein C, protein S circulates in plasma partly bound to C4b binding protein. Only free protein S, which normally constitutes about 40% of total plasma protein S, can function as an anticoagulant cofactor for activated protein C. Mutations in the protein S gene can lead to three general groups of deficiency states: types I and II are quantitative deficiencies and qualitative defects, respectively, as in antithrombin and protein C deficiency; type III is characterized by normal plasma levels of total protein S but low levels of free protein S. 43 The prevalence of protein S deficiency in the general population is unknown. In one recent study, the prevalence in the Scottish population was found to be about 0.2%, predominantly resulting from the presence of a rare polymorphism. 44 The frequency of protein S deficiency among patients evaluated for venous thromboembolism is comparable to that of protein C deficiency. Issues related to neonatal purpura fulminans and warfarin-induced skin necrosis are similar to those discussed above in protein C deficiency. Factor V Leiden (Activated Protein C Resistance) This primary hypercoagulable state was originally identified in some thrombophilic patients by the inability of APC to prolong the clotting time in vitro, a phenomenon referred to as APC resistance. 45 It was subsequently recognized that in more than 95% of individuals with APC resistance the molecular defect is a specific point mutation in the factor V gene, which replaces guanine with adenine at nucleotide 1691 (G1691A) and leads to the amino acid substitution of Arg506 by Gln (factor V R506Q, the so-called factor V Leiden). This is one of the sites within the factor V molecule normally cleaved by APC when it inactivates it as part of its physiologic anticoagulant action. The factor V Leiden polymorphism renders factor V a incapable of being inactivated by APC. 46 48 In rare individuals with thrombophilia, factor V substitutions other than factor V Leiden can cause APC resistance. 48 50 Heterozygous factor V Leiden is very common in Caucasian populations. It is present in about 5% of healthy individuals of Northern European descent, in 10% of unselected patients presenting with venous thromboembolism, and in 30% to 50% of patients referred for evaluation of thrombophilia. 24,51 Factor V Leiden is much less common in individuals of African or Asian ancestry. 52 Nevertheless, its prevalence in the United States among African Americans and Asian Americans is still about 1% and 0.5%, respectively, making it at least as common as deficiencies of antithrombin, protein C, or protein S. 53,54 While heterozygosity for factor V Leiden increases the lifetime risk of thrombosis by a factor of 5 to 10, homozygosity increases the risk by a factor of 50 to 100. A severe thrombotic phenotype and greater resistance to APC occur in individuals who are either homozygous for this polymorphism or compound heterozygotes, having inherited the factor V polymorphism in trans with a factor V null mutation (pseudohomozygous factor V Leiden). Pseudohomozygosity for factor V Leiden manifests in the laboratory as a combina-

2428 chapter 114 tion of APC resistance and reduced factor V procoagulant activity. 49,55 As with most of the other primary hypercoagulable states, factor V Leiden does not appear to be a risk factor for arterial thrombosis, including myocardial infarction or ischemic stroke. Interestingly, however, its venous thrombotic risk does not include an increased risk of pulmonary embolism. 56 Prothrombin Gene Mutation A polymorphism in the 3 untranslated region of the prothrombin gene (G20210A) is associated with a two- to threefold increased risk of venous thromboembolism. 57,58 Like factor V Leiden, this is a gain-of-function mutation; here, messenger RNA accumulation leads to increased prothrombin synthesis and increased basal levels of functionally normal prothrombin. The prothrombin gene mutation is found predominately in Caucasians, with a frequency of 1% to 6%; is uncommon in African Americans (0.2%); and is rare in other racial groups. 58,59 Increased Levels of Other Coagulation Factors There is increasing recognition that many of the primary hypercoagulable states are not all-or-none mutations in structural genes for the proteins but rather polymorphisms that increase their rate of transcription and translation. 18,60 Polymorphisms can produce varying increases in the levels of coagulation factors that may be associated with corresponding gradations in risk of thrombosis. 18 Elevated levels of coagulation factors VII, 61 XI, 62 IX, 63 and VIII, 61,64 as well as von Willebrand factor 61,65 and fibrinogen, 66 have each been associated with increased risk of venous thromboembolism. Furthermore, increased levels of factor VIII are also implicated in recurrent thromboembolism in both adults and children. 67 69 Since elevated levels of these proteins persist over time, they cannot be attributed to an acute-phase reaction. Although they have yet to be linked to specific abnormal haplotype distributions or mutations, elevated levels of these plasma proteins are likely to be under genetic influence. It is possible that they increase the risk of thrombosis via enhanced thrombin generation. 68 Other Primary Hypercoagulable States With few exceptions, causal relationships between inherited abnormalities of the fibrinolytic system and risk of thrombosis have not been convincingly demonstrated. Conflicting data have been reported for venous thromboembolism linked to hereditary plasminogen deficiency, 70 elevated levels of PAI- 1, and the PAI-1 4G/5G polymorphism. 71 Increased plasma levels of TAFI have been shown to be a mild risk factor for venous thrombosis. 72 Thrombophilic dysfibrinogenemia is a rare abnormality, in which increased tendency to thrombosis has been considered to be due to defective binding of thrombin to a qualitatively abnormal fibrin, leading to increased free thrombin levels or to impaired degradation of the abnormal fibrin clot. 73,74 Using a clot lysis assay that measures overall plasma fibrinolytic potential, a population-based case-control study on risk factors for deep vein thrombosis recently demonstrated a dose-dependent increase in risk with impaired fibrinolysis. 75 Whether this hypofibrinolytic state is determined by genetic or acquired factors, and which proteins are involved, is currently unknown. Despite strong experimental evidence for a key role in TFPI in the control of coagulation, TFPI deficiency associated with thrombotic disorders has not been found. Plasma TFPI levels below the 10th percentile were reported to be a weak risk factor for venous thrombosis in a large populationbased case-control study. 76 A recent report of stop codon mutations within the ZPI suggested an association between ZPI deficiency and thrombophilia. 77 Heparin cofactor II, like antithrombin, is a plasma protein that has heparin cofactor activity; unlike antithrombin, however, it does not inhibit other activated clotting factors. Inherited heparin cofactor II deficiency is not considered a strong risk factor for thrombosis, but may contribute to thrombotic risk when combined with other thrombophilias. Secondary Hypercoagulable States The secondary hypercoagulable states include a diverse variety of conditions that are recognized to predispose individuals to thrombosis (Table 114.3). Most are acquired but some are inherited disorders. In contrast to the primary hypercoagulable states that characteristically involve mutations and polymorphisms of specific proteins that participate in the regulation of coagulation, the pathophysiology of thrombophilia in the secondary hypercoagulable states is usually complex, multifactorial, and incompletely understood. The latter disorders may involve abnormalities of not only coagulation but also the vessel wall itself and rheology, the other factors in Virchow s triad of thrombogenesis. Therefore, the secondary hypercoagulable states often cause arterial as well as venous thrombosis. Finally, age itself is a strong risk factor for thrombosis, with a 1000-fold higher risk in octogenarians than in children. 78 80 Antiphospholipid Syndrome Two forms of antiphospholipid syndrome are recognized: a primary syndrome, with no evidence of an underlying disease, and a secondary syndrome, which occurs mainly in patients with lupus erythematosus. 81,82 Thromboembolic complications occur in up to one third of patients with antiphospholipid syndrome. 36 Antiphospholipid antibodies transiently induced by some drugs and infections are not associated with thrombosis. Both venous and arterial thrombotic events can complicate the course of antiphospholipid syndrome, with high rates or recurrence, which justify the use of long-term prophylactic anticoagulation in these patients. Unusual sites of thrombosis are often observed, including skin necrosis and livedo reticularis, as well as placental thrombosis leading to recurrent spontaneous miscarriages and fetal growth retardation. Occasional patients with catastrophic antiphospholipid syndrome manifest with thrombotic storm, a potentially fatal series of acute vascular occlusive events. 83 Antiphospholipid antibodies are a heterogeneous family of immunoglobulins that includes anticardiolipin antibodies

hypercoagulable state 2429 TABLE 114.5. Laboratory tests for evaluation of hypercoagulable state Antithrombin III deficiency Functional assay of antithrombin III (heparin cofactor assay) Protein C deficiency Functional assay of protein C Protein S deficiency Functional assay of protein S Immunologic assay of total and free protein S Activated protein C (APC) resistance/factor V Leiden Clotting assay of APC resistance, and/or DNA-based test for factor V Leiden (factor V R506Q mutation) Prothrombin gene mutation DNA-based test for Prothrombin G20210A mutation Hyperhomocystinemia Fasting plasma homocysteine Antiphospholipid syndrome Clotting assays for lupus anticoagulant (PT, aptt: inhibitor screen if prolonged; dilute Russell viper venom time; tissue thromboplastin inhibition; others), and Antiphospholipid antibodies [anticardiolipin antibodies by enzyme-linked immunosorbent assay (ELISA); others] (measured by immunoassays) and so-called lupus anticoagulants. The latter represents a particularly strong risk for thrombosis. 84 The autoantibodies in this syndrome are actually not directed against phospholipids themselves, but rather against plasma proteins bound to them, such as β 2 - glycoprotein I and prothrombin. 85 Usually no single laboratory test can make the diagnosis of antiphospholipid syndrome; when it is clinically suspected in thrombophilic patients, a battery of tests is performed (Table 114.5). Malignancy Patients with cancer are at increased risk of thrombosis. 86,87 The malignancies most strongly associated with thrombosis are pancreatic cancer, adenocarcinomas of the gastrointestinal tract or lung, and ovarian cancers. The mechanism of thrombosis usually involves a chronic state of disseminated intravascular coagulation (DIC) that is initiated directly or indirectly by tumor tissue. Other factors that contribute to the thrombotic tendency in cancer include immobility, bulky tumor mass compressing vessels, comorbid conditions such as liver dysfunction due to metastases, sepsis, surgery, long-term indwelling central venous catheters, and the thrombogenic effects of certain antineoplastic agents (e.g., l-asparaginase for acute leukemia, cyclophosphamidemethotrexate-5-fluorouracil for breast cancer, hormonal therapy for breast cancer and prostate cancer, and thalidomide in combination with other agents for multiple myeloma). As discussed below (see Clinical Approach to Patients with Suspected Hypercoagulable State), the development of thrombosis can antedate the detection of malignancy. Occult or overt cancer may be associated with some distinctive thrombotic manifestations such as migratory superficial thrombophlebitis and nonbacterial thrombotic endocarditis. The use of long-term low-molecular-weight heparin instead of oral anticoagulants can substantially reduce the risk of recurrent venous thromboembolism in cancer patients without increasing bleeding complications. 88 Pregnancy Deep vein thrombosis and pulmonary embolism are the most common thrombotic complications of pregnancy. Pregnancy and the first 6 to 8 weeks postpartum are associated with a five- to sixfold increased risk of thrombosis, occurring in about 1 in 1500 pregnancies. 18,89 Cesarean delivery, prior history of thrombosis, inherited or other acquired thrombophilias, obesity, older maternal age, multiparity, and prolonged immobilization all increase the risk of peripartum thrombosis. The pathophysiology of hypercoagulability associated with pregnancy involves a progressive state of DIC throughout the course of normal pregnancy, initiated locally in the uteroplacental circulation. At the same time, the fibrinolytic system is progressively blunted during pregnancy due to the action of placental plasminogen activator inhibitor type 2. These coagulation changes produce a precarious systemic hypercoagulable state that culminates in the peripartum period. Thrombotic risk is compounded by mechanical and rheologic factors in pregnancy, including venous stasis in the legs caused by the gravid uterus, pelvic vein injury during labor, and the trauma of cesaran section. 35 The special difficulties of diagnosing venous thromboembolism in pregnant women have been reviewed in detail elsewhere. 90 When anticoagulation is indicated during pregnancy, unfractionated or low-molecular-weight heparin should be used instead of warfarin because of the teratogenic potential of the latter. Hormonal Therapy The risk of venous thromboembolism is increased about two- to sixfold with the use of oral contraceptives and hormone replacement therapy (HRT). 91 This risk is increased markedly in women with underlying inherited thrombophilias. Second-generation oral contraceptives and HRT increase the thrombotic risk two- to fourfold; unexpectedly, thirdgeneration oral contraceptives, which contain less estrogen and a different progestin, have been found to double the risk of venous thromboembolism compared to second-generation preparations. Oral contraceptive use is also associated with increased risk of peripheral arterial disease 92 and myocardial infarction. 93 Thrombosis is a significant risk in women undergoing assisted reproductive treatment, particularly in association with ovarian stimulation (ovarian hyperstimulation syndrome). 18,94 The mechanisms by which hormonal therapy induces a prothrombotic state are not well understood. However, many of the coagulation abnormalities in this setting are similar to those found in pregnancy. Oral contraceptives and HRT induce a state of acquired APC resistance. 95 Trauma and the Postoperative State Venous thromboembolism is a common complication of major trauma. 96 In one prospective study, 59% of patients seen in a regional trauma center were found to have deep vein thrombosis, although in most cases it was asymptomatic. 97 Pulmonary embolism is the third most common cause of death in major trauma survivors, occurring in 2% to 22% of such patients. 98

2430 chapter 114 The frequency of venous thromboembolism in postoperative patients varies markedly depending on the type of surgery performed. The incidence of postoperative deep vein thrombosis ranges from about 10% after transurethral procedures, 15% to 30% after general or gynecologic surgery, up to 40% after radical prostatectomy, and 45% to 70% following total hip or knee replacement in the absence of thromboprophylaxis. 36,99 In most cases, postoperative deep vein thrombosis is asymptomatic, detected only by noninvasive studies or venography. These studies have served as the basis for current guidelines for perioperative prophylactic anticoagulation regimens. 100 In high-risk surgical patients the development of thrombosis can extend from the time the patient is on the operating table to several weeks after surgery. The mechanism of thrombosis with trauma and surgery involves hypercoagulability, which is presumably triggered by exposure to circulating blood of tissue factor from injured tissue. Mechanical factors, such as venous stasis in the lower extremities and direct trauma to blood vessels, may also contribute to the predisposition to thrombosis. Immobility and Prolonged Air Travel Prolonged immobilization is an independent risk factor for venous thromboembolism. 101,102 In the absence of thromboprophylaxis, the incidence of deep vein thrombosis in patients with acute hemiplegic stroke is about 50% within 2 weeks; 13% to 25% of early stroke deaths are due to pulmonary embolism, most often between the second and fourth weeks. 103,104 Immobility is a contributing, but not the sole, risk factor for thrombosis in hospitalized medical patients. 104 Prolonged air travel poses a small but definite increased risk of venous thromboembolism. 105,106 The risk is related to flight duration, and appears particularly with flights longer than about 6 hours or 2500 miles. Factors that have been identified to compound the risk of flight-associated venous thromboembolism include advanced age, underlying venous disease or previously venous thromboembolism, thrombophilia, use of oral contraceptives, and obesity. 106 109 The major pathophysiologic mechanism is prolonged, uninterrupted sitting in cramped quarters that leads to venous stasis, along with compression of popliteal veins on the edge of the seat. 105 The observation that prolonged sitting can cause venous thromboembolism was made during World War II by Homans, 110 who noted a significant increase in fatal pulmonary embolism in individuals who had been crowded in air raid shelters during the London blitz. In addition to simple preventive measures, such as avoidance of both prolonged sitting and leg crossing, maintenance of hydration, abstinence from alcohol, and the use of below-the-knee elastic stockings, the role of prophylactic anticoagulation in highrisk travelers has not yet been demonstrated. Hyperhomocysteinemia High plasma levels of homocysteine (hyperhomocysteinemia) result from acquired or genetic defects in homocysteine metabolism. Homocysteine is normally remethylated to methionine via pathways involving methionine synthase [using vitamin B 12 (cobalamin) as an essential cofactor] or methyltetrahydrofolate reductase (MTHFR); alternatively, homocysteine is converted to cystathionine in a reaction catalyzed by crystathionine β-synthase (CβS), for which vitamin B 6 (pyridoxine) is an essential cofactor. Marked hyperhomocysteinemia on an inherited basis results from severe deficiencies of methionine synthase, MTHFR, or CβS, and leads to premature atherosclerosis and venous and arterial thrombosis in young patients. Mild-to-moderate hyperhomocysteinemia in adults is usually caused by functional polymorphisms in MTHFR or acquired deficiencies of co - balamin, folic acid, or pyridoxine. Nutritional vitamin deficiency contributes more importantly than MTHFR polymorphisms to the hyperhomocysteinemia of young adults. 111 Prospective studies and meta-analyses have shown that mild-to-moderate hyperhomocysteinemia is weakly associated with atherosclerotic arterial disease as well as with increased risk of venous thromboembolism. 112 114 Coexistence of other thrombophilias enhances the risk of thrombosis in these individuals. 115 The mechanisms of arterial and venous thrombosis in hyperhomocysteinemia are multifactorial. Elevated homocysteine damages the thromboresistant properties of vascular endothelium, accelerates oxidation of low-density lipoprotein cholesterol, and induces vascular smooth muscle cell proliferation. Diagnosis depends on demonstration of elevated fasting homocysteine levels, which correlate with thrombotic risk more than do the findings of MTHFR polymorphisms. 116 Vitamin supplementation with cobalamin, folic acid, and pyridoxine normalizes high plasma homocysteine levels, but its long-term therapeutic efficacy in preventing thrombosis has not yet been demonstrated. Other Secondary Hypercoagulable States Thromboembolism is a common extraintestinal complication of inflammatory bowel disease, typically occurring when the disease is active. 117 It is also associated with Behçet s disease, a multisystem disorder that likewise involves intestinal mucosal inflammation. 118 Activation of the coagulation system in these disorders appears to be associated with a systemic inflammatory response. Renal vein thrombosis is the most characteristic thrombotic complication of the nephrotic syndrome, but the incidence of thrombosis in other sites is approximately 20%. 36 Acquired antithrombin deficiency results from its excessive urinary loss as part of the generalized proteinuria in this syndrome; however, there is poor correlation between the extent of urinary excretion of antithrombin, plasma antithrombin level, and clinical thrombotic complications. Other potential contributing factors include increased platelet reactivity, hyperviscosity, hypofibrinolysis, and hyperlipidemia; however, the mechanisms of thrombosis in nephrotic syndrome remain largely unclear. The myeloproliferative disorders are a group of related bone marrow stem cell disorders that include polycythemia vera, essential thrombocythemia, myelofibrosis and myeloid metaplasia, and chronic myelogenous leukemia. Thrombotic complications are major causes of morbidity and mortality in these disorders. Large-vessel venous and arterial thrombosis are most commonly encountered, but characteristic

hypercoagulable state 2431 thrombotic manifestations include portal and hepatic vein thrombosis, as well as microvascular digital ischemia. 119 The proposed mechanism of thrombosis include uncontrolled erythrocytosis (resulting in increased whole blood viscosity) in polycythemia vera, thrombocytosis, and various qualitative abnormalities of platelet function. Maintaining a normal hematocrit in polycythemia, platelet cytoreduction in highrisk patients with thrombocytosis, 120,121 and antiplatelet therapy with low-dose aspirin 122 have been recommended to prevent thrombotic complications. Thrombosis is a serious complication in patients with various types of hemolytic anemias. Cerebral thrombosis, including silent strokes, pulmonary vaso-occlusion, venous thromboembolism, and arterial thrombosis are important aspects of the clinical spectrum of sickle cell disease 123,124 and the thalassemias. 125,126 In these hemolytic disorders alterations occur in the phospholipid asymmetry of red cells, resulting in exposure of procoagulant anionic phospholipids such as phosphatidylserine, which supports increased thrombin generation. 124,125 Several lines of evidence support the existence of a generalized hypercoagulable state in sickle cell and thalassemia syndromes. 123,125 In paroxysmal nocturnal hemoglobinuria (PNH), chronic intravascular hemolysis is caused by increased sensitivity of red cells to complementmediated lysis, the result of a somatic mutation in the phosphatidylinositol glycan class A (PIG-A) gene, which is required for glycosylphosphatidylinositol glycan anchoring of proteins in the red cell membrane. Since PNH is a clonal disorder of hematopoietic stem cells, platelets are also hypersensitive to complement-mediated injury, and they shed circulating microparticles with procoagulant phospholipids; this may contribute to the thrombotic tendency of hemolysis in PNH patients. 127 Gene Gene and Gene Environment Interactions in the Hypercoagulable States Gene Gene Interactions Familial clustering of severe thrombophilia reflects the importance of gene gene interactions in the inherited hypercoagulable states. 128,129 Indeed, the inheritance of combined thrombophilic defects leads to an increased lifelong risk of thrombosis. Examples include the co-inheritance of factor V Leiden with protein S deficiency, 130 antithrombin deficiency, 131 or prothrombin 20210A. 132,133 Therefore, variability of thrombosis risk associated with individual and combined inherited thrombophilic abnormalities creates gradations of levels of lifelong hypercoagulability. The potential contribution of additional genetic risk factors in most individuals with thrombophilia remains unknown. Interactions with multiple other genetic loci may be an important determinant of thrombosis penetrance and severity. 134,135 Genome-wide scanning and experimental models will be used in the future to identify genetic modifiers of thrombosis that determine phenotype variability in thrombosis susceptibility. 136 The clinical correlate of the concept that multigene interactions determine a lifelong state of hypercoagulability is that venous thromboembolism is now recognized as a chronic disease, 137 with recurrence rates of 17.5% at 2 years and 30.3% at 8 years of follow-up. 138 The importance of systemic hypercoagulability as opposed to simply local, anatomic factors in the pathophysiology of venous thromboembolism is illustrated by the observation that when deep vein thrombosis does recur, it arises in the contralateral leg in almost half the cases. 137 Gene Environment Interactions The genetic basis of hypercoagulability confers a certain level of lifelong thrombosis risk, which is presumably constant; yet thromboembolism is an episodic event. Indeed, it is well established that many thrombotic complications in individuals with primary hypercoagulable states are precipitated by acquired, acute thrombogenic insults. These acquired triggers of thrombosis are often one of the secondary hypercoagulable states. For example, one study found synergistic thrombosis risk of oral contraceptive use in the presence of inherited thrombophilia. The prevalence of the prothrombin gene mutation was significantly higher in patients with venous thrombosis than in healthy control subjects (odds ratio of about 10). Likewise, the use of oral contraceptives was more frequent among women with thrombosis than among controls (odds ratio of 22). However, in women who were taking oral contraceptives and also had an underlying prothrombin gene mutation, the odds ratio for thrombosis rose dramatically to 150. 139 The overall risk of venous thromboembolism during pregnancy and the puerperium is about 1 in 1500; the risk with pregnancy is increased to 0.2% among carriers of factor V Leiden, 0.5% among carriers of the prothrombin gene mutation, and 4.6% among carriers of both genetic defects. 140 The risk of thrombosis in cancer patients is increased in those who are also carriers of the factor V Leiden or prothrombin 20210A mutations. 141 Other studies have demonstrated similar potentiation of thrombosis risk with acquired thrombophilic defects in patients with underlying primary hypercoagulable states. 142,143 Clinical Approach to Patients with Suspected Hypercoagulable State Clinical Assessment History A complete and careful history and physical examination are mandatory in the evaluation of patients with a suspected hypercoagulable state. 144,145 Many aspects of this initial clinical assessment will guide subsequent decisions regard - ing diagnostic testing and management. Details should be obtained regarding the age of onset and location of previous thromboembolic episodes. Objective documentation of thrombosis should be sought, particularly since the purely clinical diagnosis of deep vein thrombosis is notoriously inaccurate. Attempts should be made to identify risk factors and precipitating events for episodes of thrombosis (e.g., immobilization, surgery, underlying malignancy, oral contraceptives, HRT, other thrombogenic drugs, etc.). A family