Page 1962 - Williams Hematology ( PDFDrive )

1936 Part XII: Hemostasis and Thrombosis Chapter 113: Molecular Biology and Biochemistry of the Coagulation Factors 1937

heparin (UFH). Heparin-bridging of AT also contributes to some extent factor Xa. This step is accelerated profoundly via protein S through
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to the AT-mediated inhibition of factors IXa and Xa, but the majority of interactions with the third Kunitz domain of TFPI. 333,334 The following
rate enhancement is provided by the allosteric activation of AT. step is the inhibition of the catalytic activity of tissue factor–factor VIIa
Protease–AT complexes are cleared from the circulation by lipoprotein complexes by formation of the quaternary tissue factor–factor VIIa–fac-
receptor-related protein (LRP)-1–mediated endocytosis in the liver. 324,325 tor Xa–TFPI complex. This complex formation depends on the binding
of Kunitz 1 to the factor VIIa active site. Overall, the effects of TFPI
Gene Structure and Variations as regulator of tissue factor–initiated thrombin generation appear to
The 13.5-kb AT gene (SERPINC1) is localized on chromosome 1q25.1 depend on the fast protein S-dependent TFPI interaction with factor
and consists of seven exons. The cDNA is 1395 bp long, whereas the Xa. 333
mRNA is approximately 1.4 kb. TFPI that is truncated at the C-terminus is effective in inhibiting
Because of its essential role as an inhibitor of coagulation, indi- tissue factor–factor VIIa activity; however, this seems to occur too slow
viduals who are heterozygous for loss-of-function mutations are at to control thrombin generation at least in vitro. In contrast, inhibition
increased risk for thrombosis. The prevalence of this condition in the of tissue factor–factor VIIa activity by GPI-anchored TFPIβ is effective
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general population is approximately one in 5000 individuals, while it and independent of protein S. GPI-anchored TFPIβ also acts as an
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occurs in approximately 5 percent of patients with a history of thrombo- inhibitor of tissue factor–factor VIIa signaling by PARs, a function that
embolic disease. AT deficiency can be categorized into type I and type TFPIα seems to lack.
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II deficiencies. Type I deficiency is characterized by reduced plasma TFPI may prevent prothrombinase formation by factor Xa in the
levels of AT; however, homozygous type I deficiency is not compatible presence of the procofactor factor V, factor V that is partially activated
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with life. Type II deficiency covers all functional AT defects. by factor Xa, or platelet factor V. However, the factor Va–factor Xa
The human gene mutation database (www.hgmd.org) lists 274 prothrombinase complex is not inhibited by TFPI as a result of compe-
mutations. Mutations resulting in type I deficiency consist of large dele- tition by prothrombin.
tions, frameshift mutations, premature stop codons, splice-site muta- The heterogeneity and different activities of the multiple forms of
tions, and missense mutations. Mutations observed in type II deficiency TFPI have frustrated the measurement of TFPI for clinical purposes.
impair heparin binding or affect the overall protein structure. Chapters However, tests that estimate the free full-length form in plasma indi-
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130 and 133 provide a more detailed description of the clinical signifi- cate an association of low TFPIα levels with venous thrombosis. Low
cance of AT deficiency. levels of TFPIα are observed in protein S–deficient patients, which may
be the result of a lack of association of TFPI with protein S in the cir-
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culation and faster clearance of free TFPIα. High TFPIα levels have
TISSUE FACTOR PATHWAY INHIBITOR been observed in patients with increased expression of a splice vari-
TFPI is a Kunitz-type protease inhibitor that inhibits factor Xa and tissue ant of factor V, known as factor V-short. Factor V-short, which lacks
factor–factor VIIa activity and was discovered by Broze and Miletich in the major part of the B domain, interacts with the basic C-terminus of
1987. TFPI circulates in plasma at 2.5 nM in multiple forms of which TFPIα, most likely through the acidic B domain region in factor V. 132,133
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the majority is either truncated at the C-terminus or lipoprotein-asso- Increased factor V-short levels lead to a dramatic increase in plasma
ciated. Only 10 percent of the circulating TFPI is the full-length TFPIα TFPIα, resulting in a bleeding disorder. 133
form of 276 amino acids (Mr ≈40,000; see Table 113–1). The half-life TFPI activity is downregulated by proteolysis at the C-terminus,
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of TFPIα in the circulation is only 2 minutes because it readily associ- upon which the basic C-terminal region or the third Kunitz domain are
ates with the vessel wall endothelium. removed, thereby impairing inhibition of factor Xa and tissue factor–
factor VIIa. Complete inactivation of TFPI is observed after proteoly-
Protein Structure sis by the neutrophil derived proteases elastase and cathepsin G, which
Full-length TFPIα consists of three tandem Kunitz domains and a C-ter- also cleave in between Kunitz 1 and 2. In this way, tissue factor–fac-
minus that contains a basic region (see Fig. 113–15). However, TFPI tor VIIa activity may be protected or reactivated during inflammatory
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is very heterogeneous as a result of proteolysis and alternative splicing. processes. 338
The latter gives rise to TFPIβ that lacks the third Kunitz domain and The in vivo relevance of TFPI was shown by the sensitization of
C-terminus, but instead includes a sequence that facilitates anchorage rabbits to tissue factor–triggered disseminated intravascular coagula-
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to the endothelial cell membrane via GPI linkage. 332 tion after immunodepletion of TFPI. Furthermore, mice lacking the
Endothelial cells and platelets are the main producers of TFPI, with first Kunitz domain of TFPI are not viable. 340
endothelial cells expressing both TFPIα and β, while platelets only pro-
duce TFPIα that is secreted upon platelet activation. Although a signifi- Gene Structure and Variations
cant fraction of TFPIβ is GPI-linked to the endothelial cells, it is also The human TFPI gene (TFPI) is located on chromosome 2q31-q32.1
found in plasma. In vivo, most of the full-length TFPIα appears to be and has nine exons that span 70 kb. TFPI is synthesized in two alter-
bound to endothelial heparan sulphate proteoglycans through its posi- natively spliced forms, α and β. TFPIβ is formed by an alternative
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tively charged C-terminus. This because total plasma TFPI levels rise by splice event after exon 7 such that TFPIβ lacks the third Kunitz domain
approximately threefold upon heparin treatment, which is completely and instead has a unique C-terminus. Exon 2 appears to downregulate
attributable to an increase in TFPIα. In addition, TFPIα also circulates translation of the TFPIβ splice variant by a unique interaction with a
in complex with factor V. sequence in the 3′-end of the TFPIβ mRNA.
Homozygosity or compound heterozygosity for loss of function
Tissue Factor Pathway Inhibitor Function mutations in the gene encoding TFPI has not been described. Several
The physiologic relevance of TFPI stems from its ability to regulate genetic polymorphisms have been identified and their relationship with
tissue factor–dependent coagulation as well as its direct inhibition of venous thrombosis has been investigated. There is one report describing
factor factor Xa. TFPI inhibits the tissue factor–factor VIIa complex in that a T33C polymorphism in intron 7 is highly associated with total
a two-step mechanism. TFPI will bind via its second Kunitz domain to TFPI antigen and protects against venous thrombosis, but this rela-
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the active site of factor Xa, thereby inhibiting the proteolytic capacity of tionship with thrombosis was not confirmed in a subsequent study. 342

Kaushansky_chapter 113_p1915-1948.indd 1937 9/21/15 2:40 PM